Search Results (8,313)

Background/Objectives: HER2, a critical diagnostic marker and therapeutic target in breast cancer, is a membrane receptor that forms diverse signaling complexes, the constituents of which have not been fully characterized in actual breast cancer tissues. Methods: In this study, we applied the Rapid Immunoprecipitation Mass Spectrometry of Endogenous Proteins (RIME) method, originally developed to explore transcription factor complexes, to identify the complexes formed by HER2 in HER2-positive breast cancer specimens. Results: Through our approach, we successfully identified multiple complex components, including MARCKS, a novel HER2-interacting partner, which we verified using both proximal ligation assay in cultured cells and immunohistochemistry in tissue sections. TCGA analysis further revealed that high MARCKS expression significantly correlates with ER negativity, as confirmed by multivariate analysis, suggesting its potential role as a prognostic indicator in aggressive breast cancer subtypes. Conclusions: These results demonstrate the capability of RIME to elucidate interactomes of membrane proteins such as HER2 in clinical tissue specimens. Furthermore, this study highlights its broader applicability beyond nuclear proteins, underscoring its potential for discovering novel prognostic and diagnostic clinical markers in diverse cancer types. Full article

Drosophila melanogaster has been a cornerstone of biological research, offering critical insights into genetics, neurobiology, and disease modelling. This review examines Drosophila feeding research, including the diverse assays available to study feeding behaviour, and explores its biomedical and entomological applications. We highlight studies that have advanced our understanding of human feeding and eating disorders, vector-borne infectious diseases, and agricultural pest control. In clinical applications, we discuss a two-pronged approach: using Drosophila to model human feeding and eating disorders, as well as to study insect vectors that contribute to human disease transmission. We explore how feeding studies in Drosophila provide valuable insights into energy homeostasis, metabolic regulation, pathogen–host interactions, and vector biology. Beyond clinical relevance, the entomological applications of Drosophila feeding research extend to sustainable pest management and insecticide resistance. Finally, we identify gaps in current research and suggest promising directions for further exploration. By leveraging the genetic and behavioural tools available in this model, researchers can continue to uncover conserved mechanisms with broad implications for human health, disease control, and agricultural sustainability. Full article

Autism spectrum disorder (ASD) is a heterogeneous neurodevelopmental condition often associated with genetic syndromes. Structural variants on the long arm of chromosome 15 (15q) are recurrently implicated in syndromic ASD, yet their phenotypic spectrum remains insufficiently characterized in diverse populations. We retrospectively analyzed clinical and molecular data from three patients with ASD treated at a Brazilian public reference center who also presented neurological and systemic comorbidities. Genetic investigations included G-banded karyotyping, chromosomal microarray analysis (CMA), methylation assays, and multiplex ligation-dependent probe amplification (MLPA) when indicated. Variants were classified according to ACMG guidelines and correlated with individual phenotypes. Case 1 showed an 8.4 Mb triplication at 15q11.2–q13.1 encompassing SNRPN, UBE3A, and GABRB3, which are associated with epilepsy, delayed neuropsychomotor development, and dysmorphic traits. Case 2 presented a 418 kb duplication at 15q13.3 involving CHRNA7 and OTUD7A, a variant of uncertain significance correlated with intellectual disability, speech apraxia, and self-injurious behavior. Case 3 demonstrated extensive loss of heterozygosity at 15q11.2–q13.1 and 15q21.3–q26.2, which is compatible with maternal uniparental disomy and Prader–Willi syndrome, manifesting hypotonia, seizures, and global delay. These findings underscore the potential involvement of the 15q region in syndromic ASD and related neurological comorbidities, highlighting the diverse pathogenic mechanisms and the importance of comprehensive genomic profiling for diagnosis, counseling, and individualized care. Full article

Background: Q fever is a zoonotic disease caused by the intracellular bacterium Coxiella (C.) burnetii. In ruminants, it mainly leads to reproductive disorders. In humans, transmission typically occurs through direct contact with infected animals or inhalation of contaminated aerosols. Although it is a notifiable disease in the European Union for both humans and certain animal species, the actual incidence is likely underestimated due to the non-specific nature of clinical symptoms. Domestic ruminants are considered the main reservoirs of C. burnetii, placing farmers and veterinarians at increased occupational risk of infection. Objectives: This study aimed to assess the risk of Q fever infection in northern Italy by comparing the seroprevalence rates between professionally exposed individuals and not professionally exposed people. Methods: A total of 209 serum samples were analysed: 117 from exposed professionals (veterinarians, biologists, agronomists, laboratory technicians) and 92 from professionally unexposed people (control group). Serum samples were tested with a commercial enzyme-linked immunosorbent assay to detect the presence of IgG against C. burnetii. Positive and doubtful samples were further investigated with a commercial immunofluorescence assay for detection of IgM and IgG. Epidemiological data were also collected to explore potential risk factors. Results: In total, 10 of the 117 exposed individuals tested positive, yielding a seroprevalence of 8.6%, while only 1 of the 92 control subjects tested positive (1.1%). These findings indicate a significantly higher occupational risk of C. burnetii infection among exposed professionals compared to the general population. Conclusions: The results highlight the need for preventive measures and surveillance in at-risk occupational groups. Full article

Quality assurance in a clinical laboratory is essential to ensure reliable, accurate and precise laboratory test results all the time. A hemostasis laboratory is an important part of a clinical laboratory setting in a hospital or a healthcare center, and clinical laboratory tests play a crucial role in diagnosis and management of conditions related to bleeding or clotting of diseased individuals. This review discusses all aspects of coagulation laboratory testing from pre-analytical, analytical, and post-analytical variables as part of daily quality assurance processes undertaken as well as the quality management process of assay validation and implementation in a laboratory prior to patient testing. The internal and external quality processes that drive a hemostasis laboratory will be discussed that shows a rigorous process in assurance of testing that is reliable and accurate every time, at all times. Full article

Newcastle disease (ND) is a highly contagious and economically significant viral infection that affects poultry globally, with recurrent outbreaks occurring even among vaccinated flocks in Egypt. Caused by the Newcastle disease virus (NDV), the disease results in substantial losses due to high mortality rates, decreased productivity, and the imposition of trade restrictions. This study aimed to develop a rapid, sensitive, and field-deployable diagnostic assay based on real-time reverse transcription recombinase-aided amplification (RT-RAA) for the detection of all NDV genotypes in clinical avian specimens. Primers and an exo-probe were designed based on the most conserved region of the NDV matrix gene. After testing ten primer combinations, the pair NDV RAA-F1 and RAA-R5 demonstrated the highest sensitivity, detecting as low as 6.89 EID₅₀/mL (95% CI). The RT-RAA assay showed excellent clinical sensitivity and specificity, with no cross-reactivity to other common respiratory pathogens such as avian influenza virus, infectious bronchitis virus, Mycoplasma gallisepticum or infectious laryngotracheitis virus. All 25 field samples that were tested positive by real-time RT-PCR, including those with high CT values (~35), were detected by RT-RAA in 2–11 min, indicating superior sensitivity and speed. The assay requires only basic equipment and can be performed under isothermal conditions, making it highly suitable for on-site detection in resource-limited or rural settings. The successful implementation of RT-RAA can improve NDV outbreak response, support timely vaccination strategies, and enhance disease control efforts. Overall, the assay presents a promising alternative to conventional diagnostic methods, contributing to the sustainability and productivity of the poultry sector in endemic regions. Full article

Background: Multiple sclerosis (MS) is an immune-mediated neuroinflammatory disorder with a multifactorial etiology involving genetic susceptibility, environmental triggers, and vascular contributions. Carotid intima–media thickness (CIMT) is a significant marker of endothelial dysfunction. Endothelial cell-specific molecule-1 (endocan) and hyaluronic acid, key components implicated in endothelial and vascular remodeling, may significantly contribute to the inflammatory and vascular pathologies observed in MS. We aimed to investigate the relationship between CIMT and endothelial biomarkers, such as endocan and hyaluronic acid, in patients with MS. Methods: In this cross-sectional study, 100 patients with relapsing–remitting MS and 56 healthy controls were included. Demographic, clinical, laboratory, and imaging data were documented. CIMT was measured bilaterally using high-resolution B-mode ultrasonography. Serum endocan and hyaluronic acid levels were quantified via enzyme-linked immunosorbent assays. Results: MS patients exhibited significantly higher CIMT and serum endocan levels compared with controls (p < 0.001). CIMT values were significantly elevated in MS patients, with longer disease duration, higher expanded disability status scale scores, and an older diagnosis age (p < 0.05). However, serum endocan and hyaluronic acid levels did not significantly differ between MS subgroups based on disease duration, disability severity, and diagnosis age. Additionally, there was no correlation between CIMT and serum endocan and hyaluronic acid levels in MS patients (p > 0.05). Conclusions: Increased CIMT and serum endocan levels in MS patients may indicate endothelial dysfunction suggesting vascular involvement in MS. The lack of a correlation between CIMT and endocan and hyaluronic acid levels reveals the complexity of vascular and immune interactions in MS, which needs further research. Full article

Esophageal squamous cell carcinoma (ESCC) accounts for the majority of esophageal cancers worldwide, with a poor prognosis and increasing resistance to conventional treatments. Faced with these limitations, nanoparticles (NPs) are attracting growing interest as innovative therapeutic agents capable of improving specificity and efficacy and reducing systemic toxicity. This study critically examines the pharmacological effects, mechanisms of action, and toxicity profiles of different metallic or organic nanoparticles tested on ESCC cell lines. Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) 2020 guidelines were followed by a meticulous literature search of Google Scholar, Web of Science, PubMed/Medline, and Scopus databases to achieve this goal. The results show that the anti-tumor properties vary according to the type of nanoparticle (copper(II) oxide (CuO), silver (Ag), gold (Au), nickel(II) oxide (NiO), nano-curcumin, etc.), the synthesis method (chemical vs. green), and the biological activity assessment method (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Bromodeoxyuridine (BrdU), Cell Counting Kit-8 (CCK8) assays, etc.). NPs derived from green synthesis, such as those based on Moringa oleifera, Photinia glabra, or pomegranate bark, exhibit moderate cytotoxic activity (50% inhibitory concentration (IC₅₀) between 92 and 500 µg/mL) but show good tolerance on normal cells. In contrast, chemically synthesized NPs, such as Cu(II) complexes with 1,3,5-benzenetricarboxylic acid (H₃btc) or 1,2,4-triazole (Htrz), show lower IC₅₀ (34–86 µM), indicating more marked cytotoxicity towards cancer cells, although data on their toxicity are sometimes lacking. In addition, multifunctional nanoparticles, such as gold-based nano-conjugates targeting Cluster of Differentiation 271 (CD271) or systems combined with doxorubicin, show remarkable activity with IC₅₀ below 3 µM and enhanced tumor selectivity, positioning them among the most promising candidates for future clinical application against ESCC. The most frequently observed mechanisms of action include induction of apoptosis (↑caspases, ↑p53, ↓Bcl-2), oxidative stress, and inhibition of proliferation. In conclusion, this work identifies several promising nanoparticles (silver nanoparticles derived from Photinia glabra (PG), gold-based nano-immunoconjugates targeting CD271, and silver–doxorubicin complexes) for future pharmaceutical exploitation against ESCC. However, major limitations remain, such as the lack of methodological standardization, insufficient in vivo and clinical studies, and poor industrial transposability. Future prospects include the development of multifunctional nanocomposites, the integration of biomarkers for personalized targeting, and long-term toxicological assessment. Full article

Background/Objectives: To compare clinical features of patients with joint hypermobility syndrome/hypermobile Ehlers Danlos Syndrome (JHS/hEDS) who tested positive or negative for anti-nuclear antibodies (ANA), and to determine antibody titers, staining patterns, and reactivity to common nuclear autoantigens. Methods: ANA results were determined by Hep2 immunofluorescence assay. Reactivity to the most common nuclear autoantigens was measured by the Multiplex assay. Clinical manifestations were compared between three subgroups: total ANA+, ANA+ who did not have evidence of systemic autoimmune inflammatory disease (SAID), and ANA−. Results: Of 289 patients, 210 patients had a Beighton score > 5 and were tested for ANA antibodies. One hundred and thirty-one patients had a positive ANA test. Twenty patients in this subgroup were classified as SAID+ while the remaining 111 patients did not meet criteria for any systemic disease. Speckled staining was the most observed pattern in both ANA+SAID+ (75.00%) and ANA+SAID− (72.97%) subgroups. In the latter subgroup, the target of nuclear autoreactivity remained elusive in 80% of patients. The most common clinical manifestations were diffuse arthralgias, myofascial pain, sicca symptoms, Raynaud’s phenomenon, gastrointestinal manifestations, and chronic fatigue. Joint dislocations were observed more commonly in the ANA− subgroup compared to ANA+SAID− patients (30.38% vs. 12.61%, adjusted p < 0.05). Conclusions: Similar clinical characteristics were observed in ANA+ and ANA− subgroups of JHS/hEDS, except for joint dislocations which were more common in the ANA− subgroup. The target of ANA reactivity was unknown in 80% of ANA+JHS/hEDS patients and needs to be determined in future studies. Full article

Bacterial-mediated infections represent a major risk factor for chronic wounds. Numerous polymeric dressings have been proposed to reduce this incidence and promote wound healing. In the present investigation, chitosan/collagen/honey-based sponges were prepared by freeze-drying. The effect of honey incorporation at different concentrations on the physicochemical and antibacterial properties of the sponges was evaluated. The SEM images showed that the surface and cross-sections of all samples had a porous structure. The pore size gradually increased in the range of 78.14 to 126.9 μm due to the increase in honey content in the sponges. This property resulted in considerably higher porosity degrees (79.90–90.13%) and absorption rates (ranges of 1357–1665% in deionized water and 865–1938% in PBS solution) in honey-loaded systems. Conversely, the honey composite formulations exhibited a reduction in permeability, with WVTR values ranging from 131.01 to 99.39 gh⁻¹m⁻² and values of WVP from 0.3255 to 0.2118 gm⁻¹d⁻¹mm Hg⁻¹. The mechanical properties showed that adding honey made the sponges more flexible (12.49–7.95% MPa) but decreased elongation rates in the sponges (16.36–7.56%) due to higher pore heterogeneity. The antibacterial tests indicated that all treatments had inhibitory effects against S. aureus, P. aeruginosa, E. coli, and L. monocytogenes. The results from cells viability assays and in vitro healing models using human keratinocytes demonstrate that chitosan/collagen/honey sponges represent a potential alternative for applications such as wound dressings to help treat skin ulcers. The physicochemical, antibacterial, and biocompatibility properties of chitosan/collagen/honey sponges indicated their potential as a promising alternative for clinical use. Full article

Background: Neuroblastoma is a childhood tumor that is usually fatal after relapse in high-risk patients. Most clinical trials of CAR-T therapy for neuroblastoma are based on targeting the disialoganglioside GD2. B7-H3, a protein from the immunoglobulin superfamily, is a specific marker for neuroblastoma and a number of other solid tumors. We conducted a preclinical study of three variants of anti-B7-H3 CAR-T cells in order to justify the selection of the best candidate for subsequent clinical trials. Methods: The expression level of B7-H3 was measured in a number of cell lines and neuroblastoma tissue samples via flow cytometry. The functional activity of CAR-T cells was compared using an NFAT-inducible reporter assay, a cytotoxicity test, cytokine production, and a repeated stimulation assay. Results: The obtained CAR-T cells carrying all resulting CAR variants specifically recognized and killed B7-H3-positive tumor cells in vitro. Nevertheless, TE9-28z and 8H9-28BBz demonstrated superior activation and cytokine production compared to the second-generation 8H9-BBz construct. TE9-28z and 8H9-28BBz exhibited functional differences in expansion, exhaustion markers, and cytokine secretion in co-cultures with target cells in vitro. In particular, TE9-28z induced higher IFNγ production, while 8H9-28BBz showed increased TNFα release. Despite comparable cytotoxicity, TE9-28z and 8H9-28BBz CAR-T cells exhibited varying persistence depending on the tumor type, and showed signs of functional exhaustion upon prolonged exposure to the target antigen. Conclusion: TE9-28z and 8H9-28BBz were selected for further preclinical development as promising candidates for the effective targeting of B7-H3-expressing malignancies. Full article

Background/Objectives: We sought to evaluate the clinical predictors of underlying histologic activity in patients with lupus nephritis (LN), with a focus on urinary soluble protein CD163 (usCD163). Methods: We conducted a retrospective, cross-sectional study of forty-two consecutive LN patients with concurrent determination of usCD163 at the moment of kidney biopsy. A first morning void prior to the kidney biopsy was collected and usCD163 was measured by a commercial ELISA assay (EUROIMMUN, Lubeck, DE). Results: The study cohort had a median age at the moment of kidney biopsy of 33.5 (IQR: 24–42.7) years. The mean eGFR and median 24 h proteinuria were 76.6 ± 33.9 mL/min/1.73 m² and 1.98 (IQR: 0.83–4.52) g/day. The median activity (AI) and chronicity (CI) indices were 7 (IQR: 3–11) and 3 (IQR: 1–5), respectively. usCD163 significantly correlated with 24 h proteinuria (r = 0.7, p < 0.001), hematuria (r = 0.51, p < 0.001), and serum complement levels, C3 (r = −0.5, p = 0.001) and C4 (r = −0.32, p = 0.03), but not with eGFR (r = −0.23, p = 0.14). Regarding the histological parameters, usCD163 significantly correlated with the AI and the individual active lesions (except for fibrinoid necrosis), but not with CI or any chronic lesion. usCD163 had a higher AUC compared to the classical measures of renal involvement (proteinuria, hematuria, eGFR) for discriminating an elevated AI, but the differences between AUC reached statistical significance only for hematuria. Thus, the AUC of usCD163 was 0.74 (95%CI, 0.58–0.86) for an AI over 2, an AUC of 0.77 (95%CI, 0.61–0.88) for an AI over 3 and an AUC of 0.74 (95%CI, 0.57–0.86) for an AI of at least 9. The optimal cutoff value for usCD163 identified for all AI thresholds evaluated was 296.2 ng/mmol. Conclusions: usCD163 correlates with glomerular inflammation, being able to discriminate histologic activity from chronicity in patients with LN and identify minimal histologic activity, although it did not significantly outperform proteinuria. Full article

Background/Objectives: This study evaluated a novel PET tracer, ⁶⁸Ga-NOTA-CYT-200, which targets human granzyme B (GZB) as a biomarker for cytotoxic T-cell activation in a clinically relevant model of melanoma-bearing mice with a humanized immune system treated with immune checkpoint inhibitor (ICI) therapy. Methods: The binding affinity of the tracer was determined using an enzymatic colorimetric assay. Tumor-bearing humanized NSG mice underwent PET imaging before and during ICI monotherapy or combination therapy to assess ⁶⁸Ga-NOTA-CYT-200 uptake within tumors and other organs. The tumor growth was carefully monitored. The treatment response was evaluated based on the percentage change in tumor size at days 5 and 15 after the treatment started. A tracer biodistribution study and immunohistochemical staining of the tumors and organs were also performed. Results: The inhibition constant (Ki) of ⁶⁸Ga-NOTA-CYT-200 was estimated at 4.2 nM. PET imaging showed a significantly higher ⁶⁸Ga-NOTA-CYT-200 uptake in mice receiving the combination therapy compared to those receiving monotherapy or a vehicle (p < 0.0001 or p = 0.0005, respectively), which correlated with the greatest reduction in tumor size in the combination ICI group. Regardless of treatment, the responders presented with a significantly higher ⁶⁸Ga-NOTA-CYT-200 uptake at days 4 or 7 after the treatment began (p = 0.0002 and p = 0.0109, respectively). An increased uptake of ⁶⁸Ga-NOTA-CYT-200, especially in the intestines and liver within the combination ICI group, suggested immune-related adverse events (IrAEs). Conclusions: Our study demonstrates that ⁶⁸Ga-NOTA-CYT-200 PET imaging can predict the early treatment response in melanoma models treated with ICI and may also help in detecting IrAEs. Full article

Background: Oral cancer remains a significant global health concern due to its high incidence and mortality, as highlighted by GLOBOCAN 2022, and is characterized by poor survival rates despite available therapies. Therefore, there is an imperative need for developing novel therapeutic targets for this disease. Methods: This study investigates the oncogenic role of mortalin in oral cancer. We have used The Cancer Genome Atlas (TCGA) dataset, samples from North Eastern Region of India and tissue microarray to examine the expression of this gene/protein in patient samples. siRNA related knock down studies were carried out to determine the role of mortalin on oral cancer cell proliferation, survival, metastases, EMT, autophagy etc. Results: Analysis of TCGA dataset revealed increased mortalin expression in head and neck squamous cell carcinoma (HNSCC), which correlated with tumor grade and stage, and was associated with diminished overall survival. These findings were validated in oral cancer patient tissue samples obtained from the North East Region of India and oral cancer cell lines. Functional assays showed that mortalin knockdown via siRNA reduced cancer cell proliferation, migration, invasion, and angiogenesis while inducing apoptosis, disrupting mitochondrial membrane potential, and modulating autophagy. These effects were linked to altered expression of regulatory molecules, including p53, p21WAF1, cyclins, caspases, MMPs, Survivin, and components of the Akt/mTOR pathway, thereby alleviating key hallmarks of oral cancer. Conclusion: Collectively, these data support mortalin as a potential therapeutic target for oral cancer and warrant further studies for the development of mortalin-targeting drugs in both laboratory and clinical settings. Full article

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a widespread enzymopathy affecting approximately 500 million individuals that represents a significant global health issue. Among the more than 230 identified mutations in the G6PD gene, six class A variants—G6PD Utrecht (Pro409Ser), G6PD Suwalki (Pro409Arg), G6PD Merlo (Pro409Gln), G6PD Kawasaki (Gly410Ala), G6PD Shinagawa (Gly410Asp), and G6PD Riverside (Gly410Cys)—are located in the beta-loop near the NADP⁺ binding site. These mutations are of particular interest due to their association with severe hematologic phenotypes, including chronic hemolytic anemia, as well as their proposed role in the allosteric regulation of G6PD multimerization. This study presents a comprehensive biochemical and functional characterization of these clinically relevant G6PD variants. The variant enzymes were cloned, expressed, and purified for characterization. Kinetic parameters and thermal stability assays, complemented by molecular dynamics simulations (MDS), were employed to elucidate the structural impacts of the mutations. Our results demonstrate that these mutations significantly impair protein function, characterized by reduced affinity for glucose-6-phosphate (G6P) and NADP⁺, as well as altered thermal stability compared with wild-type G6PD. MDS revealed that point mutations in the βN- and βM-sheets in the NADP⁺_s region propagate subtle conformational changes, ultimately affecting the NADP⁺c region and the G6P binding cavity. Furthermore, secondary structure element analyses of the simulation data showed that Pro409 and Gly410 point mutations propagate several changes around residues 195–210 (G6P binding site) and 380–400 (NADP⁺_s), explaining their effect on overall catalytic performance. These findings enhance our understanding of the molecular mechanisms underlying G6PD deficiency and its clinical implications, providing a foundation for future therapeutic strategies aimed at mitigating the effects of these variants. Full article