Biomedicines doi: 10.3390/biomedicines14061256

Authors: Romina-Christiana Pavlovici Cristina-Crenguţa Albu Claudia Florina Bogdan-Andreescu Viorica Tudor Lucia Bubulac Iuliana-Raluca Gheorghe Arsenie Dan Spînu Emin Cadar Dan Alexandru Slăvescu Mariana Păcurar

Cytokines are key regulators of immune responses and tissue remodeling, playing a central role in physiological homeostasis and pathological inflammation. Dysregulation of cytokine signaling networks has been implicated in a wide range of diseases, where persistent inflammatory activation leads to progressive tissue destruction and impaired repair mechanisms. In the oral environment, cytokines critically influence the balance between tissue resorption and regeneration, particularly in processes involving dentin and alveolar bone remodeling. Pathological root resorption (PRR) represents a clinically significant model of cytokine-driven tissue destruction, characterized by the loss of dental hard tissues mediated by osteoclast-like cells within a dysregulated inflammatory microenvironment. Although mechanical, infectious, and iatrogenic factors are well-established triggers, they alone do not fully explain the variability in clinical outcomes, suggesting an important role for host-related factors. New research highlights the relationship between inflammatory signaling pathways, genetic susceptibility, and molecular biomarkers in shaping the onset and progression of PRR. In particular, the RANK/RANKL/OPG axis, cytokine networks, and gene polymorphisms have been identified as key determinants of osteoclast activation and resorptive activity. At the same time, advances in salivary and gingival crevicular fluid biomarker research provide new opportunities for early detection and real-time monitoring. Despite these advances, current knowledge remains fragmented, with heterogeneous study designs, inconsistent genetic associations, and a lack of standardized diagnostic protocols, all of which limit clinical translation. Therefore, a comprehensive and integrative synthesis of cytokine-mediated mechanisms in PRR is needed. This review aims to provide an updated and critical overview of cytokine and chemokine involvement in PRR, integrating molecular pathways, genetic determinants, and emerging biomarkers within a unified framework while highlighting translational implications for precision dentistry.

Biomedicines doi: 10.3390/biomedicines14061255

Authors: Dror Robinson Muhammad Khatib Eitan Lavon Niv Kafri Waseem Abu Rashed Hamza Murad Mustafa Yassin

Background/Objectives: Chronic low back pain (CLBP) affects approximately 20% of the global population and is a leading cause of years lived with disability. Long-term, real-world evidence for inhaled cannabis in patients refractory to conventional multimodal therapy remains scarce. We assessed the five-year efficacy and safety of inhaled cannabis in CLBP patients who had documented failure of ≥1 year of opioid analgesics, anticonvulsants, antidepressants, NSAIDs, and physiotherapy, with each patient serving as their own historical control. Methods: We analyzed prospectively collected clinical data from 241 consecutive adults with treatment-refractory CLBP (mean age 49.3 ± 14.9 years; 37.8% female; mean pain duration 15.1 years) initiated on inhaled medical cannabis (predominantly smoking, THC 4–22%, CBD 2–22%) in a single-center tertiary orthopedic clinic between 2020 and 2025 (Hasharon Hospital, Rabin Medical Center, Israel; IRB protocols 0807-21-RMC and 0634-25-RMC). Year-0 outcomes during conventional therapy were compared with outcomes at Years 1–5 on cannabis. Primary outcomes were the Numeric Rating Scale (NRS), Oswestry Disability Index (ODI), and Brief Pain Inventory severity/interference (BPI-S/BPI-I). Concomitant-medication trajectories were a secondary outcome. The primary analysis was a mixed model for repeated measures (MMRM) with random intercept and slope, REML estimation, and time as a categorical fixed effect. Multiple imputation (MAR, m = 20, Rubin’s rules) was the primary missing-data approach; complete-case and tipping-point pattern-mixture sensitivity analyses were used. A multivariate Hotelling T2 provided a joint test across the four correlated PROMs. Concomitant-medication discontinuation was modeled with GEE logistic regression and exact McNemar tests. Time to discontinuation was estimated by Kaplan–Meier and Cox regression. The Bonferroni-adjusted significance threshold for the four primary outcomes was α = 0.0125. BioWell gas-discharge-visualization (GDV) parameters were exploratory only. Results: Of 241 patients, 238 (98.8%) provided Year-5 data and 224 (92.9%) remained on cannabis at Year 5; only five patients (2.1%) discontinued for adverse events or inefficacy. All four primary PROMs improved markedly and durably. MMRM-estimated Year-5 minus Year-0 changes were: NRS −5.36 (95% CI −5.65, −5.07), ODI −17.68 (95% CI −19.73, −15.63), BPI-S −6.73 (95% CI −6.99, −6.47), and BPI-I −3.41 (95% CI −3.65, −3.16); all four contrasts had |z| ≥ 16.9 and p < 10−20. MI-pooled estimates were within 0.05 of MMRM (FMI < 0.03 for all outcomes). Hotelling T2 was F(4, 232) = 872.8, p < 10−20. At Year 5, 89.2% achieved ≥30% NRS reduction, 77.2% ≥ 50%, and 93.4% met the NRS minimum clinically important difference (MCID); ODI MCID 65.6%, BPI-S MCID (≥1 pt) 98.3%, BPI-I MCID (≥1 pt) 91.3%. Concomitant opioid use fell from 100% at baseline to 4.6% at Year 5 (within-patient absolute risk reduction 95.4%, McNemar exact p = 1.16 × 10−69), NSAID from 100% to 7.1%, SSRI/SNRI from 80.5% to 5.4%, and gabapentinoid from 38.6% to 2.5%. The ARR-derived NNT for opioid discontinuation was 1.05; this NNT is referenced to each patient’s own documented maximal-conventional-therapy state and is not equivalent to a between-arm randomized-trial NNT. Cannabis dose × time interaction was consistent with no pharmacological tolerance (β = −0.0044 per gram-month per year, p = 0.074). Across 1205 patient-years of cannabis exposure (calculated as 241 patients × 5 follow-up years from Year 1 through Year 5; baseline Year 0 represents pre-cannabis state and is not included in person-time on cannabis), 1338 organ-system AE events were recorded at 1.110/patient-year (Poisson 95% CI 1.05–1.17); 99.8% of graded events were mild (grade 1), with ocular (476 events, 0.40/PY), cognitive (460, 0.38/PY), and gastrointestinal (368, 0.31/PY) reactions predominating. The Year-3 retention dip reflected a documented telemedicine-clinic phenomenon during 2022–2024, with patients returning to in-person follow-up by Year 4–5. BioWell GDV discriminated NRS ≥ 4 only at chance level (BWS AUC 0.574, 95% CI 0.54–0.60; BWV AUC 0.51). Conclusions: In a treatment-refractory CLBP cohort with five-year longitudinal follow-up, inhaled cannabis was associated with large, sustained, and statistically robust improvements in pain, disability, and pain interference, accompanied by near-total displacement of opioids, NSAIDs, antidepressants, and gabapentinoids. These observational associations, although mechanically less susceptible to bias for the binary medication-discontinuation outcomes than for self-reported PROMs, cannot be interpreted causally in the absence of a concurrent randomized control arm and may reflect a combination of pharmacological effect, regression to the mean from a high pre-treatment baseline, expectancy and self-selection effects intrinsic to an actively chosen open-label therapy, and secular trends in pain reporting. The within-patient benefit-risk profile—ARR-derived NNT ≈ 1 for opioid sparing against a predominantly mild adverse-event burden—supports consideration of cannabis as a potentially clinically meaningful, opioid-sparing option in patients who have failed multimodal conventional therapy, pending confirmation in randomized comparative trials.

Biomedicines doi: 10.3390/biomedicines14061254

Authors: Yiwen Wu Jie Qiao Yuchun Liu Xian Yu Haifeng Wang Jianmin Zhang Yi Huang

Background/Objectives: Immune-inflammatory activation is a central feature of aneurysmal subarachnoid hemorrhage (aSAH), yet the epitranscriptomic mechanisms underlying this response remain insufficiently understood. This study aimed to investigate RNA methylation-associated immune dysregulation in aSAH and to identify potential biomarkers and signaling pathways. Methods: Four Gene Expression Omnibus datasets were analyzed to characterize RNA methylation regulator-related immune alterations in aSAH. Single-sample gene set enrichment analysis (ssGSEA), weighted gene co-expression network analysis (WGCNA), and intersection with ImmPort immune genes were used to identify candidate genes. A total of 159 machine learning combinations were evaluated for model construction and external validation. Two-sample Mendelian randomization, single-cell RNA sequencing (scRNA-seq), and CellChat analyses were further performed. Peripheral blood samples from patients with aSAH (n = 12) and matched healthy controls (n = 12) were used for total m6A quantification and quantitative real-time PCR (qRT-PCR) validation, while Western blotting and immunofluorescence were used to validate the protein expression of LIFR, GP130, IGF2BP2, and RBM15B. Results: Eleven RNA methylation regulators were differentially expressed between aSAH and controls in GSE122897. The WGCNA module most strongly associated with RNA methylation regulator-related scores was enriched in immune response and myeloid activation pathways. Intersection analysis identified 25 candidate immune-inflammatory genes associated with RNA methylation regulator-related transcriptional patterns. Among 159 algorithms, an XGBoost-LASSO pipeline selected oncostatin M (OSM) as the key variable, and the resulting RNA methylation regulator-related immune-derived gene signature (RMRIGS) showed good discrimination between aSAH and controls across training and validation cohorts. Mendelian randomization supported a protective association of genetically predicted OSM expression with subarachnoid hemorrhage risk (IVW OR = 0.66, p = 0.014). Single-cell analysis showed that Osm was predominantly enriched in infiltrating Ccr2+ macrophages, whereas Lifr and Il6st were broadly expressed in activated microglial subpopulations, indicating the presence of an Osm − (Lifr + Il6st) communication axis after SAH. Clinically, total m6A levels were increased in peripheral blood samples from patients with aSAH, and OSM, together with several RNA methylation regulators, was upregulated and associated with m6A-related changes. In experimental models, the protein expression levels of LIFR, GP130, IGF2BP2, and RBM15B were all increased after SAH-related stimulation. Conclusions: RNA methylation programs may be involved in immune dysregulation in aSAH. The OSM-centered RMRIGS was associated with disease status and may provide insight into the interaction between peripheral immune activation and post-SAH neuroinflammation. The potential involvement of the OSM–LIFR/GP130 signaling axis and its association with RNA methylation regulator-related alterations warrant further investigation.

Biomedicines doi: 10.3390/biomedicines14061253

Authors: Hong Sun Haiqing Tian Lei Feng Wuhao Ying Yikai Wang Song Gu Weihong Xu Jun Chu

Background: Pancreatic ductal adenocarcinoma (PDAC) has dismal survival, and vascular invasion is strongly associated with dissemination and poor outcomes; however, its molecular basis remains unclear. Methods: Transcriptomic and clinical data from The Cancer Genome Atlas-Pancreatic Adenocarcinoma (TCGA-PAAD) cohort were integrated with Genotype-Tissue Expression (GTEx) normal pancreas data. Vascular invasion-associated candidate genes were identified using subgroup-specific differential expression filtering. A three-gene prognostic signature was constructed using Cox and least absolute shrinkage and selection operator (LASSO) regression and validated in an independent PACA-AU cohort. Nucleoporin 35 (NUP35) was functionally evaluated by shRNA knockdown, phenotypic assays, endothelial assays using conditioned medium, and Western blotting of extracellular signal-regulated kinase (ERK)-vascular endothelial growth factor A (VEGFA) signaling. Results: We identified 172 vascular invasion-associated candidate genes and developed a three-gene model comprising NUP35, GMNN, and KLK11. The model stratified TCGA patients into risk groups with significantly different overall survival (OS; log-rank p = 0.014) and good predictive performance, with areas under the receiver operating characteristic curve (AUC) of 0.659, 0.722, and 0.796 for 1-, 3-, and 5-year OS, respectively. Consistent trends were observed in PACA-AU. Risk group transcriptomes were enriched for proliferative and tumor progression programs. Among the signature genes, NUP35 was prioritized because higher NUP35 expression was associated with poorer survival and positively correlated with VEGFA expression. NUP35 knockdown suppressed malignant phenotypes, reduced endothelial migration and tube formation, and decreased phosphorylated ERK and VEGFA without altering total ERK levels. Conclusions: A vascular invasion-related three-gene signature enables prognostic stratification in PDAC. NUP35 is associated with malignant and pro-angiogenic phenotypes and may regulate angiogenic activity partly through ERK-VEGFA signaling, supporting its potential as a prognostic biomarker and candidate therapeutic vulnerability.

Biomedicines doi: 10.3390/biomedicines14061252

Authors: Vinh Thanh Tran Linh Ha Khanh Duong Sang Doan Nien Vinh Lam Dung Ngoc Kieu Thuc Tri Nguyen

Background/Objectives: Atrial fibrillation (AF) is a prevalent arrhythmia associated with severe clinical complications. This exploratory study aimed to evaluate the prognostic value of clinical characteristics, echocardiographic parameters, and N-terminal pro-B-type natriuretic peptide (NT-proBNP) in predicting new-onset AF among patients previously diagnosed with non-AF arrhythmias. Methods: A prospective cohort study was conducted involving 232 patients who were followed for a median period of 12 months. Data collection included baseline NT-proBNP levels, demographic characteristics, medical history, presenting clinical symptoms, and various paraclinical indices, including echocardiographic measurements. Results: The most frequent presenting symptoms in the study population were syncope (59.9%) and dizziness (55.6%). Statistical analysis indicated that initial NT-proBNP levels were not a significant predictor for the development of new-onset AF (HR = 0.9995; p = 0.717). Conversely, left atrial (LA) size, a history of diabetes mellitus, and a history of stroke were identified as preliminary risk factors requiring confirmation. Specifically, a history of stroke was associated with a nearly 5-fold increase in AF risk (HR = 4.65), while diabetes mellitus increased the risk nearly 4-fold (HR = 3.84). Furthermore, each one-millimeter increase in LA size was associated with a 21% increase in the risk of developing AF (HR = 1.21). Conclusions: The findings suggest that NT-proBNP is not an effective prognostic marker for new-onset AF in patients with non-AF arrhythmias. Instead, left atrial enlargement and clinical comorbidities, such as diabetes and a history of stroke, emerged as suggestive, hypothesis-generating predictors for clinical screening and risk management in this patient population.

Biomedicines doi: 10.3390/biomedicines14061251

Authors: Mariusz Stanisław Sowa Joanna Sowa Maciej Budzanowski

Background/Objectives: Endovascular treatment of cerebral arteriovenous malformations (AVMs) and arteriovenous fistulas (AVFs) is associated with substantial radiation exposure due to procedural complexity and repeated angiographic acquisitions. This study evaluates radiation exposure during AVM and AVF embolization and establishes local diagnostic reference levels (DRLs). Methods: A single-center retrospective dose audit was conducted, encompassing 114 endovascular procedures performed using a low-dose workflow. Radiation exposure was quantified using dose area product (DAP), reference air kerma (Ka,r), fluoroscopy time (FT), and the number of digital subtraction angiography (DSA) frames per procedure. Median values were defined as the median (P50), and local DRLs as the 75th percentile (P75). Comparative analyses were conducted between AVM and AVF procedures, between male and female patients, and within selected AVM subgroups. Results: The analysis comprised 86 AVM procedures and 28 AVF procedures. For AVMs, the local DRLs (P75) were 28.9 Gy·cm2 for DAP, 400 mGy for Ka,r, 310 DSA frames per procedure, and 1619 s for FT. For AVFs, the respective values were 47.3 Gy·cm2, 465 mGy, 478 DSA frames, and 1820 s. No statistically significant differences were identified between female and male patients. However, AVF procedures demonstrated significantly higher radiation exposure than AVM procedures for all parameters except FT. Within the AVM subgroup, no significant differences were observed between single-stage and other AVM procedures or between female and male patients. Conclusions: AVM and AVF embolization procedures are dose-intensive neuroendovascular interventions. Establishing local DRLs for AVM and AVF may enhance radiation monitoring and facilitate procedure-specific dose optimization.

Biomedicines doi: 10.3390/biomedicines14061250

Authors: Hailing Lin Yuli Tang Lingfei Shi Shengjie Zhu Wenqiang Yan Weihong Chen Wancai Que

Background: Koumine is a bioactive alkaloid derived from the traditional medicinal plant Gelsemium elegans. Although it has demonstrated anti-tumor effects in various cancers, its specific role and mechanism in hepatocellular carcinoma (HCC) remain unclear. This study aims to investigate the anti-HCC effects of Koumine and elucidate the underlying molecular mechanisms. Methods: A network pharmacology approach was employed to predict potential targets and pathways of Koumine against HCC. The binding affinities between Koumine and core targets were validated using molecular docking. In vitro, the effects of Koumine on the proliferation, migration, and invasion of HCC cells were assessed, and the expression levels of key proteins were examined. In vivo, the anti-tumor efficacy and toxicity of Koumine were evaluated using a murine xenograft model. Results: Network pharmacology analysis identified 124 potential targets of Koumine against HCC, with 10 core targets (e.g., P38, JAK1, JAK2, GRB2) and key pathways involving MAP2K1, P38, JAK1, and MET being implicated. Molecular docking confirmed strong binding affinities between Koumine and these core targets. In vitro experiments demonstrated that Koumine dose-dependently inhibited the proliferation, migration, and invasion of HCC cells and modulated the expression and phosphorylation of P38. In vivo results showed that Koumine significantly suppressed tumor growth without causing notable toxicity. Conclusions: This study systematically reveals that Koumine exerts its anti-HCC effects by targeting the MAP2K1, P38, JAK1, JAK2, and MET signaling pathways. These findings highlight the potential of Koumine as a novel and safe therapeutic agent for the treatment of hepatocellular carcinoma.

Biomedicines doi: 10.3390/biomedicines14061249

Authors: Fares Jamal Abdullah Hamad Amani Elshaer Pierce L. Claassen Taylor Viggiano Michele Barnhill David M. H. Chascsa Hugo E. Vargas Bashar A. Aqel Blanca C. Lizaola-Mayo

Background: Given aspirin’s biologic plausibility for antifibrotic and antineoplastic effects, we conducted a systematic review and meta-analysis to examine the association between aspirin use and major liver-related outcomes in metabolic dysfunction-associated steatotic liver disease (MASLD). To our knowledge, this is the first systematic review and meta-analysis restricted exclusively to patients with biopsy- or registry-confirmed MASLD. Methods: A comprehensive search of Ovid MEDLINE, Ovid EMBASE, Scopus, and Web of Science was performed in October 2025. Studies enrolling adults with a confirmed MASLD diagnosis were included; those with viral hepatitis or alcohol-related liver disease were excluded. Outcomes assessed included hepatocellular carcinoma (HCC), fibrosis progression, cirrhosis, all-cause and liver-related mortality, gastrointestinal (GI) bleeding, hemorrhagic stroke, and liver disease progression. Hazard ratios (HRs) with 95% CIs were pooled using random-effects models. Heterogeneity was assessed using I2 statistics. Results: Seven studies met the eligibility criteria, with approximately 720,000 individuals included. Pooled analysis showed that aspirin use was associated with a significantly lower HCC risk (HR 0.59; 95% CI 0.43–0.81; I2 = 83%). No statistically significant association was found between aspirin use and cirrhosis incidence (HR 0.55; 95% CI 0.13–2.37; I2 = 83.4%) or GI bleeding (HR 1.11; 95% CI 0.74–1.66; I2 = 98.9%). Among the two studies that explored all-cause mortality, aspirin was associated with a modest but statistically significant reduction in all-cause mortality (HR 0.86; 95% CI 0.78–0.95; I2 = 0%). Conclusions: Aspirin use is associated with a reduced risk of HCC and all-cause mortality in MASLD without significantly increasing GI bleeding or hemorrhagic strokes. These associations may reflect aspirin’s anti-inflammatory properties in liver disease. Further RCTs are needed to verify the causal role of aspirin in MASLD management.

Biomedicines doi: 10.3390/biomedicines14061248

Authors: Elif Ozan Mehmet Cudi Tuncer İlhan Özdemir

Background/Objectives: Combination strategies involving natural compounds are increasingly being evaluated to improve the efficacy and safety of conventional chemotherapeutic agents. Quercetin (Q), a bioactive flavonoid, has been reported to regulate oxidative stress and apoptosis-associated signaling pathways. This study investigated whether Q enhances doxorubicin (DOX)-mediated cytotoxicity in OVCAR3 ovarian cancer cells, with particular emphasis on apoptosis, oxidative stress, and EGFR/FOXP3 signaling, while also assessing relative toxicity in HaCaT non-tumoral keratinocytes. Methods: Cell viability was determined using the MTT assay, and drug interactions were assessed according to the Combination Index (CI) method. Apoptosis was evaluated by Annexin V/PI flow cytometry. Caspase-3 and caspase-9 activities were measured using colorimetric assays. Intracellular reactive oxygen species (ROS) production was analyzed using the DCFH-DA assay. EGFR and FOXP3 gene expression levels were quantified by qRT-PCR, whereas caspase-9 protein expression was assessed immunocytochemically. Results: The DOX+Q combination produced synergistic cytotoxic effects in OVCAR3 cells (CI < 1). Compared with OVCAR3 cells, HaCaT cells displayed higher IC50 values following DOX treatment (7.03 µM vs. 1.42 µM) and Q treatment (183.92 µM vs. 35.94 µM), indicating relatively lower treatment sensitivity and suggesting a potentially favorable selectivity tendency; however, these findings should be regarded as preliminary. Flow cytometric findings demonstrated markedly increased proportions of both early and late apoptotic cells following combination treatment. Caspase-3 and caspase-9 activities were significantly elevated after combined exposure (p < 0.01). ROS production increased substantially in response to DOX+Q treatment, corresponding to an approximately 6.82-fold elevation relative to the control group. qRT-PCR analysis demonstrated reduced EGFR and FOXP3 mRNA expression levels in the combination-treated group. Immunocytochemical evaluation additionally revealed stronger caspase-9 staining intensity in treated OVCAR3 cells. Conclusions: These findings suggest that Q may potentiate DOX-induced cytotoxicity through mechanisms associated with enhanced oxidative stress, activation of apoptotic pathways, and modulation of proliferative signaling. The comparatively lower sensitivity observed in HaCaT cells may indicate a possible selectivity tendency; however, these observations remain preliminary and require further validation through in vivo and translational studies.

Biomedicines doi: 10.3390/biomedicines14061247

Authors: Kuo-Wei Chen Yu-Cheng Huang Yen-Heng Lin Chung-Wei Lee

Background and Purpose: The integration of flow diverter (FD) treatment into hybrid operating rooms (HORs) raises concerns regarding radiation safety, especially when transitioning from biplane systems to single-plane configurations. In this study, we evaluated the impact of distinct procedural-imaging configurations on patient radiation exposure during FD treatment for unruptured cerebral aneurysms. Methods: We retrospectively reviewed 93 patients (HOR: 22; biplane neuroangiography suite [NIS]: 71) treated between 2020 and 2024. Key metrics included fluoroscopy time (FT) and dose area product (DAP), subdivided into 2D fluoroscopy and 3D rotational angiography (3D-RA). Linear regression was used to identify independent predictors of radiation dose. Results: While the HOR significantly reduced fluoroscopy time (19.3 vs. 26.1 min, p = 0.002), it was associated with a higher total DAP compared to the NIS (299.1 vs. 96.3 Gy·cm2, p < 0.001). This increase was primarily driven by a substantially higher radiation dose delivered per 3D-RA acquisition in the HOR environment rather than an increased frequency of 3D imaging. Multivariate analysis confirmed that the surgical imaging configuration was the dominant factor influencing total radiation exposure rather than aneurysm complexity or patient characteristics. Conclusions: Hybrid ORs provide procedural efficiency but involve a significant risk of increased radiation dose due to the reliance on 3D imaging for single-plane navigation. These findings serve as preliminary institutional benchmark data, underscoring the need for adaptive radiation management and configuration-specific protocols to optimize patient safety across diverse surgical imaging suites.

Biomedicines doi: 10.3390/biomedicines14061246

Authors: Kiany Miranda Wagner Muller Estevam Daniel Sesana da Silva Késsia Cristina Carvalho Santos Luisa Martins Simmer Amanda Rangel Madureira Suellem Torezani-Sales Danilo Sales Bocalini Ana Paula Lima-Leopoldo André Soares Leopoldo

Background/Objectives: Chromium picolinate [Cr(pic)3] supplementation and strength training (ST) have been proposed as strategies to improve metabolic health in obesity; however, their combined effects on cardiac cellular function remain unclear. This study evaluated the impact of Cr(pic)3 supplementation associated with ST on body composition, metabolic parameters, cardiac morphology, and cardiomyocyte contractile function in diet-induced obese rats. Methods: Male Wistar rats were fed a high-fat diet and allocated into four groups for 8 weeks: obese sedentary (Ob), obese + ST (ObST), obese + Cr(pic)3 (ObCr(pic)3), and obese + ST + Cr(pic)3 (ObSTCr(pic)3). Chromium picolinate (80 μg/kg/day) was administered by gavage, and ST was performed using a ladder-climbing protocol three times per week. Nutritional, metabolic, cardiac morphological, and isolated cardiomyocyte contractile parameters were assessed. A significance level of 5% was set for all tests. Results: Neither ST nor Cr(pic)3, alone or combined, modified adiposity index, glucose tolerance, insulin resistance, lipid profile (except HDL), or cardiac morphology. ST improved maximal load capacity in trained groups, confirming protocol efficacy. HDL levels were higher in the combined intervention group compared with obese sedentary rats. Cardiomyocyte fractional shortening and maximal contraction and relaxation velocities were unchanged among groups. However, the association of ST and Cr(pic)3 resulted in prolonged time to 50% relaxation, indicating delayed relaxation kinetics without alterations in contractile performance. Conclusions: These findings suggest that Cr(pic)3 supplementation does not enhance metabolic adaptations to ST and may adversely affect cardiomyocyte relaxation dynamics in obesity.

Biomedicines doi: 10.3390/biomedicines14061245

Authors: Maşide Ari Emrah Ari Eray Çinar Hakan Ertürk Deniz Çelik Murat Yildiz Tarkan Özdemir Mehmet Kayadelen Derya Tüten Özdemir Tunahan Dolmuş Hasan İbiş Esma Dolmuş Ömer Faruk Tüten

Background: Pre-Chronic Obstructive Pulmonary Disease (pre-COPD) and Preserved Ratio Impaired Spirometry (PRISm) phenotypes represent important components of the early obstructive lung disease spectrum, characterized by respiratory symptoms and structural lung abnormalities prior to the development of overt airflow limitation. Emphysema is considered one of the major structural phenotypes underlying airway disease and the COPD spectrum. Although cigarette smoking is the best recognized risk factor for these conditions, non-tobacco exposures may also contribute to early structural lung changes. In this study, we evaluated the radiological features, pulmonary function parameters, and dyspnea severity of CT-detected emphysema in symptomatic patients classified as having pre-COPD or PRISm, with particular attention paid to the potential influence of smoking status on disease characteristics. Methods: In this retrospective, single-center study, symptomatic patients aged 20–50 years classified as having pre-COPD or PRISm and in whom emphysema was detected on high-resolution computed tomography (HRCT) were evaluated. Only symptomatic patients who underwent HRCT for clinical indications and in whom emphysema was identified were included. Demographic characteristics, emphysema type and quantitative emphysema severity, pulmonary function parameters, and Modified Medical Research Council (mMRC) dyspnea scores were analyzed. The PRISm and pre-COPD groups were compared in terms of clinical and symptomatic characteristics. In addition, smoking-related clinical and radiological characteristics were also evaluated. Results: A total of 232 patients were included in the study. The median age was 43 years (38–48), and 84.1% of the participants were male. Among the study population, 68.5% were classified in the pre-COPD group and 31.5% in the PRISm group. The most frequently identified emphysema patterns were paraseptal (44.4%) and centrilobular (40.5%). The median total lung emphysema area was 18% (13–22). A weak negative correlation was observed between the degree of emphysema and FEV1 (r = −0.185; p = 0.005), whereas a weak positive correlation was found between emphysema extent and the mMRC dyspnea score (r = 0.214; p = 0.001). Dyspnea severity was significantly higher in the PRISm group compared with the pre-COPD group (p < 0.001). In the smoking-status subgroup analysis, ever-smokers demonstrated significantly greater dyspnea severity and lower FEV1 values, whereas never-smokers had a significantly higher proportion of emphysema extent > 18% (all p < 0.05). Conclusions: Radiologically detected emphysema in symptomatic patients without airflow limitation was associated with statistically significant but weak alterations in pulmonary function and dyspnea burden. Dyspnea severity was significantly higher in the PRISm phenotype. In a smoking-status subgroup analysis, ever-smokers had significantly greater dyspnea severity, whereas never-smokers showed a significantly higher proportion of extensive emphysema (>18%), despite similar functional impairment across groups. These findings underscore the importance of non-tobacco exposures in the development of emphysema within pre-obstructive spirometric phenotypes. Multicenter prospective studies incorporating healthy controls and systematic exposure documentation are needed to confirm these observations.

Biomedicines doi: 10.3390/biomedicines14061244

Authors: Larissa Vieira Toledo Maíra Gabrielle de Abreu Ribeiro Thays Cristina dos Santos Maria Luiza Nonato Salvador Natália Oliveira Bertolini Jaqueline do Carmo Lima Carvalho Débora Ribeiro Orlando Rafael Neodini Remedio Alan Rodrigues Teixeira Machado Leonardo Barros Dobbss Stela Márcia Pereira Dourado Luciano José Pereira Eric Francelino Andrade

Background: Estrogen deficiency negatively affects alveolar bone by disrupting key regulators of bone remodeling. Humic acids (HAs) are natural compounds with recognized antioxidant and anti-inflammatory properties that may attenuate bone resorption. This study investigated the effects of HAs on alveolar bone in an experimental model of estrogen depletion. Methods: Female C57BL/6 mice were randomly assigned to four groups: Sham, Sham + HA, ovariectomized (OVX), and OVX + HA. Estrogen deficiency was induced by bilateral ovariectomy. HAs derived from vermicomposted biomass were administered daily by oral gavage (80 mg/kg) for 28 days. At the end of the experimental period, mandibles were collected for structural, mineral, and histological analyses. Bone elemental composition was assessed using scanning electron microscopy coupled with energy-dispersive spectroscopy (SEM/EDS). Alveolar bone loss was evaluated by histomorphometry, while RANKL and osteoprotegerin (OPG) expression were assessed by immunohistochemistry. Osteoclasts were quantified by tartrate-resistant acid phosphatase (TRAP) staining. Data were analyzed using two-way ANOVA followed by Bonferroni’s post hoc test. Results: Ovariectomy resulted in reduced calcium and phosphorus content, increased alveolar bone loss, elevated RANKL immunolabeling, increased osteoclast numbers, and a higher RANKL/OPG ratio (p < 0.05). HA treatment increased calcium and phosphorus content and attenuated alveolar bone loss in OVX animals (p < 0.05). Additionally, HA treatment partially increased OPG expression and reduced the RANKL/OPG ratio (p < 0.05), without significantly affecting RANKL immunolabeling or osteoclast numbers. Conclusions: HA therapy attenuated alveolar bone resorption in a model of estrogen depletion, possibly associated with modulation of the RANKL/OPG balance.

Biomedicines doi: 10.3390/biomedicines14061243

Authors: Kevin Triangto Bambang B. Siswanto Tresia F. U. Tambunan Teuku Heriansyah Alida R. Harahap Aria Kekalih Hajime Katsukawa Anwar Santoso Basuni Radi

Background: Skeletal myopathy is a common complication of heart failure (HF), contributing to exercise intolerance and impaired physical function. This study explores the relationship between practical skeletal muscle measurements and key biomarkers in HF patients undergoing cardiac rehabilitation. Methods: Sixty-nine stable chronic HF patients participated in a 3-month phase II cardiac rehabilitation program. Physical examinations, including the 6-Minute Walk Test (6MWT), chest expansion, inspiratory diaphragm thickness, and handgrip strength, were conducted. Blood samples were analyzed for myostatin and miRNA-133a. Data were analyzed using paired t-tests, Wilcoxon tests, Chi-square/Fisher’s exact tests, and correlation analyses. Results: Significant improvements were observed in 6MWT distance, chest expansion, and inspiratory diaphragm thickness following rehabilitation (p < 0.001). Handgrip strength also significantly improved post-rehabilitation. Myostatin and miRNA-133a levels did not change significantly post-rehabilitation. However, exploratory cross-sectional analysis revealed trends suggesting that lower myostatin levels correlated with better endurance (p = 0.036), while higher myostatin levels were also observed in patients with better 6MWT performance (p = 0.014). Higher miRNA-133a levels were potentially associated with better overall fitness, including endurance and respiratory function (p < 0.05). Conclusions: Readily performed physical assessments can serve as clinical indicators of the systemic impact of HF on skeletal muscle. The study highlights the importance of evaluating extracardiac function in HF patients, demonstrating potential exploratory associations between physical function and key biomarkers.

Biomedicines doi: 10.3390/biomedicines14061242

Authors: Sara Castiglioni Antonella Tosoni Manuela Nebuloni Jeanette A. Maier

Background/Objectives: Magnesium (Mg) is essential for neuronal maturation, yet its role in human cortical development remains poorly defined. Here, we investigated the effects of physiological (1 mM) and elevated (5 mM) concentrations of MgSO4 and magnesium pidolate (MgPid) on human brain organoids co-cultured with an in vitro blood–brain barrier (BBB) model. Methods: Human brain organoids derived from induced pluripotent stem cells were co-cultured with an in vitro BBB system and treated for 4 days with either MgSO4 or MgPid at physiological and elevated concentrations. Cortical organization was assessed by transmission electron microscopy and immunofluorescence analysis. Western blotting for neurotransmitter receptors and Mg transporters, quantification of intraorganoid Mg2+ levels, ELISA-based measurement of GABA and dopamine, and analysis of glutamate were performed. Results: High Mg exposure enhanced cortical stratification and neuronal organization, as shown by the localization of CTIP2 in the outermost layer and TBR2 in the inner layer, together with ultrastructural features consistent with advanced differentiation. Elevated Mg increased intraorganoid Mg2+ levels without altering Mg transporter abundance and selectively modulated neurotransmitter receptor expression: NMDA-R levels were reduced by MgPid, whereas GABAA-R and GABAB-R were upregulated, particularly in response to MgPid. Levels of glutamate, GABA, and dopamine remained unchanged. Conclusions: These findings identify Mg, especially in the form of MgPid, as a modulator of cortical architecture and inhibitory–excitatory receptor balance in human organoids, supporting its potential relevance for neurodevelopmental regulation and Mg-based therapeutic strategies. These results also support organoids as human-relevant, animal-free tools for neuroscience and neuropharmacological research.

Biomedicines doi: 10.3390/biomedicines14061241

Authors: SeungHwan Lee Lakkyong Hwang Sang Hoon Kim Sang Hoon Lee Jin Hee Han Jung Won Jeon Hyeong Chan Shin Il-Gyu Ko

Background/Objectives: Ulcerative colitis (UC) is a type of inflammatory bowel disease characterized by abdominal pain, diarrhea, and bleeding. Polydeoxyribonucleotide (PDRN), an adenosine A2A receptor (A2AR) agonist, exhibits anti-inflammatory properties. In the present study, we evaluated the therapeutic effects of PDRN in a dextran sodium sulfate (DSS)-induced murine model of UC. Methods: UC was induced by administering 2% DSS in drinking water for 7 days. One day after DSS administration, mice received intraperitoneal injections of PDRN (8 mg/kg) for 7 days. To investigate the involvement of A2AR, the selective antagonist 3,7-dimethyl-1-propargylxanthine (DMPX, 8 mg/kg) was co-administered with PDRN. Results: DSS administration induced colonic tissue damage and increased disease activity index (DAI) and histological scores. DSS also elevated pro-inflammatory cytokines while reducing anti-inflammatory cytokine levels. PDRN treatment reduced histological damage, restored body weight, colon weight, and colon length, and decreased DAI scores. Furthermore, PDRN treatment inhibited nuclear factor kappa B (NF-κB) activation through suppression of NF-κB inhibitor-α phosphorylation and was associated with activation of the cAMP/PKA/CREB signaling pathway. PDRN treatment attenuated inflammation and was associated with increased expression of vascular endothelial growth factor (VEGF) in colonic tissues. Given the context-dependent role of VEGF in inflammatory bowel disease, this increase is interpreted as contributing to mucosal repair rather than exacerbating inflammation. Co-administration of DMPX abolished these effects, suggesting the involvement of A2AR-dependent signaling pathways. Conclusions: PDRN attenuated colonic inflammation and improved disease outcomes in DSS-induced UC, potentially through modulation of the PKA/CREB/NF-κB signaling pathway and VEGF-mediated tissue repair mechanisms.

Biomedicines doi: 10.3390/biomedicines14061240

Authors: Mehmet Özgür Özates Kadir Çetinkaya Yasar Ünsal Hümeyra Kullukçu Oktay Gürcan Atilla Kazancı Evrim Önder Tuba Saadet Deveci Bulut Ahmet Gürhan Gürcay

Background: Cerebral ischemia–reperfusion (I/R) injury is a significant contributor to mortality and long-term disability following ischemic stroke. Despite advances in neuroimaging, there remains a critical need for non-invasive, sensitive circulating biomarkers for early diagnosis and management. Endocan, a soluble proteoglycan secreted by activated endothelial cells, has been implicated in various vascular inflammatory conditions, but its specific role as a biomarker for cerebral I/R injury in rodent models requires further elucidation. Methods: Sixteen adult male Sprague Dawley rats were randomly assigned to either a sham (n = 8) or an ischemia–reperfusion (I/R) group (n = 8). Cerebral I/R injury was induced by temporary bilateral common carotid artery occlusion for 10 min, followed by reperfusion. Serum endocan levels were quantified using ELISA at baseline (0 min) and 6, 24, and 48 h post-reperfusion. Histopathological evaluation of hippocampal neuronal degeneration was performed at 48 h using a four-point grading system by blinded neuropathologists. Statistical analyses included independent samples t-tests, one-way repeated measures ANOVA, and Spearman’s rank correlation. Results: Baseline serum endocan levels did not differ between groups (p = 0.814). However, in the I/R group, endocan concentrations were significantly elevated compared to the sham group at 6 h (p < 0.005), 24 h (p < 0.001), and 48 h (p < 0.001). Intra-group analysis of the I/R cohort revealed a significant rapid elevation in endocan levels relative to baseline at 6 h (p = 0.003), followed by a gradual decline at 24 h (p < 0.001) and 48 h (p = 0.028), remaining significantly elevated above baseline at all time points. Histopathological examination showed significantly greater neuronal degeneration in the I/R group (median score = 2.5) compared to the sham group (median score = 0; p < 0.001). A strong positive correlation was observed between serum endocan levels at 48 h and hippocampal neuronal degeneration scores within the I/R group (Spearman’s ρ = 0.857, [95% CI: 0.482–0.968]; p = 0.007). Conclusions: Serum endocan demonstrates high levels following cerebral I/R injury in a rodent model, correlating strongly with the severity of hippocampal neuronal damage. These findings suggest that serum endocan is a sensitive and biologically relevant circulating biomarker for cerebral I/R injury, holding potential for non-invasive monitoring of endothelial dysfunction and secondary injury processes in acute ischemic stroke.

Biomedicines doi: 10.3390/biomedicines14061239

Authors: Corentin Dauleac David Meyronet François Ducray Patrick Mertens François Cotton

Background/Objectives: Intramedullary tumors are uncommon spinal cord lesions that account for a small proportion of central nervous system neoplasms but are associated with a high risk of neurological morbidity. Accurate preoperative characterization is essential because therapeutic strategies, surgical planning, and functional prognosis depend strongly on tumor biology and growth behavior within the confined spinal cord environment. This study aims to characterize the radiological phenotype of intramedullary tumors and to identify imaging patterns that may assist in lesion characterization and diagnostic stratification. Methods: A retrospective analysis of preoperative MRI findings in patients with histopathologically confirmed intramedullary tumors was performed. Preoperative MRI examinations were systematically analyzed to describe imaging features according to tumor histology using conventional sequences (T1-weighted, T2-weighted, and contrast-enhanced imaging). Results: Distinct radiological phenotypes were observed across a wide spectrum of lesions. Glial tumors, including subependymoma, ependymoma, pilocytic astrocytoma, diffuse midline glioma H3K27M, glioblastoma, high-grade astrocytoma with piloid features, ganglioglioma, and diffuse leptomeningeal glioneural tumors, demonstrated variable combinations of cord expansion, margin definition, enhancement patterns, and tract involvement, reflecting differences between expansile and infiltrative growth. Secondary tumors such as metastases frequently exhibited aggressive imaging features, including extensive edema and intense or heterogeneous enhancement. Vascular lesions, including hemangioblastoma and cavernoma, showed characteristic vascular signatures, such as nodular enhancement with flow voids or susceptibility-related signal changes. Developmental lesions, such as epidermoid cysts, neurenteric cysts, and lipoma, displayed distinctive signal characteristics, especially on diffusion and T1, that aided differentiation from neoplastic processes. Conclusions: In conclusion, the structured radiological interpretation functions proposed herein are not only useful for diagnostic purposes, but could also be useful for risk stratification and therapeutic guidance.

Biomedicines doi: 10.3390/biomedicines14061237

Authors: Dilek Altun Varmış Cumali Yüksekkaya Hülya Binokay Serkan Güneş Elif Gözde Yüce Antepüzümü Yunus Kıllı Nazmiye İnce Hamide Kübra Özlük

Background/Objectives: Peripheral biomarkers for autism spectrum disorder (ASD) have shown mixed results in previous studies. In this study, complete blood count-derived immune-inflammatory markers, iron and micronutrient levels, and thyroid function were compared between drug-naïve preschoolers newly diagnosed with ASD and healthy controls. Additionally, the relationships between these markers, symptom severity, and developmental skills were examined. Methods: This retrospective case–control study included 62 children with ASD (aged 24–72 months) and 61 age-matched healthy controls. Symptom severity, behavioral traits, and developmental status were assessed using the Childhood Autism Rating Scale (CARS), Autism Behavior Checklist (ABC), and Denver II Developmental Screening Test (DDST), respectively. Composite inflammatory indices were calculated from hemogram data. Statistical analyses incorporated Holm–Bonferroni corrections for multiple comparisons and sex-stratified exploratory analyses of conditional associations using 95% bootstrap confidence intervals based on 5000 resamples. Results: Children with ASD demonstrated significantly lower mean corpuscular volume (MCV; d = 0.66, adj. p = 0.019), lower mean platelet volume (MPV; d = 0.58, adj. p = 0.034), and higher absolute lymphocyte counts (LYMPH; d = 1.10, adj. p = 0.019). Initial group differences in ferritin, serum iron, and transferrin saturation did not survive adjustment (adj. p > 0.05). Composite inflammatory indices were not significantly associated with clinical or developmental scores. Higher CARS and ABC scores correlated with lower personal–social and language scores on the DDST (p < 0.01). Furthermore, exploratory sex-stratified, conditional association analyses suggested preliminary basophil- and lymphocyte-related patterns in girls; however, these findings are strictly hypothesis-generating due to the small female sample size (n = 12). Conclusions: Newly diagnosed, drug-naïve preschoolers with ASD showed a distinct baseline blood profile, including lower MCV and MPV and higher lymphocyte counts. Clinical challenges were most evident in personal–social and language domains. While the primary diagnostic value of routine hemograms in this context appears limited, the exploratory sex-stratified basophil- and lymphocyte-related patterns require validation in adequately powered future cohorts.

Biomedicines doi: 10.3390/biomedicines14061238

Authors: Muhamad Mustaqim M Zapawi Yee Xing You Mazlyfarina Mohamad Ponnusamy Subramaniam Mohd Razif Shahril Faizah Mohd Zaki Suzana Shahar

Background/Objectives: Cognitive flexibility and working memory are regulated by the dorsolateral prefrontal cortex (DLPFC), which is closely linked to the progression of cognitive decline. The Mediterranean-DASH Intervention for the Neurodegenerative Delay (MIND) diet shows potential to lower cognitive decline risk in older adults. This study aimed to examine the association between Malaysian-MIND diet (MY-MINDD©) scores with brain activation among Malaysian older adults. Methods: A cross-sectional study was conducted among forty older adults aged 60–75 years. Subjects were stratified into quartiles of MY-MINDD© scores with ten subjects per quartile. Dietary intake was evaluated utilising a validated 124-item semiquantitative Food Frequency Questionnaire (FFQ). Brain activation was measured using task-based fMRI (N-back and Stroop Colour Word Test). DLPFC activation was analysed in Brodmann’s areas 9, 46, and the anterior cingulate cortex (ACC). ANCOVA and multiple linear regression were used to evaluate brain activation differences across MY-MINDD© quartiles, accommodating for gender, age, education, and body mass index (BMI). Results: Subjects in the highest MY-MINDD© quartile had significantly greater DLPFC activation during 0-back, 1-back, and SCWT incongruent tasks (p < 0.05). Higher MY-MINDD© adherence is linked to better task performance (p < 0.001). Multivariate General Linear Model (GLM) revealed a significant overall effect on brain activation (Pillai’s Trace = 0.544, F(8,27) = 4.11, p = 0.003). Multiple linear regression demonstrated significant positive associations between MY-MINDD© scores and DLPFC activation (p < 0.0125). Conclusions: Higher adherence to the MY-MINDD© diet was associated with greater brain activation, suggesting its relevance as a proxy for identifying risk of cognitive decline.

Biomedicines doi: 10.3390/biomedicines14061236

Authors: Baihui Hu Xiaocong Wang Xiuli Chen Tong Li Chuqiao Li Yapu Zhang Yingde Wang Jingwei Mao

Background: The Five-flavor Sophora Flavescens Enteric-coated Capsule (FSEC) is widely used to treat ulcerative colitis (UC), yet its underlying molecular mechanisms remain incompletely understood. This study aimed to systematically elucidate the therapeutic mechanisms and clinical value of FSEC by integrating network pharmacology, experimental validation, and clinical data. Methods: A multi-tiered methodological framework was implemented. First, network pharmacology analysis identified potential molecular targets and signaling pathways associated with FSEC in UC. These predictions, including the core targets and the AGE-RAGE/NF-κB signaling pathway, were subsequently validated in a dextran sulfate sodium-induced mouse colitis model in vivo and a lipopolysaccharide-stimulated RAW264.7 macrophage inflammation model in vitro. Finally, a single-center retrospective cohort study of 90 patients evaluated the efficacy and safety of FSEC as monotherapy and in combination with Mesalazine. Results: Network pharmacology identified 12 core targets and predicted the AGE-RAGE/NF-κB pathway as a key mechanism. Experimental validation demonstrated that FSEC inhibits this pathway. Both in vivo and in vitro studies consistently showed that FSEC downregulates RAGE expression and NF-κB p65 phosphorylation, reducing the production of key inflammatory mediators and thereby alleviating intestinal inflammation. Clinically, FSEC monotherapy achieved efficacy comparable to that of Mesalazine, and combination therapy was associated with higher clinical remission and mucosal healing rates, with a favorable safety profile. Conclusion: This comprehensive multi-omics investigation provides integrated evidence that FSEC exerts anti-inflammatory effects, associated with inhibition of the AGE-RAGE/NF-κB pathway. The observed synergy between FSEC and Mesalazine suggests the potential of an integrated therapeutic approach for UC.

Biomedicines doi: 10.3390/biomedicines14061233

Authors: Shanshan Zhang Jingying Jiang Shan Zheng

Organoids are three-dimensional culture systems that self-organize and partially recapitulate the architecture, cellular composition, and functional properties of native tissues. In pediatric congenital hepatobiliary diseases, persistent cholestasis, bile duct maldevelopment, epithelial injury, and progressive fibrosis often lead to cirrhosis, liver failure, or the necessity for liver transplantation. Compared with conventional two-dimensional cell culture and animal models, hepatobiliary organoids provide patient-derived, human-relevant platforms for modeling disease mechanisms, evaluating therapeutic responses, and exploring regenerative strategies. Unlike previous reviews that mainly discuss general organoid culture systems or broad liver disease modeling, this review is organized around clinically oriented translational endpoints, including mechanistic target discovery, prognostic stratification, therapeutic validation, and regenerative reconstruction. We further discuss current barriers to clinical translation, including reproducibility, scalability, vascularization, immune integration, manufacturing standardization, and patient-specific genetic, environmental, and dietary modifiers. By integrating disease-specific mechanisms with translational applications, this review provides a framework for understanding how organoid-based platforms may contribute to future diagnosis, risk assessment, therapeutic decision-making, and regenerative medicine in pediatric congenital hepatobiliary disorders.

Biomedicines doi: 10.3390/biomedicines14061235

Authors: Boris Dinkov

The global burden of type 2 diabetes mellitus (T2DM) and metabolic dysfunction-associated steatotic liver disease (MASLD) continues to rise at an alarming pace, with substantial pathophysiological overlap driven by insulin resistance, visceral obesity, and chronic low-grade inflammation. MASLD may progress to metabolic dysfunction-associated steatohepatitis (MASH), with increased risk of cirrhosis and hepatocellular carcinoma. Glucagon-like peptide 1 (GLP-1)-based therapies have transformed the management of T2DM and obesity. They exert pleiotropic effects whose basis remains incompletely understood. Concurrently, Akkermansia muciniphila has emerged as a keystone gut microbiota species with demonstrated hepatoprotective potential in preclinical models of MASLD/MASH. This narrative review positions A. muciniphila simultaneously as a target of GLP-1-mediated microbiome remodeling and as an independent modulator of hepatoprotection in MASLD/MASH. A structured search of PubMed, Scopus, and Web of Science (last searched: 12 April 2026) was conducted using terms related to Akkermansia muciniphila, GLP-1 receptor agonists, MASLD/MASH and T2DM. A total of 174 records were identified. Of these, 148 were excluded due to duplication or non-relevant study design. 26 studies (23 preclinical, 3 clinical) were included in the synthesis, directly addressing A. muciniphila. Preclinical evidence demonstrates that liraglutide, semaglutide, exenatide, and tirzepatide increase A. muciniphila abundance, while A. muciniphila in turn enhances endogenous GLP-1 secretion via the P9/ICAM-2 axis, forming a hypothetical positive feedback loop. A working mechanistic model integrating these bidirectional interactions is proposed, alongside a discussion of current limitations and future research priorities, including microbiome-guided clinical trials in MASLD/MASH populations.

Biomedicines doi: 10.3390/biomedicines14061230

Authors: Celina Logioco Anahí Lupinacci Ignacio Lischinsky Mariel A. Ytques Gabriel Bercovich María A. Moussalli Agustina De Gainza Mario O. Roux Nicolás Levaggi Ana Sanseau Karina B. Giannone Arturo Burchakchi Leila Galetto Natanael Serrano Eimi Olivares Sefair Guillermo Roux Rodrigo M. Torres

Objective: The aim of this study was to evaluate the safety and efficacy of the Paul Glaucoma Implant (PGI) in a multicenter Argentine cohort with 12 months of follow-up. Methods: This ambispective multicenter study included patients who underwent PGI implantation between November 2022 and July 2024 by glaucoma specialists across Argentina, with a minimum follow-up of 12 months. Primary outcomes were intraocular pressure (IOP) reduction and success rates, defined as complete success (≥20% IOP reduction with IOP ≥6 and ≤21 mmHg without medications), qualified success (same criteria with ≥1 medication), and failure based on predefined efficacy and safety criteria, including additional glaucoma surgery, device removal, or clinically significant hypotony. Secondary outcomes included changes in medication use, best-corrected visual acuity (BCVA), and postoperative complications. Results: Sixty-six eyes were included in the overall analysis. Mean IOP decreased from 31.2 ± 9.1 mmHg preoperatively to 12.8 ± 4.7 mmHg at 12 months (p < 0.01). Medications were reduced from 3.5 ± 0.8 to 1.3 ± 1.2 (p < 0.01). Among the 65 eyes with evaluable 12-month follow-up, 50 eyes (76.9%) achieved complete success, 14 (21.5%) qualified success, and 1 (1.5%) failure. Complications were ocular hypertension (25%), tube or plate exposure (10%), choroidal detachment (7%), and hypotony (5%). BCVA remained stable in 28 eyes (43.8%), improved in 15 (23.4%), and worsened in 21 (32.8%). Conclusions: The PGI achieved significant and sustained IOP reduction with marked medication decrease at 12 months. Most complications were mild or moderate. These findings support the PGI as an effective and safe non-valved implant for refractory glaucoma in Latin American populations.

Biomedicines doi: 10.3390/biomedicines14061234

Authors: Zhuwei Huang Yuan Wang Yixian Luo Zixu Zhang Jiaxuan Huang Shixian Zang Pei Ye Qiao Peng Ting Liu Wenmei Wang Xiang Wang Ning Duan

Background/Objectives: This study aimed to explore the potential value of narrow-band-imaging (NBI)-derived mean optical intensity (MOI) in monitoring the progression of oral squamous cell carcinoma (OSCC), from the normal oral mucosa through epithelial dysplasia to invasive carcinoma. We compared differences in the NBI MOI among distinct pathological stages, so as to provide preliminary evidence for its clinical application in auxiliary diagnosis and progression assessment for OSCC. Methods: A total of 40 human oral mucosal specimens (15 normal, 15 oral leukoplakia, 10 OSCC) were enrolled for NBI image acquisition and MOI measurements. A 4-nitroquinoline-1-oxide (4NQO)-induced mouse OSCC model (n = 34) was used to dynamically record MOI changes across different pathological stages. A syngeneic tongue tumor mouse model (n = 16) was further established to evaluate whether MOI could reflect tumor formation and growth. All MOI values were quantified using ImageJ software with standardized region-of-interest (ROI) selection and background correction. Results: In clinical samples, MOI values decreased progressively from the normal mucosa (129.6 ± 5.991 arbitrary units (a.u.)) to oral leukoplakia (OLK) subgroups, including mild dysplasia (104.6 ± 3.757 a.u.) and moderate-to-severe dysplasia (91.77 ± 4.345 a.u.), and further to OSCC (54.41 ± 14.40 a.u.). In the 4NQO model, the MOI of the lingual mucosa was highest in the healthy control group (167.3 ± 10.05 a.u.) and gradually declined with increasing dysplasia severity, reaching the lowest level at the OSCC stage (48.67 ± 10.07 a.u.). In the syngeneic tumor model, the MOI was significantly lower in tumor-bearing mice than in healthy controls (47.85 ± 10.44 a.u. vs. 119.7 ± 14.20 a.u., p < 0.001). Receiver operating characteristic (ROC) analysis demonstrated good diagnostic performance of the MOI in distinguishing healthy tissue from cancerous lesions. Conclusions: NBI-derived MOI may quantitatively reflect the dynamic alterations of the oral mucosa during oral carcinogenesis and could represent a potential biomarker enabling the non-invasive, repeatable early evaluation and dynamic monitoring of OSCC.

Biomedicines doi: 10.3390/biomedicines14061232

Authors: Walaa H. El-Maadawy Eman S. El-Wakil Marwa Hassan Gamal A. Abo Sheishaa Noha F. Zahran Mohammed S. El Faramawy Mohammed H. Abdallah Eman A. Elsayed

Background/Objectives: Cryptosporidium parvum (C. parvum), a waterborne intestinal parasite, causes severe, persistent infections in immunocompromised hosts and has been linked to the onset of ileocecal adenocarcinoma. However, the molecular pathways linking chronic infection to carcinogenesis remain unclear. Nitazoxanide (NTZ), the only FDA-approved drug for this infection, shows limited efficacy. In contrast, azithromycin (AZM) possesses both antiparasitic and anticancer activity, though conclusive evidence supporting its effectiveness against cryptosporidiosis is still lacking. This study aimed to investigate the therapeutic potential of AZM against chronic cryptosporidiosis and its associated tumorigenic sequelae. Methods: Immunosuppressed mice were infected with C. parvum and treated with NTZ or AZM. Parasite burden was assessed by quantifying fecal oocyst shedding. Ileocecal tissues were analyzed for histopathology, inflammation (IL-6 and TNF-α), autophagy markers (LC3II, Beclin-1, and Atg7), PI3K/AKT signaling, and apoptotic markers (Bcl2, Bax, cleaved caspase-3, DR4, and DR5) using ELISA, real-time PCR, and Western blot. Results: Chronic C. parvum infection induced Vienna 4.4 adenocarcinoma, activated autophagy and PI3K/AKT signaling, and suppressed intrinsic and TRAIL-mediated apoptosis. AZM significantly reduced the parasitic load by 87%, outperforming NTZ (62%). It also restored epithelial integrity, attenuated inflammation, and counteracted pro-tumorigenic effects by inhibiting autophagy, downregulating the PI3K/AKT pathway, and stimulating apoptosis. Conclusions: AZM counteracted parasite-driven tumorigenic mechanisms by disrupting survival pathways and promoting apoptosis in infected and transformed cells. These findings provide evidence that AZM exerts dual antiparasitic effects and counteracts pro-tumorigenic signaling in chronic cryptosporidiosis, highlighting its potential as a therapeutic agent to prevent infection-associated ileocecal carcinogenesis.

Biomedicines doi: 10.3390/biomedicines14061231

Authors: Liumei Yang Yiyang Shi Fangfang Han Yongming Cai

Background: Drug–drug interactions (DDIs) account for about 30% of adverse drug reactions and 5–10% of hospital deaths. Combination therapy increases DDI risks, yet extracting DDIs from biomedical text remains challenging: existing methods rely on surface co-occurrence and fail when multiple drugs and interactions coexist in a sentence. Prior multimodal approaches simply concatenate text, molecular, or knowledge features without deep alignment, leading to misclassification of structurally similar but non-interacting drug pairs. Methods: We propose MultiMod-DDI, a framework that constructs a ternary evidence chain of “molecular structure–biological entities–DDI text”. Unlike existing work, MultiMod-DDI introduces (1) PS-AEGNN, a molecular graph network with ProbSparse self-attention to capture long-range chemical dependencies; (2) an adaptive position interaction vector that dynamically weights distant semantic links between drug entities; and (3) a multi-stage adaptive fusion module that sequentially applies subgraph-molecule attention and text-guided gating. These components are co-designed to enforce structured semantic alignment among heterogeneous modalities, effectively addressing the specific challenge of matching drug pairs to their correct interaction types in complex, multi-drug sentences. Results: On SemEval-2013 Task 9, MultiMod-DDI achieves 85.57% F1macro and 85.20% F1micro, outperforming state-of-the-art models. Conclusions: Through multimodal deep semantic alignment, MultiMod-DDI effectively resolves the mismatch between drug pairs and their interaction types in complex biomedical texts. The integration of multimodal features greatly improves DDI extraction accuracy, offering a reliable method for intelligent DDI mining from biomedical literature.

Biomedicines doi: 10.3390/biomedicines14061229

Authors: Chanana D. Tsayang Emily Schanzer Bonolo B. Phinius Graceful Mulenga Kesaobaka Molebatsi Kwana Lechiile Lynnette Bhebhe Tsholofelo Ratsoma Gorata G. A. Mpebe Fredah Mulenga Basetsana K. S. Phakedi Wonderful T. Choga Madisa Mine Shahin Lockman Joseph N. Jarvis Sikhulile Moyo Motswedi Anderson Simani Gaseitsiwe

Background: Concomitant HIV/HBV infection results in worse health outcomes, with HBV reactivations being observed in immunocompromised individuals. However, data on HBV infection in people with advanced HIV disease (AHD) remains sparse in Botswana. We aimed to determine the prevalence and molecular characteristics of HBV in people living with HIV (PLHIV) with CD4+ T-cell counts ≤100 cells/µL in Botswana. Methods: Plasma samples (n = 1097) of PLHIV with CD4+ T-cell count ≤100 cells/uL collected between 2014 and 2016 were screened for hepatitis B surface antigen (HBsAg) and HBV core antibodies (anti-HBc). A 415bp region of the HBV surface gene was amplified and sequenced using Sanger sequencing. Genotypic and mutational analysis was performed using Geno2pheno. Adjusted prevalence ratios (aPRs) were estimated from a modified Poisson regression model to explore factors associated with HBV infection. p-values < 0.05 indicated statistical significance. Results: The median age was 37 years (IQR: 32–43), and 565/1097 (51.5%) were male. HBsAg prevalence was 10.6% (95%CI: 8.8–12.5%) and anti-HBc prevalence was 50.0% (95%CI:46.9–52.9%). Factors associated with HBV infection were male sex [aPR: 1.6 (p < 0.01)] and those that were ART-experienced [aPR: 1.43 (p = 0.04). Eighteen samples were successfully genotyped. The prevalence of genotype A was (12/18, 66.7%) and D (6/18, 33.3%). Sixty-three mutations were identified as associated with drug resistance and immune and diagnostic escape. Highly prevalent immune escape mutations in the surface region were S207N (12/63, 19%) and A194V (9/63, 14.3%). V163I (12/63, 19%) and M129L (12/63, 19%) were highly prevalent in the reverse transcriptase region. Two classical lamivudine-associated drug resistance mutations were observed, each occurring in one participant (L180M and V173L). Conclusions: The prevalence of HBV in people with AHD is high, highlighting the importance of HBV screening and HIV/HBV co-management in this population.

Biomedicines doi: 10.3390/biomedicines14061227

Authors: Beatrice Spedicati Domizia Pasquetti Aurora Santin Stefania Zampieri Anna Morgan Stefania Lenarduzzi Giuseppe Giovanni Nardone Elisa Paccagnella Stefania Cappellani Laura Diplotti Stefano Pensiero Fulvio Parentin Paolo Gasparini Maurizio Battaglia Parodi Giorgia Girotto

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Biomedicines doi: 10.3390/biomedicines14061228

Authors: Na Sun Ziye Huang Yu Zhan

Background: The aim of this study was to evaluate the efficacy of mepolizumab as add-on therapy to intranasal corticosteroids (INCSs) for the treatment of severe, uncontrolled chronic rhinosinusitis with nasal polyps (CRSwNPs) in a real-life setting. Methods: This prospective observational study included 60 patients with severe uncontrolled CRSwNP who received mepolizumab. Follow-up assessments were performed at baseline (T0), 3 months (T1), and 6 months (T2). At each time point, patients underwent nasal endoscopy, completed the sinonasal outcome test-22 (SNOT-22), visual analogue scales (VAS) for smell, nasal obstruction and rhinorrhoea and facial pain. Nasal secretion and blood eosinophil counts (BECs) were also evaluated. The levels of eosinophil cationic protein (ECP) in nasal secretions were measured using an enzyme-linked immunosorbent assay (ELISA). Results: Both patient- and physician-derived outcome measures showed significant improvements from baseline to 3 months, and the benefits were maintained at 6 months. No major adverse events were reported. Conclusions: Mepolizumab was associated with improvements in nasal obstruction and sense of smell, based on both patient- and physician-derived outcome measures. However, due to the single-arm design and modest sample size, these findings should be considered hypothesis-generating rather than confirmatory.

Biomedicines doi: 10.3390/biomedicines14061226

Authors: Kent W. Small

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Biomedicines doi: 10.3390/biomedicines14061225

Authors: Attila Nemes Renáta Halcsik Árpád Kormányos Nándor Gyenes Kitti Rajcsány Barbara Bordács Nóra Ambrus Mohammad Nasiri Csaba Lengyel Tibor Novák

Introduction: Gestational physiology is characterized by an expansion of plasma volume and an elevation in cardiac output. Given the scarcity of existing data on pregnancy-related left atrial (LA) volumetric and functional features, this study aims to define LA volumes, volume-based functional properties and strains in healthy subjects during mid-term pregnancy. Methods: The present study comprised 19 healthy women in mid-term pregnancy (mean age: 30.5 ± 2.7 years, weight: 81.7 ± 14.0 kg, height: 166.9 ± 5.7 cm) without any symptoms, known diseases or other conditions, which could affect the results. Their results were compared to those of 43 healthy non-pregnant women (mean age: 28.6 ± 4.9 years, weight: 59.9 ± 8.5 kg, height: 167.8 ± 7.6 cm). All participants underwent comprehensive two-dimensional Doppler echocardiography with three-dimensional speckle-tracking echocardiography (3DSTE). Results: Thicker interventricular septum, increased left ventricular ejection fraction and impaired early and late transmitral flow velocities could be detected in healthy pregnant subjects as compared to those of non-pregnant individuals. End-systolic maximum LA volume was increased with elevated stroke volume and emptying fraction. While early diastolic LA volume was preserved with elevated stroke volume and emptying fraction, late diastolic LA volume, stroke volume and emptying fraction remained unchanged. However, indexed LA volumes did not differ between the groups. Among end-systolic peak global LA strains, only LA longitudinal strain (LS) was increased, while all others remained unchanged. Among regional strains, basal, midatrial and superior LA circumferential strain (CS) and LA-LS were increased except for basal LA-CS, which was impaired. Among late diastolic LA strains at atrial contraction, none of them showed any significant changes in healthy pregnant subjects compared with those of non-pregnant women. Conclusions: With a detailed 3DSTE study, elevated end-systolic LA volume and preserved diastolic LA volumes, together with enhanced end-systolic LA reservoir and early diastolic LA conduit functional properties, could be detected with features of preserved late diastolic booster pump function in healthy women during mid-term pregnancy (second trimester). When comparing indexed LA volumes, no significant difference could be confirmed between the pregnant and non-pregnant groups. This suggests that the increased end-systolic LA volume may be an adaptation to increased body weight.

Biomedicines doi: 10.3390/biomedicines14061224

Authors: Sait Erbey Mehmet Alican Sapmaz Murat Polat Ömer Osman Eroğlu Çağanay Soysal

Background and Objectives: Despite the widespread adoption of HPV-based cervical cancer screening, the optimal triage strategy for women with low-grade cytological abnormalities and non-16/18 high-risk HPV (hrHPV) types remains debated. This study evaluated the impact of ASCCP risk-based triage strategies on colposcopy referral and biopsy outcomes in a large tertiary care center. Methods: This retrospective cross-sectional study included 2748 sexually active women aged 30–65 years who underwent colposcopy at Ankara Etlik City Hospital (January 2023–June 2025). Of these, 1932 met ASCCP criteria for cervical biopsy. Cytology results, HPV genotypes (16, 18, and other hrHPV types), and histopathological findings were analyzed. CIN3+ (CIN3, adenocarcinoma in situ, or invasive carcinoma) was the primary outcome. Multivariable logistic regression identified independent predictors, with model fit assessed by Nagelkerke R2 and the Hosmer–Lemeshow test. Results: The mean age was 42.8 ± 8.1 years. The overall CIN3+ prevalence was 15.9% (308/1932). HSIL cytology was the strongest independent predictor of CIN3+ (adjusted OR 22.41, 95% CI: 11.28–44.52). HPV16/18 combined with HSIL or ASC-H cytology conferred the highest risk (adjusted OR 17.88–21.67). Women with ASC-US or LSIL cytology and non-16/18 hrHPV types had CIN3+ rates below 10%. Irregular screening history was also an independent predictor (adjusted OR 1.38). A risk-based triage approach suggested a potential reduction of approximately 29.7% in colposcopy utilization. However, this estimate applies exclusively to the biopsied subgroup and does not account for potentially undetected lesions in the 816 non-biopsied women enrolled in surveillance follow-up. Conclusions: HSIL cytology and HPV16/18 positivity represent the highest-risk profile for CIN3+ and should remain primary indications for colposcopy. Conversely, women with ASC-US or LSIL cytology and non-16/18 hrHPV types may be candidates for surveillance-based co-testing rather than immediate colposcopy, potentially enabling a resource-efficient reduction in unnecessary procedures within the biopsied cohort studied. Prospective validation in broader colposcopy-referred populations is needed before generalizing these findings to primary screening settings.

Biomedicines doi: 10.3390/biomedicines14061223

Authors: Dmitry P. Krylov Dariya M. Badanina Dmitry S. Kozlov Peter S. Timashev Daria S. Kuznetsova Artem M. Mozherov

This review systematizes the principal methods for imaging and morphological analysis of burn wounds, ranging from light, electron, and fluorescence microscopy to tomographic techniques and mass spectrometry imaging. Light microscopy with histological staining and immunohistochemistry remains the morphological gold standard, enabling visualization of the zones of coagulation, stasis, and hyperemia, as well as molecular characterization of inflammation, angiogenesis, and fibrosis. Electron microscopy allows the study of the ultrastructure of cells and the extracellular matrix at nanometer resolution. Among optical methods, wide-field indocyanine green angiography demonstrates high accuracy in burn depth stratification, whereas fluorescence lifetime imaging microscopy assesses cellular metabolism without exogenous labels. Among tomographic techniques, high-frequency ultrasound is the most accessible bedside modality with submillimeter resolution, permitting evaluation of tissue anatomy, perfusion, and biomechanical properties. magnetic resonance imaging is limited by its high cost and long examination time, while mass spectrometry imaging is used solely for research purposes. For clinical practice, the optimal combination is high-frequency ultrasound and wide-field fluorescence imaging. All methods retain high relevance for experimental research, enabling the validation of novel therapeutic strategies.

Biomedicines doi: 10.3390/biomedicines14061219

Authors: Jialin Gao Xuee Zhou Zetao Zuo Jiahong Hong Yan Tan Xiaoxiang Rong Rui Zhou Zhenhua Huang

Background/Objectives: Prostate cancer (PCa) remains a prevalent malignancy among men, often complicated by recurrence and unfavorable clinical outcomes. Consequently, precise risk stratification and timely clinical intervention are paramount. Initially, we delineated distinct expression profiles of histone modification regulators via unsupervised clustering, identifying PCa subtypes with divergent survival probabilities and biological phenotypes. Subsequently, we sought to develop a prognostic gene signature, derived from the transcriptomic variations among these regulator-defined subtypes, to predict outcomes in PCa patients following radical prostatectomy (RP). Methods: Clinical and transcriptomic data from PCa cohorts were retrieved from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) repositories for comprehensive analysis. Subtypes driven by histone modification regulators were established using unsupervised consensus clustering, followed by in-depth characterization of their molecular features and associated pathways. A risk-scoring model was then developed to evaluate its prognostic efficacy in this patient population. Results: Stratification based on histone modification regulators yielded four distinct PCa subtypes exhibiting heterogeneous survival outcomes, functional pathways, and genomic mutational landscapes. Following rigorous feature selection, a 21-gene risk signature (HIS_score)—comprising MXD3, CCDC28B, COL11A2, SLC39A5, GPT, DNASE1L2, PIF1, KRTAP5-9, TTLL10, KRTAP5-1, KRTAP5-10, HAGHL, MSLNL, AMH, NKAIN4, CCDC114, SLC9A3, SULT1E1, ALB, SLC6A14, and RPE65—was constructed. Survival analyses demonstrated that patients assigned to the high HIS_score cohort experienced significantly worse clinical outcomes compared to their low-score counterparts. Furthermore, we integrated this signature into a novel clinical nomogram to facilitate individualized prognostic assessments. Conclusions: Derived from transcriptomic disparities between extreme epigenetic subtypes, the HIS_score and its associated nomogram serve as robust prognostic instruments. These tools effectively encapsulate the downstream transcriptional sequelae of histone modification dysregulation, offering clinicians a valuable framework to accurately predict post-RP outcomes and expedite the formulation of personalized therapeutic strategies.

Biomedicines doi: 10.3390/biomedicines14061220

Authors: Jawaher A. Alsubait Dalal S. Alshaya Fatimah F. Alghnnam Mashael J. Abu-Alola Marie Fe F. Bohol Saltana A. Alhowaiti Abdullah Al Marzan Arwa A. Al-Qahtani Esra’a Abudouleh Tarek Owaidah Fatimah Alhamlan Ahmed Al-Qahtani

Background/Objectives: Genetic variation in human leukocyte antigen (HLA) genes may contribute to inter-individual differences in infectious-disease susceptibility and clinical outcomes. This study aimed to determine the genotype frequency of HLA Class I and Class II loci in patients with COVID-19 in Saudi Arabia and to examine their associations with survival and clinical severity. Methods: A prospective observational study was conducted at King Faisal Specialist Hospital and Research Centre (KFSH&RC), Riyadh, Saudi Arabia, from January 2022 to December 2023. Genomic DNA was extracted, and polymerase chain reaction-sequence specific oligonucleotide (PCR-SSO) testing was performed to screen HLA genetic variation. Patients were grouped by survival status (recovered or deceased) and clinical severity: Stage A (asymptomatic), Stage B (mild), Stage C (moderate), and Stage D (severe). Results: In total, 123 patients with COVID-19 were included; 102 (82.9%) recovered and 21 (17.1%) died. ICU admission was more frequent among deceased patients than among recovered patients (95.2% versus 51.0%, p = 0.0001). At the locus level, HLA-DPB1 represented the largest proportion of HLA calls (21%). In call-position-specific allele-group analyses, B*15 in Allele 1 (14.3% versus 1.0%, p = 0.016), C*06 in Allele 2 (42.9% versus 18.6%, p = 0.023), DRB1*10 in Allele 1 (19.0% versus 4.9%, p = 0.045), and DQB1*05 in Allele 1 (33.3% versus 11.8%, p = 0.021) were significantly more frequent among deceased patients, whereas DQB1*03 in Allele 1 was significantly more frequent among recovered patients (45.1% versus 14.3%, p = 0.013). Severity analyses showed call-position-specific differences involving C*15, C*06, B*14, B*39, B*53, and DQB1*03. Vaccination status did not differ significantly by survival status or across the four clinical severity stages. Conclusions: Selected HLA Class I and Class II allele groups may be associated with COVID-19 survival and clinical severity patterns in this Saudi cohort. These findings should be interpreted cautiously given the cohort size and call-position-specific nature of the analyses.

Biomedicines doi: 10.3390/biomedicines14061222

Authors: Tijana Vasiljević Nikola Stevan Kokanov Bojana Kožik

Thyroid cancer (TC) is the most common endocrine malignancy, with a steadily rising global incidence. Despite most cases having a favorable prognosis, a subset of patients develops aggressive, recurrent, or radioiodine-refractory disease, demonstrating the need for improved molecular biomarkers and targeted therapies. The Forkhead box P (FOXP) transcription factors (FOXP1–FOXP4) have appeared as important regulators of tumor biology, yet their roles in thyroid cancer remain incompletely defined. This review summarizes current bioinformatic, experimental, and clinical evidence regarding FOXP expression patterns, molecular mechanisms, and clinical relevance in TC. FOXP3 and FOXP4 are mainly associated with aggressive clinicopathological features, including extrathyroidal invasion, lymph node metastasis, and distant metastases, and may serve as markers of poor prognosis. The most explored FOXP3 contributes to immune evasion and radioiodine resistance by suppressing sodium iodide symporter expression and regulating tumor-associated immune responses. FOXP4 promotes tumor progression by activating key oncogenic signaling pathways and regulating non-coding RNAs. In contrast, evidence indicates that FOXP2 primarily acts as a tumor suppressor in TC by inhibiting cell proliferation and promoting apoptosis, although it may show context-dependent functions. FOXP1, though less well studied, is also suggested to have tumor-suppressive effects in some studies, and demands additional investigation in TC. Collectively, current evidence suggests that FOXP family members may represent promising diagnostic, prognostic, and therapeutic biomarkers in thyroid cancer, although further validation in large clinical cohorts and mechanistic studies is still required.

Biomedicines doi: 10.3390/biomedicines14061221

Authors: Selva Rosyta Dewi Takashi Matsumoto Titong Sugihartono Muhammad Miftahussurur Yoshio Yamaoka

Background/Objectives: Despite increasing GERD prevalence worldwide, the role of gastroduodenal microbiota in GERD phenotypes and symptom severity remains poorly understood. This study profiled mucosa-associated microbiota from the gastric antrum and duodenum across phenotypes and examined site-specific associations with symptom severity. Methods: In this cross-sectional study, forty individuals, including 26 with erosive reflux disease (ERD), 10 with non-erosive reflux disease (NERD), and 4 participants in the endoscopically normal comparator group, underwent 16S rRNA gene sequencing. Community differences were assessed using Bray–Curtis dissimilarity, differential taxa were explored by linear discriminant analysis effect size (LEfSe), and correlations with validated symptom questionnaires were evaluated. Results: Microbial community structure differed significantly between the antrum and duodenum, with Proteobacteria and Firmicutes predominating at both sites. LEfSe suggested enrichment of Streptococcus, Haemophilus, and Enterobacter in the duodenum, whereas Sphingobium, Acinetobacter, and Aquabacterium were more abundant in the antrum. The genus Helicobacter was relatively enriched in the antrum of ERD samples, whereas Streptococcus-dominant signatures were more prominent in the duodenum. Symptom severity showed stronger associations with duodenal taxa, including Fusobacterium with odynophagia, early satiety, and globus; Aquabacterium with postnatal drip and dyspnea, whereas gastric associations were fewer. Conclusions: In this small exploratory cross-sectional cohort, gastroduodenal microbiota exhibited both site-specific and phenotype-associated differences, with phenotype-related microbial variation being more evident in the duodenum than in the antrum. These hypothesis-generating findings highlight the importance of considering both anatomical context and GERD phenotype in upper gastrointestinal host–microbe interactions, and require confirmation in larger, phenotypically well-characterized cohorts.

Biomedicines doi: 10.3390/biomedicines14061218

Authors: Yiyan Li Aoxue Xing Wenzheng Li Yiman Zhang Kejuan Zhang Tianhao Xie Gang Wu Wei Zhang

Background: CAD (Carbamoyl-Phosphate Synthetase 2, Aspartate Transcarbamylase, and Dihydroorotase) is a pivotal tri-functional enzyme complex associated with the rate-limiting steps of the de novo pyrimidine biosynthetic pathway. Despite its established metabolic role, the multi-dimensional involvement of CAD in the pan-cancer landscape—specifically regarding its regulation of the metabolic–immune axis and its impact on immunotherapy response—remains to be fully elucidated. Methods: We performed a systematic pan-cancer multi-omics analysis integrating TCGA, GTEx, and DepMap datasets to evaluate CAD expression, genomic alterations, and diagnostic potential. In addition, multiple immunotherapy cohorts were integrated for meta-analysis, and metabolomic data were incorporated to explore CAD-associated metabolic features. Results: CAD was significantly upregulated in 17 cancer types, with protein-level evidence supporting this trend. CAD also showed high diagnostic accuracy in several malignancies, particularly LAML and CHOL (AUC approximately 1.0). In immunotherapy-related analyses, CAD expression was positively associated with TMB, MSI, and initial therapeutic response, but was also linked to worse long-term overall survival in pooled cohorts (HR = 1.42, 95% CI: 1.19–1.70). Integrative metabolomic analyses further suggested that high CAD expression was associated with pyrimidine metabolite accumulation and altered amino acid metabolism, indicating a potential link between CAD-related metabolic reprogramming and the tumor immune microenvironment. Conclusions: CAD may represent a promising candidate biomarker across multiple malignancies. Notably, its association with immunotherapy-related features, together with the observed discordance between response-associated signals and long-term survival, warrants further mechanistic and clinical validation.

Biomedicines doi: 10.3390/biomedicines14061217

Authors: Fumine Tanaka Toshiaki Taoka Maki Umino Ryota Kogue Hidehiro Ishikawa Yuichiro Ii Akihiro Shindo Hajime Sakuma Masayuki Maeda

Purpose: The aim of this study was to compare the contributions of putative glymphatic-related imaging indices—diffusion-weighted image analysis along the perivascular space (DWI-ALPS) index and choroid plexus volume (CPV)—and total cortical gray matter volume (TCGMV) to cognitive function in cerebral amyloid angiopathy (CAA). Methods: Forty-four CAA patients and 22 controls underwent 3.0T MRI. Cognitive function was assessed by the Mini-Mental State Examination (MMSE). The mean DWI-ALPS index, CPV/intracranial volume (ICV), and TCGMV/ICV were compared between groups; hierarchical multivariable regression and mediation analyses evaluated MMSE correlates. Results: Compared with controls, CAA showed a lower mean DWI-ALPS index and TCGMV/ICV (both adjusted p < 0.05), whereas CPV/ICV did not differ significantly after adjustment. In hierarchical multivariable regression analysis, mean DWI-ALPS index was associated with MMSE before adjustment for TCGMV/ICV (p = 0.022), but this association was attenuated after TCGMV/ICV was added to the model (p = 0.665). CPV/ICV was not associated with MMSE in either model, whereas TCGMV/ICV was independently associated with MMSE (p = 0.013). Exploratory mediation analysis suggested an indirect association between mean DWI-ALPS and MMSE via TCGMV/ICV (indirect: p = 0.023; direct: p = 0.720). Conclusions: Cortical atrophy appeared to be the strongest imaging correlate of cognitive impairment in CAA, while the association between DWI-ALPS and MMSE in multivariable models was attenuated after accounting for cortical gray matter volume. The ALPS index may provide indirect information on glymphatic-related pathways, but its biological specificity in CAA requires cautious interpretation because ALPS measurements may be influenced by underlying microstructural alterations in white matter.

Biomedicines doi: 10.3390/biomedicines14061216

Authors: Attila Nemes Renáta Halcsik Árpád Kormányos Nándor Gyenes Kitti Rajcsány Barbara Bordács Nóra Ambrus Mohammad Nasiri Csaba Lengyel Tibor Novák

Introduction. Pregnancy is characterized by a significant expansion of plasma volume and an increase in cardiac output, necessitating structural and functional adaptations of the cardiac chambers, including the right atrium (RA). To evaluate these changes, three-dimensional (3D) speckle-tracking echocardiography (STE) was used as a validated and sophisticated modality for the concurrent assessment of RA volumetric and functional alterations. This study aimed to characterize RA volumes, volume-based functional indices, and strain parameters in healthy women during mid-gestation, compared with a cohort of non-pregnant controls. Methods. This retrospective cohort analysis included 20 healthy, asymptomatic women in their second trimester (mean age: 29.9 ± 3.0 years; weight: 81.2 ± 14.2 kg; height: 166.9 ± 5.8 cm; body surface area [BSA]: 1.95 ± 0.17 m2). The control group consisted of 30 age-matched healthy non-pregnant women (mean age: 29.9 ± 4.1 years; weight: 58.7 ± 6.5 kg; height: 166.0 ± 5.4 cm; BSA: 1.68 ± 0.11 m2). All subjects underwent comprehensive two-dimensional Doppler echocardiography and 3DSTE. Results. Early and late diastolic RA volumes were significantly reduced, despite preserved end-systolic RA volume. Pregnant subjects exhibited reduced active RA stroke volume and increased passive RA emptying fraction, while all other parameters remained comparable between the groups. No significant differences were observed between groups in end-systolic peak RA global or mean segmental strains, nor in RA strains measured during atrial contraction. However, end-systolic peak regional RA strain analysis revealed decreased basal RA circumferential strain (CS) and increased superior RA-CS in pregnant participants compared with controls. Furthermore, during late diastole (at atrial contraction), superior RA-CS, RA-3D strain, and RA-area strain were significantly higher in healthy pregnant subjects than in controls. Conclusions. Substantial regional changes in RA function were detected by 3DSTE, likely reflecting adaptation to pregnancy-induced plasma volume expansion, and resulting in significant RA volumetric changes.

Biomedicines doi: 10.3390/biomedicines14061215

Authors: Stefanie Bonini Patricia Fuentes Richard Brannon Claytor

Introduction: Chronic wounds and pathologic scars remain a persistent challenge in plastic surgery. Conventional treatments can be costly and inconsistent, prompting interest in regenerative approaches that utilize autologous tissue. Emulsified fat produces nanofat through mechanical processing and contains adipose-derived stem cells, stromal vascular fractions, extracellular matrix proteins, cytokines and growth factors. The purpose of this systematic review is to evaluate the use of autologous nanofat for wound healing and scar management, with emphasis on preparation techniques, treatment indications, and outcomes. Methods: A comprehensive PubMed search with no date restrictions was conducted in January 2026 using MeSH terms and keywords related to nanofat and wound-healing applications. Studies were screened independently by two reviewers using the Rayyan platform. Eligible studies evaluated nanofat for wound healing in human or animal subjects; non-English articles, studies not involving nanofat, editorials, and conference abstracts were excluded. The extracted data included study characteristics, participant numbers, treatment details, indications, adjunct therapies, follow-up duration, outcomes, and complications. Studies were grouped by clinical application, with individual reports included in multiple categories when relevant. Results: The search identified 53 records, of which 22 studies met the inclusion criteria after screening. These included 20 human and two animal studies spanning randomized controlled trials (n = 3), prospective trials (n = 6), retrospective analyses (n = 6), case series (n = 4), and case reports (n = 3). Mechanical emulsification was the predominant autologous nanofat preparation method (91%), often combined with filtration or centrifugation. Clinical indications in human studies were diverse, most commonly including scar treatment (n = 14) (acne, burns, depressed, and post-surgical), followed by chronic wounds (n = 3) and reconstructive applications (n = 3). Nanofat was administered via injection in 86% of studies (n = 19), typically using fine-gauge needles or microcannulas with intradermal or subdermal placement, while three studies used non-injection approaches such as topical, membrane, or dressing-based delivery. Scar or aesthetic parameters, measured using VSS, POSAS, physician grading, photography, pigmentation analysis, or clinical appearance, were evaluated in 73% of studies (n = 16), and all reported improvement in variables such as pigmentation, pliability, thickness, texture, or overall appearance. Wound-healing endpoints were assessed in 36% (n = 8), with 100% (n = 8) demonstrating accelerated healing, improved epithelialization, or defect closure. Patient-reported outcomes, including satisfaction or quality of life, were measured in 32% (n = 7), and all showed improvement. Objective imaging modalities (e.g., 3D imaging, ultrasound, angiography, digital analysis) were used in 23% (n = 5), each confirming structural or physiologic improvement. Histologic or biomolecular analyses were performed in 27% (n = 6) and uniformly demonstrated regenerative changes, such as increased angiogenesis, collagen remodeling, or growth factor expression. Treatment was well tolerated, with 77% of studies (n = 17) reporting minimal or no complications and only transient mild adverse effects, including mild pain, bruising, erythema, and edema. Conclusions: Current evidence suggests that autologous nanofat is a promising regenerative therapy for wound healing and scar modulation. Across diverse clinical applications, nanofat has been associated with improved tissue quality, enhanced healing, and favorable patient-reported outcomes, with minimal complications. The mechanical processing of autologous tissue may also involve fewer regulatory concerns compared with more extensively manipulated cellular products.

Biomedicines doi: 10.3390/biomedicines14061214

Authors: Hatice Kumru Aina Ros-Alsina Agustín Hernandez-Navarro Eloy Opisso Margarita Vallès Jesus Benito-Penalva Joan Vidal Miquel Sarrio Loreto García-Alén

Background: Transcutaneous spinal cord stimulation (tSCS) is a promising approach to enhance functional recovery after spinal cord injury (SCI). However, evidence on repeated sessions and their effects on gait and lower-limb strength remains limited. This study evaluated the effects of multisegmental tSCS on walking ability, muscle strength, and functional independence in individuals with SCI. Methods: In this randomized controlled trial with a partial crossover design, twelve individuals received tSCS combined with gait rehabilitation, while ten underwent gait rehabilitation alone, for three weeks. Four participants crossed over to the tSCS group after a minimum one-week washout period following the control intervention. We assessed the American Spinal Injury Association Impairment Scale (AIS), Total Motor Score (TMS), Lower Extremity Motor Score (LEMS), Walking Index for Spinal Cord Injury II (WISCI-II), 10- and 6-Meter Walking Tests (10MWT, 6meterWT), Timed Up and Go (TUG) test, maximal voluntary contraction (MVC) of the quadriceps (QM) and tibialis anterior (TA), and the Spinal Cord Independence Measure (SCIM-III); tSCS was applied at three spinal segments during gait rehabilitation over 15 sessions. Results: tSCS significantly improved MVC in both muscles, as well as SCIM-III and TUG, and these improvements were maintained at follow-up, with no significant adverse events reported. Other clinical assessments also showed significant improvement in both groups. Conclusions: tSCS was well tolerated and conferred additional benefits in lower-limb muscle strength, walking ability (as assessed by TUG), and functional independence, supporting its potential as a valuable adjunct to rehabilitation.

Biomedicines doi: 10.3390/biomedicines14061207

Authors: Liwen Zhang Tianqi Liu Na Yu Ruijian Zhang Zhepeng Luo Xiaoqing Zhang Jiani Wang

Background: Intestinal fibrosis is a severe complication of Crohn’s disease (CD) with no effective therapies currently available. Muscleblind-like protein 1 (MBNL1) is an RNA-binding protein that has been implicated in fibrosis across multiple organs, but its role in CD-associated intestinal fibrosis remains unexplored. This study aims to investigate the expression, functional role, and underlying mechanism of MBNL1 in intestinal fibrosis. Methods: MBNL1 expression was examined in a TNBS-induced mouse model and in stenotic intestinal tissues from CD patients. In vitro, human colonic fibroblasts (CCD-18Co) were stimulated with transforming growth factor-β1 (TGF-β1) to model fibrosis. MBNL1 was knocked down or overexpressed to assess its effects on fibroblast activation, proliferation (5-ethynyl-2′-deoxyuridine, EdU; Cell Counting Kit-8, CCK-8), and apoptosis (flow cytometry). Potential downstream pathways were predicted using BioGRID and DAVID analyses and validated by Western blot. A rescue experiment with the RAS activator ML-097 was performed to confirm pathway dependency. Results: MBNL1 expression was significantly upregulated in fibrotic tissues from both the mouse model and CD patients, as well as in TGF-β1-stimulated CCD-18Co. MBNL1 knockdown suppressed TGF-β1-induced fibroblast activation and proliferation while promoting apoptosis, whereas MBNL1 overexpression had the opposite effect. Mechanistically, MBNL1 positively regulated the RAS-MAPK signaling pathway. Reactivation of this pathway with ML-097 reversed the inhibitory effects of MBNL1 knockdown on fibroblast activation and proliferation. Conclusions: MBNL1 promotes colonic fibroblast activation and proliferation by activating the RAS-MAPK signaling pathway, establishing it as a potential therapeutic target for intestinal fibrosis in Crohn’s disease.

Biomedicines doi: 10.3390/biomedicines14061209

Authors: Murong Wang Chunying Liu Xin Wei

Glaucoma is a chronic progressive optic neuropathy and one of the leading causes of irreversible blindness worldwide. Although elevated intraocular pressure remains the most important modifiable risk factor, increasing evidence suggests that immune dysregulation and autoimmune responses also contribute substantially to disease onset and progression. Clinical studies across different glaucoma subtypes have identified subtype-dependent immune abnormalities, including altered serum autoantibody profiles, dysregulated cytokine and chemokine expression, and changes in peripheral immune cell subsets. Experimental and translational studies further indicate that multiple immunopathogenic mechanisms are involved in glaucomatous neurodegeneration, including glial cell-mediated immune responses, activation of pattern recognition receptor signalling pathways, adaptive immune responses, and complement cascade dysregulation. These processes may interact to sustain chronic neuroinflammation, promote retinal ganglion cell injury, and accelerate optic nerve degeneration. Importantly, a better understanding of immune involvement in glaucoma has generated growing interest in immunomodulatory therapy as a potential strategy beyond intraocular pressure lowering. Targeting microglial activation, inflammatory signalling pathways, adaptive immune imbalance, and complement-mediated injury has shown neuroprotective potential in animal or in vitro models, whereas clinical evidence in glaucoma patients remains limited. These findings may provide preliminary directions for future therapeutic development. In this review, we summarise the current clinical evidence linking glaucoma with autoimmunity, discuss the major immune mechanisms implicated in disease pathogenesis, and highlight recent advances in immunomodulatory therapeutic strategies. Elucidating the immune basis of glaucoma may help pave the way for more precise and effective treatments for this complex optic neuropathy. We believe that immune dysregulation in glaucoma functions as a context-dependent amplifier of retinal ganglion cell injury rather than a uniform primary driver, with innate (microglia/astrocytes), adaptive (T/B cells, HSP-specific immunity), and complement pathways interacting to sustain neuroinflammation and neurodegeneration. This integrated immune response contributes to subtype- and stage-specific vulnerability, and targeting these maladaptive immune mechanisms represents a promising, precision-guided strategy for neuroprotection beyond intraocular pressure lowering.

Biomedicines doi: 10.3390/biomedicines14061213

Authors: Errikos Petsas Gerasimos Siasos Thomas Mavromoustakos Christos T. Chasapis

Βackround/Objectives: The rising global burden of cardiometabolic disorders and chronic low-grade inflammation underscores the need for therapies capable of modulating multiple interconnected pathways. Methods: In this work, a ligand-based virtual screening campaign centered on a previously reported scaffold (compound 1a) was combined with molecular docking, 200 ns molecular dynamics simulations and ADMET prediction to identify and prioritize small-molecule multitarget candidates against PCSK9, GLP1R, FGFR1, GIPR, NF-κB and NLRP3. Results: Among the screened analogs, D4Z emerged as the most balanced lead, displaying consistently favorable binding profiles, stable interactions within functionally relevant pockets and a drug-like physicochemical and pharmacokinetic profile with high predicted oral absorption. Conclusions: Although these findings remain purely computational, they support D4Z as a prioritized multitarget lead for synthesis and experimental validation and illustrate the potential of rational multitarget design for addressing the cardiometabolic–inflammatory axis.

Biomedicines doi: 10.3390/biomedicines14061212

Authors: Guangyuan Cheng Suting Sun Guoshu Deng Ying Luo Miao Li Hang Zhao Xiaofan Yang Ruiping Wang Le Kuai Ying Zhang Bin Li Yi Ru Jiankun Song

Atopic dermatitis (AD) is a chronic inflammatory skin disease driven by immune dysregulation and epidermal barrier dysfunction, in which eosinophils act as key effector cells contributing to tissue damage and persistent inflammation. This comprehensive review elucidates the multifaceted contributions of eosinophils to the progression of AD. Driven by key type 2 cytokines (notably IL-4, IL-5, and IL-13) and specific chemokines, eosinophils infiltrate lesional skin and undergo IgE-mediated degranulation. The subsequent release of cytotoxic granule proteins, including major basic protein (MBP), eosinophil cationic protein (ECP), eosinophil-derived neurotoxin (EDN), and eosinophil peroxidase (EPX), directly induces keratinocyte apoptosis, exacerbates tissue remodeling, and sustains the local inflammatory cascade. Furthermore, we explore the intricate crosstalk between eosinophils and sensory neurons, which, alongside cytokines like IL-31, profoundly aggravates chronic pruritus. Consequently, modulating eosinophil activation and recruitment has emerged as a vital therapeutic approach. We systematically evaluate current and emerging pharmacological interventions, ranging from conventional topical corticosteroids to advanced targeted therapies. Particular emphasis is placed on the mechanistic impact of novel biologics and small-molecule Janus kinase (JAK) inhibitors, demonstrating how they attenuate eosinophilic inflammation. By identifying current gaps in this field, this review provides valuable insights for future research and clinical practice in the field of AD.

Biomedicines doi: 10.3390/biomedicines14061205

Authors: Michael A. Leone Georgia Kurman Madeline Bright Peter K. Gregersen Christine N. Metz

Quercetin is a naturally occurring flavonoid found in fruits, vegetables, and teas that is widely available as a dietary supplement. Numerous studies have investigated quercetin’s therapeutic potential across a broad range of diseases and conditions. Collectively, these studies reveal its anti-inflammatory, antioxidant, anti-proliferative, anti-cancer, anti-fibrotic, antibacterial, endocrine-modulating, and senolytic properties, establishing quercetin as a polypharmacologic agent with diverse biological activities. This review describes quercetin’s biochemical properties, bioavailability, and proposed mechanisms of action. It highlights the unique characteristics of the human uterus vs. other species and evaluates published evidence from pre-clinical and clinical studies supporting quercetin’s pleiotropic effects and potential therapeutic benefits for six uterine-related conditions: endometrial cancer, endometriosis, adenomyosis, uterine infections, uterine fibroids, and polycystic ovary syndrome (PCOS). The findings support that quercetin targets multiple endometrial and other uterine cell types and may attenuate key pathological processes relevant to uterine disease. However, robust human clinical evidence supporting quercetin’s efficacy is generally lacking. Critical knowledge gaps and translational barriers to advancing quercetin from a ‘promising preclinical candidate’ into an ‘evidence-based therapeutic’ for improving uterine health are discussed.

Biomedicines doi: 10.3390/biomedicines14061210

Authors: Monica Loguercio Domenico Mario Giamundo Alessia Giglio Emanuela Buda Marco Ambrosetti Francesco Perone

The gut microbiota has emerged as a key regulator of cardiovascular health, influencing metabolic, inflammatory, and vascular pathways. Growing evidence indicates that gut dysbiosis, characterized by reduced microbial diversity, depletion of beneficial short-chain fatty acid–producing bacteria, and enrichment of pro-inflammatory taxa, is associated with major cardiovascular risk factors and disease progression. Microbial-derived metabolites, including trimethylamine/trimethylamine N-oxide, short-chain fatty acids, amino acids and bile acids, may play a central role in modulating lipid metabolism, endothelial function, inflammation, and thrombosis, although the underlying mechanisms remain incompletely understood. Recent multi-omics approaches have expanded this understanding by identifying personalized microbiome–metabolome signatures linked to cardiovascular risk, supporting a shift toward precision medicine. In this review, we summarize current evidence on the composition and functional role of the gut microbiota in cardiovascular disease and critically discuss emerging microbiota-targeted strategies. These include dietary interventions, prebiotics, probiotics, synbiotics, antibiotics, enzyme inhibitors, and fecal microbiota transplantation, which may contribute to both the prevention and adjunctive treatment of cardiovascular conditions. In addition, we address the challenges of integrating gut microbiota management into clinical practice and highlight the importance of tailored strategies, including exercise-based interventions, microbial enzyme inhibitors, and postbiotics. Despite promising preclinical and early clinical data, the translation of microbiome-based therapies into routine practice remains limited by heterogeneity in study design, the lack of standardized protocols, and incomplete mechanistic understanding. Overall, targeting the gut microbiota represents a novel and potentially complementary approach for cardiovascular disease prevention and management, warranting further well-designed clinical studies.

Biomedicines doi: 10.3390/biomedicines14061211

Authors: Viviana A. Ruiz-Pozo Santiago Cadena-Ullauri Rafael Tamayo-Trujillo Patricia Guevara-Ramírez Elius Paz-Cruz Alejandro Cabrera-Andrade Ana Karina Zambrano

Telomere length (TL) is a key determinant of cellular aging and genomic stability, influenced by genetic, molecular, and environmental factors. Progressive telomere shortening has been associated with degenerative and cardiovascular diseases, whereas longer telomeres have been linked to an increased risk of cancer, highlighting a dual and context-dependent relationship between TL and disease susceptibility. Evidence from genome-wide association studies (GWAS) and Mendelian randomization analyses indicates that TL is a highly heritable and polygenic trait, regulated by variants in genes such as TERT, TERC, RTEL1, and POT1, as well as components of the shelterin and CST complexes. This study integrates genetic variants associated with telomere shortening and elongation, including their functional classification, proposed molecular mechanisms, and ACMG/AMP categorization, together with global and Latin American allele frequency data. These variants may participate in key processes such as telomerase activity, telomerase RNA stability, and telomere replication, thereby influencing susceptibility to multiple diseases. However, current evidence is largely derived from European and Asian populations. Given the highly admixed nature of Latin American populations, population-specific studies are required to identify unique genetic determinants and to improve the application of precision medicine.

Biomedicines doi: 10.3390/biomedicines14061208

Authors: Tianfu Liu Yichen Huang Yizhe Wang Rui Zhao Haili Shen

Background: Recruitment and activation of monocyte-derived macrophages (MoMFs) sustain cholangitis in primary biliary cholangitis (PBC), but whether MoMFs amplify inflammation through ferroptosis remains unclear. We defined ferroptotic programs in MoMFs and evaluated the calpain/ACSL4 axis as a regulatory and therapeutic node. Methods: We analysed a public human liver single-cell RNA sequencing (scRNA-seq) dataset and examined MoMF-associated ACSL4 and 4-hydroxynonenal (4-HNE) signals in CD11b+CD68+ cells by multiplex immunofluorescence. We used a 2OA–BSA-induced PBC-like mouse model to assess liver injury, inflammation and ferroptosis-related markers and tested Liproxstatin-1 (Lip-1), rosiglitazone (ROSI) or the calpain inhibitor PD150606. Bone marrow-derived macrophages (BMDMs) from control and PBC mice were profiled and challenged with RSL3, with or without Ferrostatin-1 (Fer-1), ROSI or PD150606. Results: MoMFs were expanded in PBC livers and showed the strongest induction of ferroptosis signatures, centered on ACSL4, with enhanced inflammatory crosstalk with cholangiocytes. Human PBC tissues showed increased CD11b+CD68+ cells positive for ACSL4 or 4-HNE. In PBC-like mice, malondialdehyde (MDA) increased and glutathione (GSH) decreased, and macrophages showed greater colocalization with ferroptosis markers; Lip-1, ROSI or PD150606 improved liver biochemistry, reduced inflammation scores and limited macrophage infiltration. PBC-derived BMDMs upregulated ACSL4 and CAPN1/2 and were more sensitive to RSL3; Fer-1, ROSI or PD150606 attenuated ferroptosis-associated molecular changes. Conclusions: MoMF ferroptosis is prominently engaged in PBC, and our findings implicate a pharmacologically tractable calpain/ACSL4 axis that may contribute to macrophage ferroptotic susceptibility and inflammatory liver injury.

Biomedicines doi: 10.3390/biomedicines14061206

Authors: Jan Soukop Martin Poruba Zuzana Rácová Iveta Zapletalová Hana Malínská Martina Hüttl Irena Marková Rostislav Večeřa

Background: Metabolic syndrome (MetS) is a cluster of risk factors increasing the likelihood of cardiovascular and metabolic diseases. Objectives: This study investigated the relative mRNA expression of key hepatic and intestinal phase II drug-metabolising enzymes, specifically UDP-glycosyltransferases (UGTs) and sulfotransferases (SULTs), in four non-obese rat models of MetS characterised by different dominant traits: the hereditary hypertriglyceridaemic (HHTg) rat, spontaneously hypertensive rat (SHR), SHR-expressing transgenic human C-reactive protein (SHR-CRP) rat, and bilaterally ovariectomised female Wistar (W-OVX) rat, compared to Wistar controls. Methods: Gene expression was quantified by RT-PCR with data normalised using the ΔΔCt method. Results: Measurements showed significant model-specific differences, especially in the liver. HHTg rats exhibited significant hepatic suppression of Ugt1a9 (−90%) and undetectable Ugt2b transcripts, alongside compensatory upregulation of Sult1a1 (196%) and Sult1b1 (277%). The SHR model showed significant hepatic upregulation of Ugt1a1 (330%), Sult1a1 (266%), and Sult1b1 (328%). Chronic inflammation in SHR-CRP rats caused a significant decrease in hepatic Ugt1a1, whereas a significant induction occurred in the intestine. Oestrogen deprivation (W-OVX) led to significant downregulation of hepatic Ugt1a6 and Ugt1a9. Conclusions: These findings highlight that the alterations in phase II metabolism strongly depend on the pathophysiological context, potentially affecting drug disposition in preclinical models.

Biomedicines doi: 10.3390/biomedicines14061204

Authors: Baglan Kazhiyakhmetova Nursulu Altaeva Meirat Bakhtin Pavel Tarlykov Yasutaka Omori Shinji Tokonami Chutima Kranrod Radhia Pradana Saowarak Musikawan Anel Lesbek Danara Ibrayeva Elena Saifulina Dana Auganova Moldir Aumalikova Madina Kairullova Aigerim Shokabayeva Dinara Bizhanova Yerlan Kashkinbayev

Background: Radon exposure is the second most important risk factor for lung cancer after tobacco smoking and represents a significant but often underestimated public health problem. Due to the absence of specific clinical manifestations at early stages, the identification of molecular biomarkers reflecting early radon-induced carcinogenic processes is of particular importance. The aim of this study was to identify protein biomarkers associated with radon exposure in lung cancer patients residing in settlements of the Akmola and North Kazakhstan regions of Kazakhstan. Methods: Indoor radon exposure was assessed using CR-39 detectors to measure radon concentrations in residential dwellings during summer and autumn periods. The study included 57 lung cancer patients and 73 control subjects residing in areas characterized by varying levels of radon exposure. Plasma samples were collected and analyzed using liquid chromatography–tandem mass spectrometry (LC–MS/MS) to identify differentially expressed proteins associated with lung cancer and radon exposure. Statistical analyses were performed to evaluate differences between groups and associations between radon exposure and molecular biomarkers. Results: Seasonal variability in indoor radon concentrations was observed, with several settlements demonstrating levels exceeding international reference values. Proteomic analysis identified multiple proteins differentially expressed between lung cancer patients and controls, as well as between radon-exposed and non-exposed lung cancer patients. Several proteins involved in inflammation, lipid metabolism, oxidative stress, and immune regulation pathways demonstrated significant differences in expression levels, suggesting potential associations with radon-induced carcinogenic mechanisms. LC–MS/MS proteomic profiling identified multiple differentially expressed proteins associated with lung cancer and radon exposure after false discovery rate correction. Proteins involved in inflammation, oxidative stress, immune regulation, and lipid metabolism, including ORM2, AZGP1, PRDX2, IRF7, and APOC3, demonstrated significant expression differences between radon-exposed and low-exposure groups. Conclusions: The identified protein biomarkers demonstrated significant associations with both radon exposure and lung cancer status, indicating their potential relevance for early detection and risk assessment of radon-induced lung cancer. The integration of environmental exposure assessment with proteomic profiling may provide new insights into the molecular mechanisms of radon-associated carcinogenesis and support the development of preventive strategies.

Biomedicines doi: 10.3390/biomedicines14061203

Authors: Patrycja Wszelaki Aleksandra Karczmarska Szymon Szymoniuk Grzegorz Grześk Zbigniew Włodarczyk Joanna Sikora

The article presents the results of a structured literature review of the past decade and focuses on the crucial role of platelets in the pathogenesis of pulmonary arterial hypertension (PAH). It explores how endothelial dysfunction initiates early prothrombotic signals that activate platelets, which, in response, adopt a pro-inflammatory phenotype, release cytokines and chemokines, and form aggregates with leukocytes, thereby modulating their migration and activity. A key feature of PAH is the “platelet paradox,” in which chronic in vivo activation coexists with reduced ex vivo reactivity due to functional exhaustion. Prolonged stimulation and disease progression lead to complex hemostatic dysregulation, characterized by heterogeneity in platelet phenotypes. At the same time, platelets undergo immunometabolic reprogramming, with a predominance of glycolysis, over oxidative phosphorylation, mitochondrial dysfunction, altered fatty acid oxidation (FAO), increased lactate production, and enhanced vesicle release. These phenomena sustain inflammation and promote pulmonary vascular remodeling. This study aims to review the current mechanisms of immunometabolic platelet activation in pulmonary arterial hypertension. It primarily focuses on platelet aspects as key elements in disease progression and as potential sources of new biomarkers and therapeutic targets.

Biomedicines doi: 10.3390/biomedicines14061202

Authors: Ahmed Massoud Amina E. Essawy Mohammed A. Alfredan Ashraf M. Abdel-Moneim Rehab A. Gomaa Sherine Abdel Salam

Background/Objectives: Betanin (BET), a prominent phytochemical mainly derived from Beta vulgaris, exhibits strong anti-inflammatory and antioxidant activities owing to its distinctive chemical structure. Nevertheless, its potential analgesic effect in the context of inflammatory pain remains insufficiently explored. Accordingly, this study investigated the analgesic effects of BET in a complete Freund’s adjuvant (CFA)-induced rat model of inflammatory pain. Methods: Rats received a single subcutaneous injection of 100 µL CFA to induce inflammatory pain, followed by oral administration of BET at doses of 40 or 80 mg/kg/day for 14 days. Results: BET treatment significantly reduced paw edema and improved HPL (hot plate latency) in CFA-injected rats. Biochemically, in the ipsilateral spinal cord of rats, BET at both 40 and 80 mg/kg significantly increased IL-4, and only the 80 mg/kg dose significantly reduced oxidative stress (MDA) and IL-1β. TNF-α levels were slightly reduced at both doses and did not reach statistical significance versus CFA. At the molecular level, miR-107 was significantly downregulated by BET at 80 mg/kg (but not 40 mg/kg), while miR-145 was significantly upregulated by both 40 mg/kg and 80 mg/kg compared to CFA. Pearson’s correlation indicated that miR-107 was positively correlated with MDA, IL-1β and TNF-α but negatively with IL-4, whereas miR-145 was positively correlated with IL-4 but negatively with IL-1β. PCA biplot analysis corroborated these findings, showing simultaneous presence of MDA, IL-1β, TNF-α, and miR-107 with CFA, and IL-4 and miR-145 were only related to control and CFA+BET80 groups. In addition, using transmission electron microscopy imaging, we found that BET alleviated neuronal damage in CFA-treated rats. Furthermore, molecular docking analysis predicted that BET may exhibit stable binding interactions with several inflammation- and apoptosis-related targets, including AKT1, mTOR, IKKβ, TNF-α, IL-1β, COX-2, caspase-3, caspase-7, and caspase-8, supporting its multi-target anti-inflammatory and antiapoptotic effects. Conclusions: Overall, our data suggest that BET can possibly exert analgesic effects in CFA-induced inflammatory pain by modulating oxidative stress and favoring a shift toward an anti-inflammatory status. These effects coincided with downregulation of miR-107, overexpression of miR-145, and improvements in inflammatory pain behaviors. Further investigations are required to validate the involvement of specific miRNA- and pathway-mediated effects. Nevertheless, our findings highlight BET as a promising natural candidate for future development of anti-inflammatory and analgesic strategies.

Biomedicines doi: 10.3390/biomedicines14061201

Authors: Rahima Aftab Asher Fawwad Eraj Abbas Ruqaya Nangrejo Fasiha Fatima Syed M. Shahid

Background/Objectives: The rising global burden of type 2 diabetes mellitus (T2DM) demands multifaceted and more effective treatment strategies beyond monotherapy to achieve optimal metabolic control. The study aimed to evaluate the integrated effects of SGLT2 inhibitors and metformin in newly diagnosed T2DM patients on biochemical parameters, clinical outcomes and hormonal changes. Methods: This prospective longitudinal study was conducted at the Department of Biochemistry, Baqai Medical University, in collaboration with the Baqai Institute of Diabetology and Endocrinology. A total of 120 newly diagnosed T2DM patients were enrolled and stratified into three groups (n = 40): Group 1 (SGLT2 inhibitors only), Group 2 (SGLT2 inhibitors + metformin), and Group 3 (metformin only). Patients were followed for six months with data collection at baseline, at 3 months and 6 months. Anthropometric indices (weight, BMI, waist and hip circumferences, WHR), biochemical markers (FBS, HbA1c, lipid profile, uric acid, serum creatinine, HOMA-IR), and hormonal levels (insulin, glucagon) were assessed at baseline, first follow-up, and second follow-up. ANOVA, post hoc, Bonferroni and Tukey’s tests were applied; p-value < 0.05 was considered significant. Results: The findings indicate that Group 2 showed the greatest improvement in anthropometric parameters, particularly waist and hip circumferences (p < 0.01). Group 3 demonstrated the most significant improvement in glycemic indices and lipid profile (p < 0.01). HOMA-IR significantly decreased in Group 3 from baseline to the first follow-up (p < 0.01). While insulin levels remain insignificantly different among all groups. Glucagon levels declined significantly from baseline to the second follow-up in all groups, with a more pronounced decrease in Group 3 (p < 0.01). Serum creatinine and uric acid levels showed significant reductions from baseline to the second follow-up in Group 1 and Group 2 (p < 0.05). However, given the observational design, these associations should not be interpreted as causal evidence of renoprotection. Conclusions: Within the limitations of this observational study, early differences among treatment regimens were observed, though metabolic outcomes became statistically comparable across groups by six months. These hypothesis-generating findings suggest potential benefits of early combination therapy that require confirmation in randomized controlled trials. Given the substantial within-group variability and non-randomized design, no definitive conclusions about therapeutic associations can be drawn from these data.

Biomedicines doi: 10.3390/biomedicines14061200

Authors: Zorana Maric Ostovic Katarina Mijacic Isidora Kostic Nevena Gajovic Milena Jurisevic Bojana Simovic Markovic Vladimir Markovic Sanja Zornic Snezana Jovanovic Stevic Bojan Kujundzic Srdjan Masic Dragana Drakul Ivan Jovanovic

Background: The newly synthesized palladium(II) complex [Pd(L1)Cl]Cl (where L1 = N2,N6-bis(5-methylhthiazol-2-yl)pyridine-2,6-dicarboxamide) has demonstrated significant in vitro antitumor activity. In this study, the effects of this complex on the immune response and its antimicrobial potential were evaluated. Methods: Splenocytes isolated from mice were treated with lipopolysaccharide (LPS)/Concanavalin A (ConA) along with the Pd(II) complex. The concentrations of IFN-γ, IL-17, IL-4, TNF-α, IL-1β, and IL-10 were measured using commercial ELISA kits. The antimicrobial effect was tested against reference strains of Gram-positive and Gram-negative bacteria, as well as yeast. The Minimal Inhibitory Concentration (MIC) was determined via the broth microdilution method, followed by the determination of Minimal Bactericidal/Fungicidal concentrations (MBC/MFC). Results: The Pd(II) complex induced an increase in the concentration of all tested cytokines compared to untreated cells. Co-treatment with Pd(II) complex and LPS significantly increased the levels of IFN-γ, IL-1β, and IL-17 compared to the LPS-only-stimulated group. Co-treatment with ConA and the Pd(II) complex resulted in a significant increase in TNF-α and IL-17 levels, whereas a significant decrease was observed in the concentrations of IL-10, IL-4, and IFN-γ compared to the ConA-only-stimulated group. The tested complex showed weak to moderate antimicrobial activity, Gram-positive bacteria showed better susceptibility to the examined complex compared to Gram-negative. Conclusions: Results of the study indicate that the Pd(II) complex exhibits a significant immunomodulatory effect on splenocytes, alongside weak to moderate antimicrobial activity.

Biomedicines doi: 10.3390/biomedicines14061199

Authors: Bulat I. Yalaev Rita I. Khusainova Ildar R. Minniakhmetov Dmitry D. Panteleev Madina I. Yevloyeva Minara S. Shamkhalova Yulia Y. Golubkina Ekaterina A. Dobreva Marina V. Shestakova Natalia G. Mokrysheva

Background: Diabetic retinopathy (DR) and chronic kidney disease (CKD) are among the leading causes of disability in individuals with type 1 diabetes mellitus (T1DM). However, the genetic architecture of these complications remains poorly understood. Genome-wide studies demonstrate significant interpopulation heterogeneity, while candidate gene studies yield conflicting results due to limited power. Independent replication of previously obtained results in separate cohorts, with appropriate intergroup comparisons, is essential for identifying the most significant biomarkers of microvascular complications in T1DM. Purpose: To search for associations of the most significant polymorphic variants rs55703767, rs72831309, rs118124843, rs9344715, rs183937294, rs4293393, rs12917707, rs77924615, rs11864909, rs9622363, rs73885319, rs2523989, rs3825932, rs763361, rs12708716, rs2292239, and rs4948088 with the risk of developing T1DM and its complications—DR and CKD. Methods: The study involved 618 individuals, including 522 patients with T1DM undergoing inpatient treatment at the Endocrinology Research Centre, as well as 96 control individuals without T1DM. Among the T1DM patients, 232 had concurrent CKD and retinopathy, while 80 were free of both microvascular complications. A comparison of allele and genotype frequencies of 17 single-nucleotide polymorphisms (SNPs) was conducted between the T1DM group and the control group, as well as an intergroup comparison between individuals with and without complications. Results: The rs2292239 (ERBB3) locus is associated with an increased risk (pbonf = 0.001, OR = 2.02), while rs55703767 (COL4A3) is associated with a decreased risk of developing T1DM in general (p = 0.01846, OR = 0.42). rs9344715 (AKIRIN2) is associated with the risk of diabetic nephropathy (p = 0.03996, OR = 1.29), while PDILT variants rs77924615 (OR = 0.57, pbonf = 0.045) and rs11864909 (OR = 0.41, pbonf = 0.0105) with DR. Conclusions: The study identified potential genetic markers for the risk of type 1 diabetes and its microvascular complications. The results require further verification in an independent, expanded cohort. Consideration of genetic factors confirmed the independent contribution of the identified variants, supporting their value as promising biomarkers for risk stratification and personalized prevention of T1DM complications.

Biomedicines doi: 10.3390/biomedicines14061198

Authors: Manjur Kolhar Raisa Nazir Ahmed Kazi Ahmed M. Al Rajeh

Background/Objectives: Although deep learning methods have achieved promising performance in recent years, comparatively less attention has been given to explicitly modeling periodic and multi-scale temporal dynamics for ECG-specific representation learning within TimesNet-based frameworks. In this work, we propose an ECG-specific TimesNet-based framework for multi-label classification of multi-lead ECG recordings that incorporates periodicity-aware temporal modeling. Methods: The proposed framework utilizes Fast Fourier Transform (FFT)-guided temporal decomposition to identify dominant frequency components and reshapes ECG sequences into period-aligned representations to better capture intra-period morphological patterns and inter-period rhythm dependencies. Multi-scale convolutional TimesBlocks are further employed to learn rhythm-aware and morphology-aware temporal representations. Results: The proposed framework was evaluated on the PTB-XL dataset using two experimental settings: Three-Class classification (NORM, AFIB, PVC) and Five-Class classification (NORM, AFIB, MI, PVC, STTC). Experiments were conducted using a one-vs-rest multi-label learning strategy with independent class probability estimation. The framework achieved mean one-vs-rest test AUC values of 0.956 and 0.913 for the Three-Class and Five-Class settings, respectively. Experimental results indicated that the reduced classification complexity in the Three-Class setting was associated with improved feature separability, more stable decision boundaries, and enhanced discriminative representation learning. Latent-space visualization using UMAP and PCA demonstrated clearer clustering in the Three-Class configuration, while gradient-based interpretability analysis highlighted physiologically relevant ECG waveform regions contributing to model predictions. In addition, computational profiling demonstrated practical feasibility with approximately 1.957 million trainable parameters, 13.14 GFLOPs computational complexity, 5.230 ms average inference latency per ECG recording, and a throughput of approximately 191 ECG recordings per second on GPU hardware. Conclusions: These findings suggest that periodicity-aware temporal modeling can improve ECGF representation learning while demonstrating practical potential for computationally efficient and interpretable automated ECG analysis applications.

Biomedicines doi: 10.3390/biomedicines14061197

Authors: Imane Achir Alispahic Josefin Eklöf Pradeesh Sivapalan Alexander Ryder Jordan Zitta Barrella Harboe Tor Biering-Sørensen Katja Biering Leth-Møller Allan Linneberg Jens-Ulrik Stæhr Jensen

Background: Chronic obstructive pulmonary disease (COPD) is a progressive respiratory condition where many patients are given antibiotics like amoxicillin and macrolides (clarithromycin, azithromycin, roxithromycin) for bacterial infections. Recent concerns about clarithromycin’s potential link to cardiovascular events have arisen, despite its effectiveness against respiratory pathogens. This study aims to compare the cardiovascular risk of macrolide antibiotics versus amoxicillin in suspected COPD patients. Method: We used the Danish National Health Service Prescription Database (DNHSP) to identify COPD patients and their use of antibiotics. The included COPD patients were divided into four groups: amoxicillin users, roxithromycin users, clarithromycin users and azithromycin users. Data from multiple registries were merged to track hospitalizations, causes of death, and major adverse cardiovascular events (MACEs) as the primary endpoint. Patients were followed for a 3-year period. We applied adjusted Cox regression and sensitivity analyses with IPTW and IPCW to address confounders and censoring. Results: Our study involved 45,869 patients who were prescribed a long-acting muscarinic antagonist, over the age of 40 years old and who received one of the following antibiotics: amoxicillin, azithromycin, clarithromycin, or roxithromycin. No increased risk of MACEs was observed in macrolide-treated patients compared to those treated with amoxicillin (azithromycin: HR 0.97: 95% CI 0.83–1.13 p = 0.69, clarithromycin: HR 1.06 95% CI 0.87–1.28 p = 0.57, roxithromycin: HR 1.04 95% CI 0.91–1.18 p = 0.60), as confirmed by the sensitivity analysis (azithromycin: HR 0.95 95% CI 0.82–1.11 p = 0.52, clarithromycin: HR 1.05 95% CI 0.87–1.27 p = 0.60, roxithromycin: HR 1.05 95% CI 0.92–1.19 p = 0.48). Similarly, hazard ratios for all-cause mortality and cardiovascular death among the antibiotic groups showed no significant statistical differences. Conclusions: These findings suggest that there is no difference in the risk of MACEs, all-cause mortality, or cardiovascular death between the amoxicillin group and the macrolide group in a large and unselected population of COPD patients.

Biomedicines doi: 10.3390/biomedicines14061196

Authors: Jiaxun Zhang Akezhouli Shahatiaili Yuhan Hou Ning Zhou Ke Huang Xiaojun Wang Dongmei Wang Zhentao Yu Xiaoli Feng Yibo Gao

Background: As the most lethal neuroendocrine tumor, small cell lung cancer (SCLC) can drive its progression by hijacking neuronal mechanisms. At the core of this neural integration is the N-methyl-D-aspartate receptor (NMDAR) complex. However, its pan-cancer expression and clinical significance in SCLC remain poorly understood. Methods: We characterized NMDAR transcriptomic profiles across human cancers to develop the NMDAscore, and analyzed three independent European and Asian SCLC cohorts to identify prognostic biomarkers. Furthermore, we investigated the molecular mechanisms of GRIN2A and evaluated the efficacy of GluN2 inhibitors. Results: The developed NMDAscore exhibited significant prognostic correlations in ACC, COAD, KIRC, UVM, KIRP, OV, PCPG, UCS, THCA, THYM, HNSC, KICH, LGG, and PAAD. Focusing on the SCLC cohorts, we identified GRIN2A (encoding the GluN2A subunit) as a statistically relevant prognostic biomarker associated with poor survival. Mechanistically, GRIN2A upregulation correlates with the activation of neuro-synaptic signaling, metabolic reprogramming, genomic instability, and an immune-cold microenvironment characterized by CD8+ T cell exclusion. Pharmacological inhibition of GluN2 using dizocilpine and the FDA-approved antagonist memantine suppressed SCLC proliferation and tumorigenicity in vitro, in 3D tumor spheroids and in vivo xenograft models. Conclusions: Collectively, these findings establish GRIN2A as a prognostic biomarker, linking synaptic hijacking, metabolic plasticity, immune evasion, and drug resistance, and identify the therapeutic potentials of the GluN2 inhibitors dizocilpine and memantine for SCLC.

Biomedicines doi: 10.3390/biomedicines14061195

Authors: Peng-Yuan Chang Mao-Hsien Wang Yu-Ling Yeh Kuo-Chi Chang Hung-Sheng Soung

Background/Objectives: Rotenone (RT)-induced neurotoxicity is widely used to model Parkinsonism-like nigrostriatal injury and recapitulates several PD-relevant pathological features, including oxidative stress, mitochondrial dysfunction, neuroinflammation, and dopaminergic neurochemical disturbance. Loganin (LG), an iridoid glycoside isolated from Cornus officinalis, has been reported to possess antioxidant, anti-inflammatory, anti-apoptotic, and neuroprotective properties. However, its protective effects in a unilateral stereotaxic RT lesion model have not been fully elucidated. This study aimed to investigate the neuroprotective potential of LG against RT-induced Parkinsonism-like pathology in rats and to explore the possible involvement of antioxidant-related signaling mechanisms. Methods: Adult male Wistar rats were randomly assigned to twelve experimental groups (n = 8/group), including control, sham, RT, sham + LG, RT + LG, RT + trigonelline (TG) + LG, and RT + selegiline (SL). RT was stereotaxically injected once into the right substantia nigra pars compacta (SNpc) on Day 0 to induce unilateral nigrostriatal injury. LG was administered orally once daily from Day 1 to Day 21 at doses of 3, 10, and 30 mg/kg. TG was given intraperitoneally 30 min before LG treatment, while SL served as a reference antiparkinsonian drug. Behavioral assessments and biochemical analyses were conducted to evaluate motor dysfunction, oxidative and nitrosative stress, endogenous antioxidant status, mitochondrial dysfunction, inflammatory and apoptotic responses in the SNpc, and striatal catecholamine disturbances. Results: RT lesioning produced significant motor deficits, oxidative and nitrosative stress, depletion of endogenous antioxidant defenses, mitochondrial dysfunction, inflammatory and apoptotic activation in the SNpc, and abnormalities in striatal catecholamine levels. LG treatment significantly attenuated these pathological changes, with more pronounced protective effects observed at 10 and 30 mg/kg. Co-administration of TG partially weakened the beneficial effects of LG, suggesting the possible involvement of antioxidant defense-related signaling while not providing direct proof of a single pathway. SL also ameliorated RT-induced behavioral and biochemical abnormalities. Conclusions: These findings suggest that LG confers multi-target neuroprotective effects against RT-induced Parkinsonism-like features in rats. The protective actions of LG were associated with attenuation of oxidative stress, mitochondrial dysfunction, neuroinflammation, apoptosis, and catecholaminergic disturbances. Because the pathway analysis remains pharmacological and indirect, additional studies using direct molecular validation are warranted before LG can be considered a disease-modifying candidate for PD-related neurodegeneration.

Biomedicines doi: 10.3390/biomedicines14061194

Authors: Rachael E. Clifford Sulaimaan Hannan Hamish W. Clouston Victoria Lavin Claire Arthur Paul A. Sutton

Background: Local recurrence of rectal cancer is a challenging problem for patients and clinicians. Surgical resection is associated with good outcomes if R0 margins are achieved; however, it is often complex, requires suitable patient fitness, and is associated with long term physical and psychological consequences. Meanwhile, continuing technical advances in radiotherapy have enabled the delivery of highly conformal treatment, thereby enabling dose escalation or pelvic reirradiation to be safely considered—either as definitive management or in the neoadjuvant setting—for patients with locally recurrent rectal cancer. Pelvic reirradiation may refer to patients who have received primary rectal radiotherapy with the aim of neoadjuvant downstaging or reducing the risk of locoregional recurrence, versus radiotherapy for a previous unrelated non-rectal pelvic malignancy. Methods: A literature search of pelvic reirradiation for non-metastatic, locally recurrent rectal cancer was conducted for full text articles published over the last 20 years. Additional papers were identified within the references of these papers. Studies focusing on non-rectal cancers, and patients having primary radiotherapy for locally recurrent rectal cancer were excluded. Due to the heterogenicity of the data, no meta-analysis was performed. Results: A total of 15 papers were included, containing a cohort of 840 patients. Several reirradiation modalities were reported, including external beam radiotherapy, brachytherapy, stereotactic ablative radiotherapy and heavy particle therapy (carbon ion). Carbon ion radiotherapy was the most common reirradiation treatment modality utilised with a median cumulative dose of 70.4 Gray (Gy). Treatment response, defined as either complete or partial improvement in tumour size, was only reported in seven studies, and varied from 14 to 88%. Overall 3-year survival was also variable with rates reported between 18 and 85%. These observations may be due to variation in patient selection, treatment intent, and technique. Pelvic reirradiation was associated with acceptable toxicity, low rates of G3+ toxicity, and improved symptom control. Conclusions: Our review describes the multitude of approaches to pelvic reirradiation for locally recurrent rectal cancer. Reviewing the radiobiological and patient outcomes is challenging in view of the degree of heterogeneity in patient selection, treatment approach, and reported outcomes. However, there is consensus that pelvic reirradiation—either for long term control or to downstage prior to definitive surgery—is feasible with potential utility in this setting.

Biomedicines doi: 10.3390/biomedicines14061193

Authors: Tony D. Baldini Soren D. Johnson Aneesh S. Bhat Mengyao Liu Andrea C. Filler Mark A. Lee J. Kent Leach Maryam Rahmati Augustine M. Saiz

Background: Fracture healing requires a coordinated inflammatory response, and its dysregulation, as seen in polytrauma, can impair bone regeneration. Human mesenchymal stem cells (hMSCs) play a central role in fracture repair through osteogenic differentiation and also via their secretome, which regulates local inflammation, angiogenesis, and tissue regeneration. Interleukin-6 (IL-6), an early pro-inflammatory cytokine, contributes to fracture healing by promoting MSC recruitment and osteogenic differentiation, whereas bone morphogenetic protein-2 (BMP-2) is a key osteoinductive factor that drives bone formation. However, the combined effects of IL-6 and BMP-2 on the hMSC secretome remain poorly understood. Methods: We cultured hMSCs in osteogenic media supplemented with recombinant IL-6 (1–20 ng/mL) alone or combined with recombinant BMP-2 (1 ng/mL) on tissue culture plastic (TCP) and within gelatin methacryloyl (GelMA) hydrogels of low (~3 kPa), medium (~15 kPa), and high (~30 kPa) stiffness. Osteogenic differentiation was assessed by alkaline phosphatase (ALP) activity and calcium deposition; cytokine profiling was performed using a multiplex antibody array. Results: When cultured on TCP, IL-6 suppressed ALP activity by day 21. Co-treatment with IL-6 and BMP-2 induced a dysregulated secretome with concurrent upregulation of pro-inflammatory markers (MIP-1α, TNF-α, and GM-CSF) and anti-inflammatory mediators (IL-10, TGF-β1, and VEGF). This hyperinflammatory response was attenuated when hMSCs were encapsulated in GelMA, with high-stiffness gels most effectively suppressing pro-inflammatory chemokines and medium-stiffness gels yielding the highest ALP activity. Conclusions: These findings suggest that mechanically tuned GelMA hydrogels modulate immune and osteogenic responses of hMSCs in vitro, warranting further investigation in the context of scaffold design for fracture care.

Biomedicines doi: 10.3390/biomedicines14061192

Authors: Alexandra E. Meyer Natalie G. Alexander Elisa M. Tucker Hallie E. Knight Benjamin T. Klemp Bryan J. Estrada Sarah F. Hathcock Henry D. Mauser Kylie A. Buchanan Brandon J. Kim

Background: The blood–brain barrier (BBB) separates the circulation from the central nervous system (CNS) and serves to maintain brain homeostasis. The BBB comprises highly specialized brain endothelial cells (BECs) with unique properties that allow the BBB to maintain strict regulation of molecules entering and exiting the CNS. These characteristics include tight junctions, low endocytosis rates, and efflux and nutrient transporters. Breast cancer resistance protein (BCRP) is an efflux transporter found at the BBB that plays a key role in protecting the CNS. Together with other efflux transporters, BCRP contributes to multidrug-resistant cancers and difficulty delivering drugs and therapeutics to the brain and other organs. Methods: Using the hCMEC/D3 line, we utilized BCRP substrate rosuvastatin to effectively select for cells expressing high amounts of BCRP, thus generating hCMEC/D3-BCRP. To assess protein abundance, we utilized flow cytometry and confirmed expression via qPCR. To investigate BCRP efflux function in evolved hCMEC/D3-BCRP, we performed substrate accumulation assays with BCRP and P-gp substrates. Results: We found hCMEC/D3-BCRP had increased BCRP abundance and expression relative to parent hCMEC/D3. We also observed an increase in BCRP function via substrate accumulation of two BCRP substrates compared to parent hCMEC/D3. Conclusions: BCRP serves a protective role within the BBB and is a major hurdle in drug delivery. We generated a BCRP overexpression BEC cell line (hCMEC/D3-BCRP) under the influence of endogenous promoters. This cell line can be used to further investigate the role of BCRP in BECs and utilized in efflux transport studies.

Biomedicines doi: 10.3390/biomedicines14061191

Authors: Hasan Zeybek Tugrul Hosbul

Objectives: This study aimed to evaluate very low HBV DNA viral load below the limit of quantification and to identify correlational factors in different patient groups, including individuals with chronic hepatitis B (CHB), occult HBV infection (OBI), and others. Methods: We retrospectively analyzed 390 patients with very low-level viremia (VLLV). HBV DNA levels were measured in plasma samples using real-time quantitative PCR (qPCR). Serological markers were evaluated in serum samples using chemiluminescence microparticle immunoassay (CMIA). Demographic variables, HBV serological markers (anti-HBs, anti-HBe, anti-HBc), and DNA results were evaluated. Results: The study included 193 CHB patients with maintained virological suppression and 197 patients in the other group; of which, 60 patients had occult hepatitis B infection (HBV DNA positive, HBsAg negative) and 137 had no occult hepatitis B infection. Very low viral load was more common in men (53.3%) and in individuals aged ≥50 years (63.3%). In univariate analysis, OBI was associated with anti-HBe (odds ratio (OR) = 2.874, 95% CI: 1.255–6.579, p = 0.013), and anti-HBc seropositivity (OR = 5.750; 95% CI: 2.626–12.591, p < 0.001). In multivariate analysis, anti-HBe positivity and anti-HBc positivity were independently associated with OBI. Anti-HBs positivity was independently and inversely associated with OBI. Conclusions: In patients with VLLV cohort, anti-HBc and anti-HBe seropositivity were independently associated with detectable but unquantifiable HBV DNA. Although anti-HBe positivity reflects reduced viral replication, it does not indicate complete viral suppression and may be detected at very low viremia levels, especially in occult HBV infection. These findings highlight the complex interplay between viral replication dynamics and host immune responses across the VLLV spectrum, characterize the serological landscape associated with detectable but unquantifiable HBV DNA, and warrant validation in prospective studies.

Biomedicines doi: 10.3390/biomedicines14061190

Authors: Yulia A. Tunakova Svetlana V. Novikova Vsevolod S. Valiev

Background: Lipid peroxidation is a primary driver of biological membrane damage and mediates the relationship between environmental exposure and adverse health outcomes. Malondialdehyde (MDA) is a widely recognized biomarker for quantifying oxidative stress intensity. Despite numerous studies on oxidative stress and metal exposure, nonlinear relationships between physiological characteristics, serum metal profiles and MDA levels in pubertal children remain insufficiently studied. Methods: The study included 105 conditionally healthy children aged 12–14 years from urban and rural regions of Tatarstan, Russia. Serum MDA concentrations were determined spectrophotometrically using the thiobarbituric acid assay, while Zn, Cu, Fe, Sr and Pb concentrations were measured by atomic absorption spectrometry. A multilayer perceptron neural network was applied to model nonlinear relationships between MDA levels, environmental exposure indicators and morphophysiological characteristics. Because the original relational dataset contained partially replicated participant-derived relational structures, primary validation was performed using independently reconstructed datasets without repeated observations. Additional repeated cross-validation and SHAP-based feature importance analysis were performed. Results: Urban-residing children demonstrated significantly higher serum MDA levels than rural counterparts, independent of sex, with girls consistently showing higher values. Reduction of predictor dimensionality improved model generalization behaviour. Validation using independently reconstructed datasets without repeated observations demonstrated reproducible exploratory predictive behaviour of the reduced neural network model, with independently reconstructed validation datasets yielding mean R2 values of 0.901 ± 0.052 and 0.914 ± 0.046, respectively. SHAP analysis demonstrated that zinc, copper and iron consistently represented the dominant contributors to the nonlinear model, although substantial variability in the relative ranking of zinc and copper was observed between validation datasets. Conclusions: The proposed neural network model demonstrated the ability to capture reproducible nonlinear relationships between oxidative stress markers and environmental exposure parameters in a limited biomedical dataset. The model should primarily be interpreted as an exploratory explanatory tool rather than an individual clinical prediction instrument. Because of the limited dataset size, partially reconstructed relational structure and exploratory study design, the findings require cautious interpretation and further external validation.

Biomedicines doi: 10.3390/biomedicines14061189

Authors: Shiwang Tan Ju Lai Heng Zhi Wei Tang Ling Jin Shaoqing Yu

Background/Objectives: Chronic rhinosinusitis with nasal polyps (CRSwNP) is a heterogeneous inflammatory disease in which eosinophilic subclassification is widely used for clinical stratification. However, it remains unclear how closely nominal histologic eosinophilic labels reflect the broader molecular organization of diseased tissue. Methods: We performed an inference-based integrative analysis of public datasets spanning discovery single-cell RNA sequencing (scRNA-seq), independent scRNA-seq validation, GeoMx digital spatial profiling, and bulk transcriptomic replication cohorts. A sample-level molecular burden framework was constructed using four dimensions: type 2 inflammation, epithelial injury/remodeling, extracellular-matrix remodeling, and barrier/defense impairment. Composite burden and component-level features were then examined across nominal eosinophilic categories, epithelial states, spatial compartments, and independent bulk cohorts. Results: Nominal eosinophilic labels were directionally informative but incompletely concordant with molecular burden. In the discovery cohort, eosinophilic CRSwNP samples were enriched toward the higher-burden end, whereas nominally non-eosinophilic CRSwNP samples extended across the intermediate-to-high burden range. Across discovery and validation scRNA-seq datasets, GeoMx spatial profiling, and independent bulk cohorts, the most reproducible burden-associated signals centered on epithelial injury/remodeling-like programs and related remodeling features. In the epithelial compartment, higher burden was associated with epithelial state reorganization, stronger wounding-associated activity, and trajectory-linked glandular/secretory remodeling. Independent validation and spatial analyses further supported epithelial wounding-, barrier-, and myeloid remodeling-related features, whereas type 2 context signals were directionally consistent but less uniform across platforms. In bulk replication, composite burden, epithelial wounding, and myeloid remodeling were more consistent across cohorts than type 2 context alone. Conclusions: Nominal eosinophilic labels in CRSwNP capture clinically relevant but incomplete information about underlying tissue biology. Epithelial injury/remodeling-like programs and remodeling-linked myeloid features emerged as the most stable organizational axes of molecular burden across public multimodal datasets. These findings support a graded, multidimensional view of CRSwNP and may complement, rather than replace, conventional pathology-based eosinophilic subclassification.

Biomedicines doi: 10.3390/biomedicines14061188

Authors: Litong Zhu Taoyan Lin Lai Yee Cheong Jason K. H. Sher Irene Y. L. Yam Wynn Cheung Susan Yung Tak Mao Chan Desmond Y. H. Yap

Aim: Disturbances in exhausted and classical memory B cells have been implicated in the pathogenesis of systemic lupus erythematosus (SLE) and lupus nephritis (LN), but the genetic regulation of their homeostasis remains poorly understood. Methods: We analyzed the single-cell RNA-seq data of peripheral blood mononuclear cells (PBMCs) from the NIH SLE dataset (GSE135779) and another published LN single-cell RNA-seq dataset (dbGAP database accession code phs001457.v1.p1). Overlapping differentially expressed genes (DEGs) in exhausted and classical memory B cells from SLE and LN patients were identified, and their altered expression was validated in B cells obtained from LN patients. GO and KEGG analyses were used to analyze associated pathways. The relationships between exhausted and classical memory B cells and cellular metabolic pathways were also assessed. Results: Three DEGs (IFI44L, XAF1, and MX1) were detected in both exhausted and classical memory B cells, and their increased expression was verified in classical and exhausted memory B cells obtained from LN patients during remission. The protein–protein interaction network of the DEGs suggested that STAT1 showed the highest eigenvector centrality for these DEGs. IFI44L, XAF1 and MX1 were involved in distinct biological processes and immune pathways (especially JAK-STAT). Classical memory B cells showed higher expression of genes involved in sulfur metabolism (SQRDL and TST), amino sugar metabolism (GFPT1 and UAP1), and butanoate metabolism (ACADS and ACAT1), while exhausted B cells exhibited inverse relationships with these metabolic pathways. Conclusions: Altered expression of IFI44L, XAF1 and MX1 is associated with distinct metabolic signatures and immune pathways in exhausted and classical memory B cells in SLE and LN.

Biomedicines doi: 10.3390/biomedicines14061187

Authors: Filippo Capilupi Valentino Condoleo Giandomenico Severini Giuseppe Armentaro Corrado Pelaia Ilaria Gareri Pasquale Loiacono Maria Rosangela Scarcelli Francesco Maruca Alberto Panza Marilisa Panza Sofia Miceli Raffaele Maio Angela Sciacqua

Introduction: Obstructive sleep apnea (OSA) is highly prevalent, affecting up to 50% of individuals over 65 years. Elderly patients often present with atypical, fewer and less severe symptoms, suggesting age-specific phenotypes. However, comprehensive clinical phenotyping that incorporates cognitive outcomes remains limited. This study aimed to characterize OSA phenotypes through cluster analysis and evaluate their association with cognitive impairment, independently of age. Materials and Methods: Between 2020 and 2024, 409 adults with moderate-to-severe OSA were enrolled and stratified into three age groups (<65, 65–74, ≥75 years). All underwent home sleep apnea testing (HSAT), comprehensive symptom assessment, Epworth Sleepiness Scale (ESS), and Montreal Cognitive Assessment (MoCA, pathological ≤ 25 pts). Hierarchical cluster analysis (Ward’s method) used AHI, T90, BMI, and ESS. Logistic regression identified independent predictors of cognitive impairment. Results: Older groups showed lower BMI, higher comorbidity burden, fewer symptoms, and greater cognitive impairment prevalence (4.5% vs. 9.7% vs. 45.9%; p < 0.001), despite comparable polysomnographic severity across age groups. Cluster analysis identified three phenotypes: Cluster 1 (classical OSA: high AHI, BMI, T90, ESS); Cluster 2 (geriatric phenotype: low AHI, BMI, T90, ESS, highest cognitive impairment rate: 27.7%); Cluster 3 (hypersymptomatic: low AHI and T90, high sleepiness and asthenia, prevalent depression). On multivariate regression, age (OR 1.155; p < 0.001), male sex (OR 2.223; p = 0.034), and Cluster 2 (OR 3.131; p < 0.001) were independent predictors of cognitive impairment. Conclusions: Three clinically distinct OSA phenotypes were identified regardless of age and severity. The geriatric phenotype was associated with three-fold increased risk of cognitive impairment, supporting routine cognitive screening and age-adapted diagnostic strategies in elderly OSA patients.

Biomedicines doi: 10.3390/biomedicines14061186

Authors: Renata Jabłońska Robert Ślusarz Agnieszka Królikowska Karolina Filipska-Blejder Magdalena Zając Paweł Sokal

Background/Objectives: Early functional status at hospital discharge is a clinically relevant outcome after aneurysmal subarachnoid hemorrhage (aSAH), but early prognostic assessment remains challenging. We evaluated whether admission inflammatory blood cell ratios were associated with discharge independence and added prognostic information beyond established neurological severity scales. Methods: In this retrospective single-center cohort study, 252 consecutive adults with aSAH were screened, and 144 endovascularly treated patients with available admission complete blood count with differential were included. Discharge independence was defined as a Barthel Index score ≥60 at hospital discharge. A clinical reference model included age, World Federation of Neurosurgical Societies (WFNS) grade, and Hunt–Hess grade. Multivariable logistic regression was used to assess associations between inflammatory ratios and discharge independence. Discrimination was assessed using receiver operating characteristic analysis with DeLong’s test, and the final model was internally validated by bootstrap resampling. Results: Forty-one patients (28.5%) achieved discharge independence. Higher admission neutrophil-to-lymphocyte ratio (NLR) was independently associated with lower odds of discharge independence (adjusted odds ratio 0.47 per interquartile range increase, 95% CI 0.24–0.90; p = 0.022). Adding NLR to the clinical reference model improved discrimination (AUC 0.790 vs. 0.737; p = 0.039), with an optimism-corrected AUC of 0.767 after bootstrap validation. Other inflammatory indices did not significantly improve discrimination. Conclusions: In this single-center retrospective cohort of endovascularly treated patients with aSAH, admission NLR was independently associated with discharge independence and provided modest incremental prognostic information beyond established neurological severity scales.

Biomedicines doi: 10.3390/biomedicines14061185

Authors: Dominik Jucha Michał Klimas Dominika Wiśniewska Martyna Winiarska Mateusz Szczupak Jacek Kobak Sabina Krupa-Nurcek

Background/Objectives: Bipolar disorder affects about 40 million people worldwide, and the greatest burden of the disease is associated with depressive episodes. About 25% of patients experience drug-resistant depression, in which standard treatment turns out to be insufficient, and monotherapy often does not bring full remission. Despite the use of second-generation antipsychotics, the effectiveness of therapy in TRBD remains limited, which necessitates rational polypharmacotherapy and augmentation strategies. The paper discusses the receptor mechanisms of drug combination, current therapeutic regimens and new interventions such as ketamine acting on the glutamate anergic system. The aim was to synthetically compare the efficacy and safety of available augmentation strategies and polypharmacotherapy. Methods: The material consists of published clinical, observational and randomized trials on pharmacotherapy of drug-resistant bipolar depression, including atypical neuroleptics, ketamine, pramipexole, modafinil, lamotrigine, celecoxib and memantine. The authors analyze receptor mechanisms, neurobiological data and clinical trial results, comparing them with current definitions of TRBD according to ISBD and CINP. Biomarker data, such as the Systemic Immune-Inflammation Index, and the results of neuroimaging and metabolomic studies were also used in the work. Results: The analysis showed that atypical neuroleptics showed limited efficacy and high rates of side effects, while ketamine has the fastest and most pronounced antidepressant effect with a low risk of phase change. Pramipexole has shown promise in terms of long-term efficacy, but its use reduces the high risk of induction of mania and impulse control disorders. Celecoxib as an anti-inflammatory therapy significantly increased response and remission rates compared to escitalopram alone, and memantine showed only an early, short-term antidepressant effect. The results highlight that TRBD requires targeted polypharmacotherapy, with the most promising directions being glutamatergic modulation and anti-inflammatory therapies. Conclusions: Drug-resistant bipolar depression requires a departure from classical monotherapy in favor of rational, mechanistically justified polypharmacotherapy, targeting complex monoaminergic, glutamatergic and neuroinflammatory disorders. Available data indicate that ketamine has the greatest clinical potential among the current strategies, characterized by a rapid onset of action and a favorable safety profile compared to atypical neuroleptics or dopamine agonists. Modulation of inflammatory processes with the use of celecoxib also has promising results, which highlights the importance of biomarkers and personalization of therapy. However, further, large, and well-designed studies are needed to unambiguously determine optimal treatment strategies for TRBD and to verify the effectiveness of new pharmacological interventions.

Biomedicines doi: 10.3390/biomedicines14061184

Authors: Belgheis Ebrahimi Xu Cindy Yang Carol Wang Yiming Yue Johnson Liu Jun Lu John A. Taylor

Background/Objectives: Marine-derived bioactive compounds have attracted increasing interest due to their potential antiviral properties. This study investigated in vitro antiviral activity of oil extracted from the green-lipped mussel (Perna canaliculus, GLM) and low-molecular-weight (LMW) fucoidan from Undaria pinnatifida against three human viruses in mammalian cell systems. herpes simplex virus-1 (HSV-1), respiratory syncytial virus (RSV), and SARS-CoV-2. These marine compounds were selected with the longer-term aim of evaluating their combination as a potential synergistic antiviral strategy. Methods: Antiviral efficacy was assessed using complementary assay platforms, including plaque reduction assays in mammalian cell systems and a lentiviral pseudovirus system delivering a bioluminescent reporter gene in HEK293/ACE2 cells pseudotyped with the SARS-CoV-2 spike glycoprotein. Cytotoxicity was assessed in parallel, and the selectivity index (SI) was calculated as the ratio of CC50 to IC50 for each compound and virus tested. Results: GLM oil showed potential antiviral activity against SARS-CoV-2 pseudovirus (SI > 6.20), with limited activity against RSV (SI > 3.48) and HSV-1 (SI > 2.28). In contrast, LMW fucoidan did not demonstrate antiviral activity against any of the tested viruses. Conclusions: These findings support further investigation of GLM-derived bioactive compounds as potential antiviral agents, including studies to elucidate their mechanisms of action and in vivo studies to confirm their antiviral efficacy. Combination studies were not pursued in the present work as both compounds require further optimisation individually; however, future studies should evaluate their combined antiviral potential, as synergistic or additive effects remain plausible.

Biomedicines doi: 10.3390/biomedicines14061183

Authors: Deyan Donchev Ralitsa Nikolova Katya Vaseva Hristo Taskov Mariana Murdjeva Michael Maes Ivan Nikolaev Ivanov

Background: Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) and long COVID-19 syndrome (LC) show substantial clinical overlap, but direct comparative microbiome studies remain limited. Methods: In this cross-sectional study, we compared the fecal gut microbiome of patients with ME/CFS, LC, and healthy controls (HC) within a unified analytical framework using 16S rRNA profiling, differential abundance testing, and multivariate modeling. We also examined associations between microbiome variation and questionnaire-derived symptom-domain scores. Results: Alpha-diversity did not differ significantly among groups, whereas beta-diversity analyses showed small but significant disease-associated community differences with broad overlap between cohorts. Differential abundance analysis identified stronger signals in disease-versus-control contrasts than in the direct ME/CFS vs. LC contrast. Both ME/CFS and LC shared enrichment of Sutterella and depletion of Terrisporobacter and Lachnospiraceae relative to HC. Predicted functional profiling showed shared disease-versus-control changes in pathways related to anaerobic acetate/H2 carbon flow, inositol/polyol degradation, phosphonate/C1-related metabolism, and lysine-derived fermentation. Regression analyses showed the strongest microbiome associations with fatigue-related and physiosomatic domains, while affective, cognitive, and gastrointestinal outcomes showed weaker signals. Conclusions: Overall, these findings support the presence of overlapping but non-identical gut microbiome alterations in ME/CFS and LC. The results provide a basis for future longitudinal and multi-omics studies aimed at clarifying the stability, functional relevance, and clinical utility of these microbial patterns.

Biomedicines doi: 10.3390/biomedicines14061182

Authors: Olga Butranova Yury Kustov Anna Abramova Sergey Zyryanov Irina Asetskaya Elizaveta Terekhina Vitaly Polivanov

Background: Pulmonary fibrosis (PF) is a major global health issue associated with substantial morbidity across all age groups. One of the important etiological factors contributing to PF is drug-induced lung injury, which can result from both direct and indirect damage to the pulmonary parenchyma caused by various pharmacological agents, including chemotherapeutics, antirheumatic drugs, cardiovascular medications, and certain antimicrobial agents. The aim of our study was to assess the structure of antibacterials involved in drug-induced PF (DIPF) and analyze signals of DIPF, calculating the reporting odds ratio (ROR) and proportional reporting ratio (PRR) using spontaneous reports (SRs) extracted from the Russian National Pharmacovigilance database. Methods: A retrospective, descriptive pharmacoepidemiological analysis of SRs from the AIS database for the period 1 April 2019–31 March 2025 was conducted. Results: A total of 130 SRs with data on DIPF associated with antibacterial agents were identified, with patients’ mean age of 59.1 ± 14.46 years. Death was reported in 65 SRs (50%) with a mean age of 53.0 ± 13.66 years. Next, antibacterials were identified as leaders: sulfamethoxazole (used alone or in combination with trimethoprim, 20.7% (n = 50)), azithromycin (18.2%, n = 44), levofloxacin (12.4%, n = 30), doxycycline (11.6%, n = 28), and cefuroxime (10.7%, n = 26). Disproportionality analysis performed with PRR and ROR calculation revealed the strongest association with DIPF for cefuroxime (PRR = 15.11, 95% confidence interval, CI: 10.25–22.27; ROR = 15.31, 95% confidence interval, CI: 10.33–22.68). Conclusions: Cefuroxime was revealed as a drug with an unexpected but robust safety signal for DIPF, warranting heightened clinical awareness and further investigation. The observed associations between antibacterial agents and DIPF should be interpreted with caution, as they may reflect protopathic bias (antibiotics prescribed for early symptoms of unrecognized pulmonary fibrosis) or context-dependent biological effects rather than true pro-fibrotic drug properties. Our findings do not establish causality but rather generate safety signals that warrant validation through prospective studies with detailed clinical phenotyping and mechanistic investigations using human cell lines.

Biomedicines doi: 10.3390/biomedicines14061181

Authors: Xuanyu Jin Junkai Yang Daojia Miao Wei Xiong Zhiyong Xiong

Renal cell carcinoma (RCC) is a predominant malignancy of the urinary system, with clear cell renal cell carcinoma (ccRCC) representing 75–85% of clinical cases. Since the early stages are often asymptomatic, nearly 30% of patients present with metastases at diagnosis, which significantly complicates the prognosis. The diverse mechanisms and clinical indications of current strategies, despite recent breakthroughs in immunotherapy, pose a major challenge for systematic application. This review employs the cancer-immunity cycle as a framework to evaluate four critical steps: antigen presentation, T-cell activation, reversal of exhaustion, and immune evasion in the tumor microenvironment. We introduce the major immunotherapy strategies in RCC in this cycle and summarize their clinical position. Combining immune checkpoint inhibitors (ICIs) with tyrosine kinase inhibitors (TKI) has redefined the first-line standard for advanced RCC by addressing both T-cell infiltration barriers and functional suppression. Standalone approaches such as tumor vaccines and cytokines in contrast have shown limited efficacy in advanced settings. In this context, we further propose emerging research directions, such as individualized immunotherapy and multi-target blockade, and point out the relevant biomarkers, offering an integrated perspective of the RCC immune landscape and providing insights for both clinical practice and future research.

Biomedicines doi: 10.3390/biomedicines14061180

Authors: Wei Wang Jinhui Zhao Ang Li Chen Chen Weitao Jia Xiaolin Li

Background: We previously reported that knocking down the ubiquitin E3 ligase LNX2 in bone marrow monocytes by shRNAs attenuated osteoclastogenesis in vitro. However, the role of LNX2 in the regulation of osteoclasts and bone homeostasis in vivo remains unknown. Methods: In this study, we generated myeloid Lnx2 conditional knockout mice by crossing Lnx2-flox mice with LysM-Cre mice. The role of LNX2 was verified through in vitro osteoclast induction experiments using mononuclear macrophages and experiments on estrogen-deficient osteoporosis models. Results: Micro-CT and histological analysis unveiled that loss of Lnx2 in osteoclast precursor cells decreased osteoclast numbers and increased trabecular bone mass in mice. Moreover, Lnx2 deficiency prevented bone loss in an ovariectomized mouse model of postmenopausal osteoporosis. In vitro mechanistic studies identified that the loss of Lnx2 had little effect on cell proliferation but significantly inhibited the formation of osteoclasts and bone resorption. Furthermore, the deletion of Lnx2 decreased the expression of NOTCH2 and its downstream HES1 via enhancing the level of the NOTCH2 inhibitor, NUMB. Conclusions: Our findings elucidate an important role of Lnx2 in the regulation of osteoclasts and bone metabolism and indicate that Lnx2 is a potential therapeutic target for the treatment of osteoporosis.

Biomedicines doi: 10.3390/biomedicines14061179

Authors: Jingxian Wu Xueying Qin Shuting Xie Liuyan Zheng Huan Yu Huairong Wang Yalin Chen Teng Li Tao Wu Dafang Chen Yonghua Hu Yiqun Wu

Background/Objectives: Atrial fibrillation (AF) is a complex polygenic disorder; its genetic architecture remains challenging to fully elucidate. Methods: In this study, we leveraged the extensive genetic overlap between AF and a spectrum of cardiometabolic and behavioral factors—collectively defined by Life’s Essential 8 (LE8)—to advance our understanding of its etiology. Results: We first estimated significant genetic correlations between AF and all LE8 components (rg: −0.11 to 0.19) using LD score regression. We then applied conditional false discovery rate analysis and detected 970 pleiotropic loci associated with AF and at least one LE8 trait. Subsequent colocalization analysis identified 179 loci harboring shared causal variants between AF and one or more LE8 components, which were further refined into 137 distinct colocalized regions. Through region-based annotation and functional predictors, we finally prioritized 164 candidate genes from these colocalized loci, including 40 novel genes. These candidate genes were enriched in pathways related to heart development and regulation of cardiac contraction, and were also enriched among molecular targets of otological agents. Among all LE8 components, blood pressure demonstrated the most extensive shared genetic architecture with AF, supported by the strongest genetic correlation, highest pleiotropic enrichment, and the greatest number of colocalized loci with AF. Polygenic risk scores constructed from these colocalized loci demonstrated significant associations not only for AF but also for arrhythmia and heart failure. Conclusions: Our findings establish a genetic pleiotropy-informed framework that enhances the discovery of novel risk loci of AF and advances our understanding of the shared genetic architecture and potential biological mechanisms between AF and LE8 components.

Biomedicines doi: 10.3390/biomedicines14061178

Authors: Young Lee Je Hyun Seo Sung Pyo Park

Background/Objectives: To investigate the causal relationships linking body mass index (BMI) and circulating insulin-like growth factor 1 (IGF-1) levels with diabetic maculopathy risk using two-sample Mendelian randomization (MR). Methods: A two-sample MR framework was applied, utilizing genetic instruments for BMI and IGF-1 derived from the UK Biobank. Summary-level diabetic maculopathy data were obtained from the FinnGen consortium. Genome-wide significant single-nucleotide polymorphisms (SNPs, p < 5.0 × 10−8) independently associated with each exposure were employed as instrumental variables. Primary causal estimates were obtained using the inverse-variance weighted (IVW) method. Sensitivity analyses, including MR-Egger regression, weighted median methods, and the MR-Pleiotropy RESidual Sum and Outlier (MR-PRESSO), were conducted to evaluate robustness and potential pleiotropy. Results: Genetically predicted BMI was positively associated with diabetic maculopathy risk in both the IVW analysis (odds ratio [OR] = 1.16 (95% confidence interval [CI]: 1.04–1.30), p = 0.008) and MR-PRESSO (OR = 1.16 (95% CI: 1.04–1.28), p = 0.006). MR-PRESSO exhibited a significant relationship between higher IGF-1 levels and increased diabetic maculopathy risk (OR = 1.09 (95% CI: 1.01–1.18), p = 0.025), whereas the IVW method indicated only a suggestive association (OR = 1.08 (95% CI: 0.99–1.18), p = 0.087). Conclusions: The genetic evidence supports a causal role of insulin resistance-related traits in diabetic maculopathy development, with higher BMI and IGF-1 levels increasing diabetic maculopathy risk. These results underscore the potential contributory role of IGF-1 in disease pathogenesis and suggest that insulin resistance-related traits may represent preventive therapeutic targets.

Biomedicines doi: 10.3390/biomedicines14061177

Authors: Madina Khalmirzaeva Gulfiruz Urazbayeva Almagul Kurmanova Nagima Mamedalieva Gaukhar Kurmanova Damilya Salimbayeva Ainur Veliyeva Gaini Anartayeva Zhanar Kypshakbayeva Shugyla Amirtayeva Altynay Nurmakova

Background: Recurrent pregnancy loss (RPL) remains etiologically unexplained in 40–50% of cases following standard diagnostic workup. Non-criteria antiphospholipid antibodies (non-criteria aPL) are increasingly considered potential markers of seronegative obstetric antiphospholipid syndrome (APS); however, their diagnostic value in this clinical setting requires further investigation. Objective: To assess the diagnostic value of non-criteria aPL in women with RPL and to construct an exploratory immunological scoring model for diagnostic stratification. Methods: Antiphospholipid antibody detection was performed using a single-measurement semi-quantitative line immunoblot assay (Anti-Phospholipid 10 Dot, Generic Assays, Germany). Statistical analysis included χ2, Fisher’s exact test, Mann–Whitney U test, binary logistic regression, and ROC analysis. Results: Statistically significant associations with RPL were observed for anti-prothrombin antibodies (OR = 11.1; 95% CI 1.8–68.0; p = 0.022 [Haldane–Anscombe correction]), anti-annexin V (OR = 4.28; 95% CI 1.18–15.6; p = 0.023), and anti-β2GP I (OR = 3.31; 95% CI 1.18–9.28; p = 0.019). The exploratory composite immunological score demonstrated moderate discriminatory performance (AUC = 0.701; 95% CI 0.588–0.814; p = 0.005). The overall logistic regression model was statistically significant (χ2 = 8.564; p = 0.036), although none of the individual predictors retained independent significance, indicating a contribution of cumulative immunological burden rather than any single marker. Conclusions: In this single-center cross-sectional study, non-criteria aPL were frequently detected in women with RPL and were statistically associated with the condition. The findings should be interpreted as hypothesis-generating only, given the cross-sectional design, single-measurement immunoblot, small control group, and absence of external validation. Confirmation in larger prospective multicenter cohorts using ELISA-based assays with the internationally recommended 12-week repeat measurement is required before any clinical implementation.

Biomedicines doi: 10.3390/biomedicines14051176

Authors: Alexandra-Cristiana Gache Elena Danteș Ariadna-Petronela Fildan Andreea-Cristina Postu Viorica Zamfir Adina-Milena Man Nicoleta-Larisa Șerban Irene Rășanu Any Axelerad

Background/Objectives: Respiratory dysfunction in Parkinson’s disease (PD) is a clinically relevant but frequently underrecognized manifestation associated with functional impairment and increased risk of respiratory complications. This study compared spirometry and impulse oscillometry (IOS) in the assessment of respiratory function in PD, with particular focus on the detection of subtle or peripheral airway abnormalities. Methods: A prospective, single-center, cross-sectional study was conducted, including 108 participants (55 patients with PD and 53 control subjects). Pulmonary function was evaluated using standardized spirometry and IOS protocols. Group comparisons were performed using non-parametric tests, while multivariable regression analyses adjusted for potential confounding factors, including age, body mass index, smoking status, pollutant exposure, and cardiovascular comorbidities. Results: IOS identified a higher frequency of abnormal categorical findings compared with spirometry, including among subjects with normal spirometric values. Although dyspnea was more frequent in patients with PD in unadjusted analyses, multivariable regression demonstrated that PD was not an independent predictor of respiratory dysfunction. Pollutant exposure was significantly associated with abnormal IOS findings (p = 0.011). No significant differences were observed between PD and control groups regarding continuous spirometric or oscillometric parameters. Only a weak association between disease severity and FEV1 (%) was identified, whereas no significant correlations were observed for oscillometric parameters. Conclusions: IOS may provide complementary information regarding subtle or peripheral respiratory abnormalities in patients with PD. The findings suggest that respiratory alterations in this population are likely multifactorial and not independently determined by PD itself. Incorporating oscillometric assessment into respiratory evaluation may contribute to the identification of subtle respiratory mechanical alterations in patients with PD.

Biomedicines doi: 10.3390/biomedicines14051175

Authors: Wojciech Danysz Paulina Nunez-Badinez Andreas Gravius Klaus Fink Jens Nagel

Background/Objectives: Trigeminal neuralgia (TN) is a debilitating neurological condition characterized by recurrent, severe pain linked to peripheral and central sensitization within trigeminal pathways. Current pharmacologic treatments are limited by inadequate efficacy or dose-limiting side effects, and botulinum neurotoxin type A (BoNT/A) has emerged as a viable option. However, its potential use in the management of TN is hampered by methodological limitations in existing studies and a lack of pivotal clinical trials. This study investigated the efficacy, optimal treatment site, preventive utility, and duration of effect of incobotulinumtoxinA (Inco/A), a BoNT/A, in a model of TN. Methods: An infraorbital nerve chronic constriction injury model was used to induce mechanical allodynia in male Sprague–Dawley rats, reproducing the trigeminal sensitization seen in TN. The effects of subcutaneous Inco/A (1, 2, and 4 U) were measured using the mechanical sensitivity (von Frey) test to evaluate the dose response, effect of injection location, potential preventive nature of treatment, and duration of benefit. Results: Inco/A produced a robust, dose-dependent reduction in mechanical allodynia, predominantly via a local mechanism of action. Both preventive and therapeutic administration of Inco/A was efficacious, with significant reduction in allodynia even when administered up to 28 days before nerve injury. The anti-allodynic effect persisted up to 56 days post-injection. Conclusions: Inco/A is highly effective in alleviating mechanical allodynia in a validated rat model of TN. The findings highlight Inco/A as a promising candidate for clinical translation in TN and related neuropathic pain syndromes and support systematic investigation in well-controlled human trials.

Biomedicines doi: 10.3390/biomedicines14051174

Authors: Karolina Merecz Kinga Suska Olga Biniszewska Mikołaj Hirsa Aneta Wojdyło Aleksandra Tarasiuk-Zawadzka Jakub Fichna

Background: Inflammatory bowel disease (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), is a group of chronic gastrointestinal (GI) diseases with complex and multifactorial pathophysiology. The global prevalence of IBD is increasing, highlighting the need to develop new therapeutic approaches. Plant-derived extracts have recently gained prominence due to their anti-inflammatory properties. Methods: This study investigated: apricot leaves (ALE), peach leaves (PLE), black chokeberry fruit (BCHE), rosehip seeds (RSE), passion fruit seeds (PSE), and linden blossom (LBE) (all at the concentration 10–200 µg/mL) in RAW 264.7 mouse macrophages. Cytotoxicity was assessed using the neutral red uptake (NRU) assay, and anti-inflammatory activity was assessed using Griess assay in the lipopolysaccharide (LPS)-induced inflammation. Additionally, the mRNA expression levels of key inflammatory genes (interferon-γ (Ifn-γ), interleukin-6 (Il-6), nitric oxide synthase (Nos2), and tumor necrosis factor-α (Tnf-α)) were analyzed. Results: ALE and PLE exhibited minimal cytotoxicity and strong anti-inflammatory activity, reducing the expression of all analyzed genes. PSE demonstrated anti-inflammatory activity in the Griess assay, but did not alter mRNA expression. Conclusions: ALE and PLE exhibit promising anti-inflammatory properties and warrant further preclinical investigation. Comprehensive in vitro and in vivo studies are necessary to confirm these results.

Biomedicines doi: 10.3390/biomedicines14051173

Authors: Paweł Kubik Wojciech Gruszczyński Aleksandra Pawłowska Maciej Malinowski Brygida Baran Agnieszka Pawłowska-Kubik Łukasz Kodłubański Dariusz Grzanka Paulina Antosik Bartłomiej Łukasik

Background: Minimally invasive aesthetic procedures using atmospheric plasma devices are increasingly applied to improve skin laxity and age-related loss of firmness. These systems generate a localized plasma arc at the tissue surface, enabling controlled and spatially confined tissue interaction; however, quantitative histological data on the extent of plasma-induced tissue effects remain limited. Materials and Methods: This ex vivo study evaluated freshly collected porcine kidney, liver, and skeletal muscle tissues (n = 3 per tissue type). Tissue sublimation defects were produced using the Plasma IQ device under conditions representative of standard clinical use, applying two predefined settings (“LOW” and “HIGH”). Immediately after treatment, specimens were fixed in 10% neutral buffered formalin and processed into formalin-fixed paraffin-embedded (FFPE) blocks. Sections were stained with hematoxylin and eosin (H&E), and the diameter and depth of the sublimation zones were measured by light microscopy. Results: Plasma IQ exposure consistently produced well-demarcated superficial sublimation defects in all tissues. The HIGH setting increased the diameter of the sublimation zones compared with the LOW setting across all tissue types, whereas the depth differences were smaller and tissue-dependent. Lesions exhibited a characteristic flattened, cone-shaped morphology, with diameter exceeding depth. No histologically detectable collateral damage was observed beyond the immediate sublimation zone. Conclusions: Atmospheric plasma treatment induces controlled and spatially confined tissue sublimation with clearly defined histological boundaries and limited penetration depth. These findings provide quantitative histological support for the localized tissue effects of plasma-based devices and their rationale in aesthetic procedures.

Biomedicines doi: 10.3390/biomedicines14051172

Authors: Jonas Wallinder Anders Wanhainen Helena Åkerud Dick Wågsäter Martin Björck

Background/Objectives: The etiology behind sex differences in the prevalence of abdominal aortic aneurysm (AAA) can only partly be explained by environmental factors such as smoking. Genetic factors are also likely to be part of the explanation since family history is common. We hypothesized that genetic factors on AAA prevalence might be different between the sexes. Methods: This study is designed as a case–control study with 83 female AAA patients, 101 female controls, 97 male AAA patients, and 196 male controls. Single nucleotide polymorphism (SNP) analysis was performed comparing 13 different SNPs. The selection of SNPs was based on previous SNP association studies, estrogen receptors, and SNPs important to inflammation and lipid metabolism, as these processes are modulated by estrogen. Results: A multivariable logistic regression resulted in significant differences in SNP association with AAA development between men and women in two SNPs (rs2010963 and rs8113877). Significant differences were found between cases and controls, using univariate analysis, in four SNPs: rs8113877 among women, and in rs6511720, rs2010963 and rs4988300 among men. No SNPs were significantly different compared to controls in both men and women. SNP rs8113877 is located in the promotor of the MMP-9 gene. Levels of circulating MMP-9 were measured in a subgroup of the study participants: an association between MMP-9 and AAA was found, and the association between rs8113877 and MMP-9 was sex-dependent. Conclusions: Genetic variability associated with AAA differs between men and women; these differences should be accounted for in future research.

Biomedicines doi: 10.3390/biomedicines14051171

Authors: Guanglan Li Jiayi Chen Chenfeng Xu Ganyuan He Feng Yu Wei Liu Yanhua Wu Wenke Hao Wenxue Hu

Background: The immune mechanism of diabetic kidney disease (DKD) has not yet been fully elucidated. This study aimed to characterize circulating lymphocyte subsets in patients with type 2 diabetes mellitus (T2DM), with a particular focus on DKD-related immune alterations and prognosis. Methods: Circulating T cells, B cells and NK cells were identified by flow cytometry. The primary endpoint was all-cause mortality, and overall survival was defined as the time from enrollment to death from any cause or last follow-up. Associations between lymphocyte subsets, inflammatory indices and renal function parameters were analyzed. Cox regression was used to identify factors associated with overall survival in patients with DKD and in the whole T2DM cohort. A prognostic nomogram was developed in the whole T2DM cohort to estimate 1-, 2-, 3-, and 5-year overall survival (OS) probabilities. Model performance was evaluated using the concordance index (C-index), calibration curves, receiver operating characteristic (ROC) curves, and decision curve analysis (DCA). Mendelian randomization (MR) was performed as a further exploratory analysis to assess whether immune-related traits were genetically associated with DKD susceptibility, with inverse variance weighting (IVW) as the primary analytical method. Results: In total, 74 T2DM patients were divided into DKD (stage 3–4 of chronic kidney disease) and non-DKD groups. Median follow-up duration was 34.6 months. DKD patients exhibited elevated levels of NK cells, the monocyte-to-lymphocyte ratio (MLR), neutrophil-to-lymphocyte ratio (NLR), and platelet-to-lymphocyte ratio (PLR). In patients with DKD, higher PLR and serum creatinine (SCr) were associated with poorer overall survival, whereas CD4+CD25+ T cell frequency was not significant after adjustment. In the whole T2DM cohort, higher frequency of circulating CD4+CD25+ T cells were associated with improved survival (HR 0.920, 95% CI 0.858–0.986, p = 0.019), whereas elevated PLR and SCr were linked to poorer outcomes. The exploratory nomogram incorporating CD4+CD25+ T cells, PLR, and SCr, showed acceptable internal performance in this cohort. As a separate exploratory analysis, MR suggested that genetically proxied CD4 expression on activated CD4 regulatory T cells was associated with a lower risk of DKD. Conclusions: DKD was associated with higher mortality and elevated MLR-, NLR-, PLR-, and NK cell levels in patients with T2DM. In patients with DKD, PLR and SCr were associated with overall survival, supporting the prognostic relevance of systemic inflammation and renal dysfunction. Individual lymphocyte subsets were not independently associated with survival in the DKD cohort after adjustment, whereas CD4+CD25+ T cell frequency provided additional prognostic information in the whole extended T2DM cohort analysis. Further validation is warranted.

Biomedicines doi: 10.3390/biomedicines14051170

Authors: José Manuel Martinez Ana Espírito Santo Pedro Leite Ana Pinho Ana Rita Carneiro Ana Maria Oliveira Diana Ramada Rui Medeiros

Background: Early identification of bacteremia in immunosuppressed cancer patients remains difficult, especially in neutropenia. This study evaluated the diagnostic accuracy of NLR, PLR, and NLPR for identifying bacteremia and sepsis in patients undergoing blood culture episode. Methods: We conducted a retrospective diagnostic accuracy study at a tertiary oncology center between January 2023 and December 2024. All bacteremia identified were included as cases. Culture-negative episodes were subsequently sampled as controls using a frequency-matching strategy. Hematological parameters were obtained within ±24 h of first blood culture episode. Diagnostic performance was assessed using ROC curve analysis and multivariable logistic regression. Results: Of 369 screened episodes, 337 from 323 unique patients were included after excluding 31 records. NLPR showed the highest accuracy for bacteremia (AUC 0.730; 95% CI 0.671–0.788). The optimal cut-off was 0.038 (sensitivity 69.2%, specificity 72.3%) and remained consistent after excluding episodes with antibiotic therapy (AUC 0.768), corticosteroids (AUC 0.708), or growth factor use (AUC 0.718). In severe neutropenia, NLPR showed the highest accuracy (AUC 0.887; 95% CI 0.797–0.978). In multivariable analysis (n = 304), NLPR remained independently associated with bacteremia (p < 0.001), with good model discrimination (AUC 0.815; 95% CI 0.763–0.866). Diagnostic performance for sepsis was lower and not statistically significant. Conclusions: These findings suggest that NLPR may represent a simple, inexpensive, widely accessible adjunctive biomarker to support early bacteremia risk stratification in immunosuppressed cancer patients, particularly in patients with severe neutropenia. Although its overall discrimination was comparable to isolated lymphocyte count, NLPR may provide clinically relevant contextual information by integrating multiple dimensions of immune dysregulation. Further prospective multicenter validation is warranted.

Biomedicines doi: 10.3390/biomedicines14051169

Authors: Jae-Heon Kim Ah-Rim Moon Miho Song Kwang-Woo Lee Soo Min Suh Hui Ji Kim Luis Alfonso Pefianco Kevin Andrean Seongho Ryu Yun-Seob Song

Purpose: Prostate cancer is one of the most common malignancies in men, yet current prognostic methods remain suboptimal. Emerging evidence indicates that microRNAs (miRNAs) play critical roles in prostate cancer progression. This study aimed to identify miRNAs associated with adverse clinical outcomes by comparing miRNA expression profiles between prostate tumors with unfavorable versus favorable prognostic features. Materials and Methods: High-throughput next-generation sequencing (NGS) was used to analyze miRNA expression in formalin-fixed, paraffin-embedded prostate cancer tissue samples. Patients were classified into favorable or unfavorable prognosis groups based on risk stratification scores, Gleason grade group, and biochemical recurrence. Differentially expressed miRNAs were identified using a fold-change threshold ≥2 and a false discovery rate (FDR) <0.05. Predicted target genes and pathway analyses were conducted to generate candidate regulatory hypotheses rather than confirm mechanistic relationships. Results: Several miRNAs were differentially expressed according to prognostic category. miR-206 was significantly downregulated in high-risk tumors compared with low-risk tumors. High-Gleason-grade tumors showed reduced expression of miR-7704 and miR-4454, while miR-25-3p and let-7f-5p were upregulated. In patients with early biochemical recurrence, miR-7704 and miR-10400-5p were downregulated relative to those with prolonged recurrence-free survival. Target prediction analysis identified CPEB3, HAND1, PTAR1, and SPRYD4 as shared candidate targets, with CPEB3 emerging as a prioritized candidate supported by consistency in external datasets rather than a confirmed molecular target. Conclusions: Distinct miRNA expression patterns correlate with prostate cancer aggressiveness and clinical outcomes. miR-206, miR-7704, miR-4454, miR-25-3p, and let-7f-5p represent candidate prognostic biomarkers. Their shared target CPEB3 should be interpreted as a prioritized candidate for future investigation. Given the very small sample size and the lack of qRT-PCR and functional validation, these findings should be considered preliminary and hypothesis-generating, requiring validation in larger independent cohorts and experimental studies.

Biomedicines doi: 10.3390/biomedicines14051168

Authors: Jing-Hong Hu Ming-Ling Chang Tung-Jung Huang Nai-Jen Liu Jui-Hsiang Tang

Background and Aims: Type 2 diabetes mellitus is a recognized risk factor for hepatocellular carcinoma (HCC), particularly in the setting of metabolic dysfunction-associated steatotic liver disease (MASLD), chronic viral hepatitis, advanced fibrosis, and cirrhosis. Beyond hyperglycemia and insulin resistance, diabetic hepatocarcinogenesis is shaped by metabolic inflammation, lipotoxicity, oxidative stress, fibrogenic remodeling, and the cirrhosis-dysplasia-HCC continuum. Sodium-glucose cotransporter-2 inhibitors (SGLT2i) may influence several hepatometabolic pathways, but the epidemiologic evidence linking SGLT2i use to HCC risk remains heterogeneous. Methods: We conducted a systematic review and meta-analysis of observational studies evaluating SGLT2i exposure and incident HCC in adults with type 2 diabetes. PubMed, Embase, and the Cochrane Library were searched up to 15 March 2026. Adjusted time-to-event estimates were pooled using a restricted maximum likelihood (REML) random-effects model. The certainty of evidence was assessed using the GRADE framework and judged to be very low. Results: Six observational studies including 526,446 participants were included. SGLT2i exposure was associated with a lower observed risk of incident HCC (pooled HR 0.59, 95% CI 0.45–0.77), but between-study heterogeneity was substantial (I2 = 75.2%, τ2 = 0.074). The association remained directionally similar after exclusion of Huynh et al. (HR 0.61, 95% CI 0.45–0.81) and in a DPP-4 inhibitor-restricted active-comparator analysis (HR 0.60, 95% CI 0.39–0.92). However, the 95% prediction interval crossed the null (0.25–1.37), indicating that future comparable studies may plausibly show no protective association. Conclusions: SGLT2i exposure was associated with a lower observed risk of incident HCC across available observational studies. However, the certainty of evidence was judged to be very low, and substantial heterogeneity, comparator variation, mixed time-to-event estimands, residual confounding, and a prediction interval crossing the null preclude causal interpretation. These findings should be considered hypothesis-generating rather than practice-changing evidence and support further hepatology-oriented validation.

Biomedicines doi: 10.3390/biomedicines14051167

Authors: Chen Liu Xueling Wang Jiaqi Lu

Background: Although emerging evidence indicates that major depressive disorder (MDD) raises the risk of developing ovarian cancer (OC) and worsens survival, the biological mechanisms underlying this relationship remain unclear. This study explores the MDD-OC association using single-cell transcriptomics and genetic approaches. Methods: Using single-cell RNA-seq profiles of peripheral blood from MDD and OC patients, we compared shifts in immune cell subsets and selected the consistently expanded CD8+ effector memory (CD8_EM) T cells population for follow-up, validated using flow cytometry. We integrated expression quantitative trait loci (eQTL) data from CD8_EM T cell-specific genes with OC genome-wide association study (GWAS) summary statistics through two-sample Mendelian randomization (MR). In vitro experiments were additionally conducted to assess CLSTN3’s role in OC cell proliferation. Results: Among the 554 differentially expressed genes (DEGs) identified in CD8_EM T cells, MR showed a nominal association between CLSTN3 and ovarian cancer risk (OR 1.21, 95% CI 1.03–1.43), though this did not withstand correction for multiple comparisons. Colocalization analysis confirmed that CLSTN3 expression, regulated by the genetic variant rs3759416, shares a causal variant with the OC GWAS signal (PPH4 = 99.99%). Functionally, siRNA-mediated CLSTN3 silencing in HOC7 cells significantly reduced cell viability (CCK-8 assay). Conclusions: By focusing on CD8_EM T cells shared by MDD and ovarian cancer, we identified CLSTN3 as a candidate molecule through nominated by the convergence of genetic, transcriptomic, and functional evidence. These findings provide preliminary insights into the connection between depression and OC, though further validation is warranted.

Biomedicines doi: 10.3390/biomedicines14051166

Authors: Worarat Boonpech Pemikar Srifa Dhassida Sooksawat Praopim Limsakul Jirakrit Saetang Varomyalin Tipmanee Krit Charupanit Chaitong Churuangsuk Kantida Juncheed

Background/Objectives: Hepatocellular carcinoma (HCC) exhibits enhanced glycolytic activity, primarily facilitated by Class I glucose transporters (GLUTs), particularly GLUT-2. Phloretin, a natural polyphenol, is known to modulate glucose transport; however, its isoform-specific interactions and functional impact on HCC metabolism remain unclear. This study compared phloretin’s inhibitory effects on glucose uptake in HCC cells versus normal liver cell models and assessed its binding affinity across Class I GLUTs using molecular docking. Methods: Cytotoxicity was evaluated in HepG2 (HCC) and THLE-2 (normal hepatocyte) cells using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays to determine biologically relevant concentrations. Glucose uptake at sub-cytotoxic levels was quantified using the fluorescent analog 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-Deoxyglucose. To elucidate the molecular mechanism, in silico docking simulations were performed to compare the binding affinities of phloretin, glucose, and reference inhibitors (glutor and cytochalasin B) with the outward-facing conformations of GLUT-1 through GLUT-4. Results: Phloretin induced dose- and time-dependent cytotoxicity, with HepG2 cells exhibiting significantly higher sensitivity than THLE-2 cells. Functionally, phloretin markedly reduced glucose uptake in HepG2 cells, whereas THLE-2 cells showed minimal inhibition. Molecular docking revealed that phloretin occupies the central substrate-binding cavity of Class I GLUTs, forming its most stable interaction network with GLUT-2. Conclusions: These results demonstrate that phloretin selectively inhibits glucose uptake in liver cancer cells, likely through its high-affinity interaction with GLUT-2. Collectively, these findings highlight phloretin’s potential as a metabolic therapeutic agent and support GLUT-2 as a viable target for HCC intervention.

Biomedicines doi: 10.3390/biomedicines14051165

Authors: William Sosa Lucas Camargo Felipe Fregni

Background: Chronic low back pain (CLBP) is a leading cause of disability worldwide, with exercise endorsed as first-line treatment and non-steroidal anti-inflammatory drugs (NSAIDs) among the most used pharmacologic options. These interventions are frequently combined in clinical practice, yet their synergistic effects remain unclear. To evaluate whether NSAID use modifies the association between physical activity (PA) and CLBP using nationally representative data from NHANES 2009–2010. Methods: We analyzed 988 adults aged ≥20 years with complete data on chronic low back pain, physical activity, medication use, and modeled covariates. Results: Among participants not using NSAIDs, moderate recreational physical activity was associated with lower odds of CLBP (adjusted OR = 0.47, 95% CI 0.25–0.91; p = 0.029). Active transport showed a similar direction but was not statistically significant (OR = 0.38, 95% CI 0.13–1.12; p = 0.074). In interaction models, active transport x aspirin was associated with higher odds of CLBP (OR = 2.24, 95% CI 1.02–4.90; p = 0.044), and moderate recreational PA x any NSAID use was also associated with higher odds of CLBP (OR = 2.26, 95% CI 1.01–5.06; p = 0.047). Subgroup analyses were exploratory and heterogeneous, including a significant potential protective interaction (OR ≈ 0.19, 95% CI 0.06–0.69; p = 0.015). Conclusions: In a nationally representative sample, NSAID use appeared to modify the association between physical activity and chronic low back pain. These findings are exploratory and hypothesis-generating. Therefore, longitudinal studies are needed to clarify the temporal and causal relationships and the potential influence of NSAIDs.

Biomedicines doi: 10.3390/biomedicines14051164

Authors: Tamara Pawlaczyk-Kamieńska Tomasz Kulczyk

Juvenile idiopathic arthritis (JIA) is the most common systemic chronic inflammatory connective tissue disease in children, characterized by joint inflammation lasting at least six months. Temporomandibular joint (TMJ) involvement can occur in conjunction with other joints and may often be asymptomatic in its early stages. Objective: This study aims to evaluate the relationship between clinical symptoms of the stomatognathic system and radiologically confirmed cone beam computed tomography (CBCT)-detected structural TMJ changes in children with JIA. The research hypothesis posits that specific clinical symptoms are more prevalent in patients with CBCT-confirmed structural TMJ changes. Methods: A cohort of children diagnosed with JIA was examined. Clinical symptoms, including facial asymmetry, limited mandibular movement, and joint and masticatory muscle pain upon palpation, were assessed. CBCT imaging was performed to assess osseous TMJ structural changes. Results: The frequency of orofacial clinical symptoms was assessed and compared between patients with and without radiological evidence of TMJ involvement. Children with CBCT-confirmed TMJ changes demonstrated significantly higher rates of facial asymmetry, reduced maximum mouth opening, mandibular deviation during opening, and limitations in lateral or protrusive movements compared with those without TMJ involvement. Pain-related symptoms (TMJ pain, muscle tenderness, and pain during movement) and joint sounds occurred at similar frequencies in both groups. Conclusions: Facial asymmetry, mandibular deviation during opening and reduced mandibular mobility are the clinical signs most strongly associated with structural TMJ involvement in JIA and should prompt targeted imaging. Pain-related symptoms show limited diagnostic value, highlighting the need for focused clinical assessment and future studies integrating CBCT and MRI to refine early screening protocols.

Biomedicines doi: 10.3390/biomedicines14051163

Authors: Ruxandra Ștefănescu Amelia Tero-Vescan Camil-Eugen Vari Dragoș Sita Bianca-Eugenia Ősz

Background: Periodontitis is a chronic inflammatory disease characterized by dysbiotic biofilm formation, progressive destruction of periodontal tissues, and alveolar bone resorption. Conventional periodontal therapy primarily focuses on mechanical biofilm removal; however, adjunctive therapeutic approaches targeting host inflammatory responses and microbial activity have gained increasing attention. Cannabidiol (CBD), a non-psychoactive phytocannabinoid derived from Cannabis sativa, has demonstrated anti-inflammatory, antimicrobial, and immunomodulatory properties that may be relevant in periodontal disease management. Objective: This systematic review aimed to evaluate the available evidence regarding the potential role of CBD in modulating periodontal inflammation, microbial biofilms, and bone resorption processes. Methods: A systematic literature search was conducted in Web of Science, Cochrane, PubMed, Scopus, and Google Scholar. The review was conducted in accordance with PRISMA guidelines. Studies investigating the effects of CBD on periodontal inflammation, oral biofilms, or bone remodeling were included. Both preclinical (in vitro and animal) and clinical studies were considered. Results: Evidence from experimental studies consistently demonstrated that CBD modulates inflammatory signaling pathways, including inhibition of the TLR4/NF-κB pathway and a reduction in pro-inflammatory cytokine expression, but some results are contradictory. Animal studies reported reduced alveolar bone loss and decreased osteoclast activity following CBD administration. Several studies also demonstrated antimicrobial and antibiofilm effects of CBD against oral microorganisms. Conclusions: While preclinical evidence is promising, the current body of clinical data remains limited. Further well-designed randomized clinical trials are required to determine the efficacy, type of formulation, optimal dosing, and long-term safety of CBD as an adjunctive therapy in periodontal treatment.

Biomedicines doi: 10.3390/biomedicines14051161

Authors: Anastasia Voznesenskaya Alyona Sorokina Marina Shestakova Ekaterina Shestakova Ildar Minniakhmetov Anna Ivanova Sergey Rumyantsev Natalia Mokrysheva Vladimir Chekhonin Marina Loguinova

Of the many clinical phenotypes of obesity, the most prevalent are metabolically “healthy” (MHO) and metabolically “unhealthy” (MUO) obesities, the latter being associated with a range of comorbidities, including type 2 diabetes mellitus (T2DM). The underlying causes of different obesity phenotypes and the mechanisms of conversion of one phenotype into another have yet to be fully elucidated. However, increasing evidence suggests the key role of low-grade metabolic inflammation (metaflammation) in the pathogenesis of obesity and metabolic dysfunction. The review presents a comprehensive description of changes in immune cell populations and pro-inflammatory mediators, as well as a detailed comparative mapping of the adipose tissue immune landscape during MHO/MUO transition. Based upon a conceptual model for the intensification of metaflammation during MHO progression and conversion to MUO, a pattern of dynamical changes that accompany MHO/MUO transition is described. Though many parameters demonstrate significant differences in multiple cross-sectional and some longitudinal studies, only a few of them (CRP, IL-6, IL-17A, absolute counts of leukocytes and neutrophils) meet the criteria of a validated biomarker in clinical setting. A lack of standardization in MHO definition and heterogeneity in the severity of MUO make the search for predictive biomarkers a challenge. The review also discusses the mechanisms underlying metabolic memory and the incomplete reversibility of metabolic disturbances after bariatric surgery.

Biomedicines doi: 10.3390/biomedicines14051162

Authors: Zhiqiang Shen Linlin Ma George D. Mellick

Background: Parkinson’s disease (PD) has become the fastest-growing neurodegenerative disorder worldwide. A valuable approach for unraveling the disease’s mechanisms and new therapeutic targets involves investigating the PD-causing genes identified in families exhibiting the Mendelian inheritance of parkinsonism. Methods: In this article, we review how genetically modified mouse models can be employed to decipher the genetic architecture of PD. Results: We first discuss how well the human motor and non-motor symptoms of PD are currently evaluated in these PD mouse models, highlighting limitations. The pathogenic roles of five inherited PARK genes in PD are then extensively examined through their respective genetic mouse models in terms of phenotypic and cellular impacts. Furthermore, we discuss the strengths and weaknesses of existing transgenic mouse models and highlight significant accomplishments and advancements in this field from 2018 to the present. Conclusions: Building upon the current understanding of PD, we propose potential directions for enhancing genetic mouse models to further unveil the underlying mechanisms of PD and advance therapeutic research.

Biomedicines doi: 10.3390/biomedicines14051160

Authors: Diana-Ruxandra Hădăreanu Flavia-Mihaela Stoiculescu Călin-Dinu Hădăreanu Maria-Livia Iovănescu Anca Mihu-Marinescu Georgică-Costinel Târtea Ionuț Donoiu Oana Munteanu-Mirea Răzvan-Ilie Radu Eugen-Nicolae Țieranu Octavian Istrătoaie Cristina Florescu

Background/Objectives: Atrial fibrillation (AF) is common in heart failure with preserved ejection fraction (HFpEF) and is associated with worse symptoms and prognosis. Emerging evidence suggests that sex modifies the AF–HFpEF relationship through differences in atrial remodeling, comorbidity burden, and hemodynamic vulnerability. This study aimed to evaluate how sex and AF jointly relate to differences in cardiac structure, clinical characteristics, and outcomes in HFpEF. Methods: We retrospectively analyzed 622 patients with HFpEF admitted between January 2019 and May 2023. Patients were categorized into four predefined clinical subgroups: women without AF, women with AF, men without AF, and men with AF. The primary endpoint was first rehospitalization for HF decompensation. Results: Over a mean follow-up of 48.6 ± 16.4 months, 181 patients (29.1%) were rehospitalized for worsening HF, with the highest event burden observed in men with AF. Sex and AF were each associated with distinct clinical and remodeling profiles, without significant sex-by-AF interaction effects. AF was independently associated with a higher risk of HF rehospitalization (HR 1.45, 95% CI 1.06–1.99, p = 0.021), whereas female sex was protective (HR 0.71, 95% CI 0.53–0.97, p = 0.032). Men with AF exhibited the most adverse remodeling profile, characterized by the largest unindexed left atrial and left ventricular dimensions, the highest prevalence of significant tricuspid regurgitation, and the lowest event-free survival (HR 1.92, 95% CI 1.23–2.99, p = 0.004). In contrast, women with AF more frequently displayed concentric remodeling and significant mitral regurgitation. Independent predictors of rehospitalization included higher NYHA functional class and lower left ventricular EF within the preserved EF range. Conclusions: Sex and AF were independently associated with substantial differences in cardiac structure, clinical characteristics and prognosis in HFpEF. Men with AF represent the highest-risk subgroup, driven by more advanced structural remodeling and valvular dysfunction. These findings suggest that simple sex- and rhythm-based classification may provide complementary information for risk stratification and management in HFpEF. Further validation in independent cohorts is warranted.

Biomedicines doi: 10.3390/biomedicines14051159

Authors: Michele Di Crosta Francesca Chiara Ragone Rossella Benedetti Gabriella D’Orazi Roberta Santarelli Maria Saveria Gilardini Montani Mara Cirone

In the original publication [...]

Biomedicines doi: 10.3390/biomedicines14051158

Authors: Lyubov V. Machekhina Alexandra A. Melnitskaya Mikhail S. Arbatskiy Anna V. Permyakova Alexey V. Churov Irina D. Strazhesko Olga N. Tkacheva

Background/Objectives: Population aging necessitates a shift from disease-focused paradigms to a holistic characterization of biological aging processes. While chronological age remains the primary metric, it poorly captures inter-individual variability in physiological resilience and health trajectories. This study aimed to identify robust, multidimensional aging phenotypes independent of chronological age and sex using integrative factor analysis of heterogeneous biomedical data from a Russian cohort—a population underrepresented in aging research. Methods: We analyzed data from 1201 conditionally healthy adults (aged 18–99 years) enrolled in the RUSS AGE study. A comprehensive dataset comprising 118 variables across 11 modalities—including biochemical markers, anthropometry, physical function, cognitive-emotional assessments, lifestyle factors, and psychosocial indicators—was integrated using Multi-Omics Factor Analysis v2 (MOFA2). Following the extraction of 16 latent factors and residualization for demographic confounders, consensus clustering was performed to identify distinct aging phenotypes. Phenotype stability was internally recapitulated using gradient-boosting classifiers (XGBoost, CatBoost) in a stratified five-fold cross-validation and on a held-out test set. Results: MOFA2 identified 16 stable latent factors, explaining 21.3% of the total variance and capturing coordinated variation across metabolic, inflammatory, cardiovascular, cognitive, and behavioral domains. Consensus clustering revealed five reproducible phenotypes—Anemic (n = 82), Metabolically Subcompensated (n = 99), Metabolically Decompensated (n = 304), Overloaded (n = 302), and Balanced (n = 414)—characterized by distinct multisystem profiles independent of age (p > 0.05 after FDR correction) and sex. Supervised classification achieved high discriminative performance (macro F1-score = 0.75, OvR ROC-AUC = 0.93 on the held-out test set), quantifying the internal reconstructability of the phenotype labels from the original feature space rather than external generalization to an independent cohort. Conclusions: This study demonstrates the feasibility of data-driven, biologically coherent phenotyping of healthy aging using integrative factor analysis. The identified phenotypes represent stable configurations of physiological, functional, and psychosocial characteristics that transcend chronological age, providing a foundation for the future development of risk-stratification tools, preventive interventions, and biological-age calculators, subject to subsequent validation in longitudinal and independent external cohorts.

Biomedicines doi: 10.3390/biomedicines14051157

Authors: Cristina Iosif Anca Labunet Andreea Kui Stanca Cuc Marioara Moldovan Sorina Sava

Background: Durable adhesion between orthodontic brackets and enamel is essential for successful fixed orthodontic therapy. Despite simplified adhesive systems being available, conventional etch-and-rinse adhesives remain widely used due to their reliable enamel bonding. Methods: This in vitro study evaluated the bonding performance of three orthodontic adhesive strategies in combination with Transbond XT composite resin for metal bracket fixation. Thirty extracted human premolars were randomly allocated to three groups according to the adhesive system applied: OptiBond Solo Plus (etch-and-rinse method), SafeBond Universal DC (selective enamel etching method) and Transbond XT primer (control method). Shear adhesion resistance, maximum force and breakout force were measured and statistically analysed. Results: No statistically significant differences were observed between the OptiBond and SafeBond groups for any of the evaluated mechanical parameters (p > 0.05), although the OptiBond group exhibited higher mean values. The Transbond XT primer group showed significantly lower adhesion resistance and debonding forces than both of the other groups (p < 0.05). SafeBond demonstrated lower variability of results compared with OptiBond. Conclusions: When used with Transbond XT composite resin, both OptiBond Solo Plus and SafeBond Universal DC provided comparable mechanical performance for metal bracket bonding. While OptiBond yielded higher mean bond strength values, SafeBond exhibited more consistent behaviour. The Transbond XT primer alone resulted in inferior bonding performance.