Cells doi: 10.3390/cells15110974

Authors: Menachem Hanani Rachel Feldman-Goriachnik Suhail Aamar

Fibromyalgia (FM) is a complex syndrome associated with chronic widespread pain and with various other symptoms, including sleep and mood disturbances. Its underlying causes are not fully understood, and the lack of diagnostic blood tests and imaging, along with the absence of definitive treatments, makes management challenging. Recent studies showed that passive transfer of immunoglobulins from FM patients into mice activated satellite glial cells (SGCs) in mouse dorsal root ganglia (DRG), leading to pain behaviors. Here, we aimed to determine whether whole serum from FM patients activates mouse SGCs in DRGs and other ganglia that may be involved in FM’s diverse symptoms. Serum from FM patients (N = 15) and healthy controls (HCs, N = 8) was collected. Sera were incubated with different types of mouse sensory ganglia: DRG, trigeminal ganglion (TG), the nodose ganglion (NG), and the superior cervical sympathetic ganglion (Sup-CG). SGC activation was assessed by immunostaining of SGCs for the glial activation marker glial fibrillary acidic protein (GFAP). All the ganglia tested, DRG, TG, NG, and Sup-CG, displayed induced upregulation of GFAP labeling in SGCs after incubation with FM serum compared with HCs, indicating SGC activation by the serum. Similar responses were observed in both male and female mice. We conclude that serum from FM patients contains factors that can activate SGCs across various types of mouse ganglia, which may reflect the diverse symptom profile of FM. These findings provide evidence for pathogenic factors that could serve as a foundation for a diagnostic method for FM and require further purification and identification, hopefully paving the way for future targeted FM therapy.

Cells doi: 10.3390/cells15110973

Authors: Borghini Palazzo Gorini

Selenium (Se), a vital trace element, plays a significant role in maintaining vascular health and may offer protective effects against atherosclerosis. Its actions are mediated through Se-dependent selenoenzymes and selenoproteins, which enhance antioxidant defense, modulate inflammatory responses, and promote autophagy. These processes collectively help prevent cellular senescence—a state associated with age-related vascular decline characterized by oxidative stress, DNA damage, pro-inflammatory activity, and endothelial dysfunction. Epidemiological evidence consistently shows that low Se status is associated with increased risk of atherosclerotic cardiovascular disease within a narrow concentration range. However, clinical trials have not demonstrated clear reductions in cardiovascular events or mortality with Se supplementation alone. Overall, current evidence indicates that Se modulates key mechanisms involved in vascular aging and atherosclerosis, particularly redox balance, immune activation, and vascular cell homeostasis. This comprehensive review summarizes current epidemiological, clinical, and experimental research on the role of Se in cardiovascular health. It underscores Se’s potential as a promising strategy for the prevention and treatment of atherosclerosis, while also acknowledging the complexities and nuances of its effects on vascular health. A deeper understanding of the cellular and molecular mechanisms involved could pave the way for targeted interventions aimed at reducing the burden of atherosclerotic cardiovascular disease.

Cells doi: 10.3390/cells15110972

Authors: Muhammad Sohail Khan Muhammad Faizan Gabsik Yang Ki Sung Kang

Major depressive disorder (MDD) is a mental illness with high mortality, suicide, and relapse rates that could become the leading cause of health problems worldwide by 2030. The microbiota–gut–brain axis involves bidirectional communication between the human gut microbiota and the central nervous system (CNS). The gut microbiome is a complex ecosystem of approximately 100 trillion microorganisms, including viruses, bacteria, and fungi. The gut microbiota has recently been recognized for its impact on various diseases and health concerns. Several factors influence the composition and structure of gut microbes, ultimately affecting human physiology, with the nervous system being particularly vulnerable. The gut–brain–microbiota axis influences several important brain functions through numerous pathways, including vagus nerve signaling, gut microbial synthesis of metabolites, and immune-related chemicals. These factors can influence neurotransmitter activity, neuroinflammation, behavior, and mental health. Despite increased interest, the possibility of modifying the gut microbiota as a therapeutic approach remains unclear. Although numerous studies suggest that microbiota play an important role in many illnesses, the precise mechanisms are yet to be elucidated, and there are currently no evidence-based, microbiota-focused treatments for these illnesses. Recent research indicates that gut dysbiosis (GD) causes increased intestinal permeability (leaky gut), initiates systemic inflammation, and contaminates the blood. Opportunistic microbial metabolites cross the blood–brain barrier, triggering a neuroinflammatory cascade and apoptotic pathways while affecting neurogenesis and neurotransmitters, ultimately resulting in the development of MDD and anxiety. This review examined the factors influencing normal gut microbiota and GD-mediated MDD, as well as possible therapeutic options. The study outlines its objectives and methodological approaches, including the screening and filtering of research on GD-induced depression. Furthermore, it explored the daily use of dietary supplements, revealing new paths for clinical and preclinical research.

Cells doi: 10.3390/cells15110971

Authors: Young A Choi Ryan Konrad Elliot S. Pohlmann Eric Abenojar Agata Exner Edward Medof

Solid tumors typically expand in a “cold” immunosuppressive tumor microenvironment (TME) and resist killing by CAR T cells or conventional therapy. Herein, we show that intratumoral injection of C3a and C5a receptor 1 (C3ar/C5ar1) pharmaceutical antagonists in an in situ forming implant (ISFI) can robustly suppress such tumors. Antagonizing autocrine C3ar/C5ar1 signaling in eight human and murine cancers of diverse lineages was universally anti-mitotic and pro-apoptotic in vitro, and growth-repressive in vivo. In contrast to i.p. administration of C3ar/C5ar1 antagonists to tumor-bearing mice, injecting the antagonists intratumorally in slow release poly (lactic-co-glycolic acid) (PLGA) polymer caused near-complete tumor elimination. The focused blockade of C3ar/C5ar1 GPCR signaling in an intratumoral ISFI opposed solid cancers by jointly repressing cancer cell viability/growth, tumor-associated angiogenesis, and myeloid-derived suppressor cell (MDSC) recruitment. Thus, the sustained blockade of C3ar/C5ar1 signaling in an intratumoral ISFI uninterruptedly disrupts three processes essential for solid cancer growth while avoiding adverse effects on other cell types. Our findings may apply to multiple cancer types in which discrete tumor masses can be targeted.

Cells doi: 10.3390/cells15110969

Authors: Hyun Mi Kim Eun Jung Oh Suin Kwak Se Ok Han Ho Yun Chung

Hypertrophic scars are characterized by excessive collagen deposition, fibrotic remodeling, and functional impairment. However, the ability of current models is limited in recapitulating human pathology. This study presents a novel approach using induced pluripotent stem cell-derived scar organoids to model hypertrophic scar characteristics in vitro. Following established protocols, human pluripotent stem cells were differentiated into skin organoids and induced fibrotic transformation by treatment with TGF-β1 (10 ng/mL) and hypoxia (5% O2) from day 45 onward. Scar organoids exhibited significant contraction and increased collagen I deposition compared with skin organoids. Immunofluorescence analysis showed reduced LHX2 expression, indicating loss of hair follicle development, while collagen I expression was significantly elevated. Dark-field imaging revealed marked morphological divergence between skin and scar organoids. RNA sequencing revealed distinct transcriptomic profiles. Expression of hair follicle-associated gene families (KRT and KRTAP) was upregulated in scar organoids, whereas epidermal structure-related genes (KRT4, KRT7, CLDN7, and WNT7) were downregulated. These findings demonstrate that iPSC-derived scar organoids successfully recapitulate key features of human hypertrophic scars, including excessive collagen production, loss of skin appendage development, and contractile behavior. This platform offers potential for future applications in drug screening, precision medicine, and understanding the molecular mechanisms underlying scar formation.

Cells doi: 10.3390/cells15110970

Authors: Maria Derkaczew Kamila Zglejc-Waszak Piotr Podlasz Marcin Jozwik Joanna Wojtkiewicz

Background: Anxiety is a frequent clinical problem that becomes disabling when excessive or persistent. Cyclitols are naturally occurring polyhydroxy compounds, and inositols are the most abundant cyclitols in eukaryotic cells; several stereoisomers have been proposed as candidates for CNS-relevant effects. Methods: A narrative review was conducted using a structured search of biomedical bibliographic databases. The search was centered on myo-inositol, scyllo-inositol, and D-chiro-inositol in relation to anxiety-related outcomes. Results: The retrieved literature suggests some biological plausibility for anxiolytic effects of selected inositol stereoisomers through pathways related to intracellular signaling and neurotransmission. However, the available evidence is uneven and remains limited. The most informative findings concern myo-inositol and include both preclinical and clinical studies, whereas data on scyllo-inositol and D-chiro-inositol are scarce, particularly in relation to anxiety-related outcomes. Conclusions: Current evidence suggests a possible anxiolytic role of selected inositol stereoisomers; however, the existing data are limited and heterogeneous, and do not allow for definitive clinical conclusions. Further research is required.

Cells doi: 10.3390/cells15110968

Authors: Steven Timmermans Céline Van Dender Maxime Roes Elise Moens Tineke Vanderhaeghen Jolien Vandewalle Claude Libert

Sepsis, which affects 49 million people yearly, killing 11 million of them, is known to induce severe liver dysfunction. It is characterized by extensive metabolic reprogramming, resulting in acute metabolic loss of function and maladaptive repair that can prime the organ for fibrosis rather than functional regeneration. To understand how intercellular communication dictates these outcomes, we performed cell type-specific bulk RNA-sequencing on hepatocytes (HEP), hepatic stellate cells (HSCs), liver sinusoidal endothelial cells (LSECs), Kupffer cells (KC), and CD45+ leukocytes (CD45) from mice following polymicrobial sepsis. Cell-cell communication analyses using CellChat and NicheNet revealed a clear reorganization of the hepatic environment. While HSCs remain largely quiescent during homeostasis, after sepsis, they become the liver’s central signaling hub and broadcast potent fibrogenic and chemotactic signals (e.g., Ccl7) to surrounding cells. This actively suppresses hepatocyte metabolic functions, promotes leukocyte infiltration, and may further initiate early fibrogenic priming. Our findings highlight HSCs as regulators during septic acute liver injury, revealing communication nodes that could be targeted to constrain fibrosis responses and promote normal functions and repair.

Cells doi: 10.3390/cells15110967

Authors: Bethany P. Torr Jennifer P. Craig Simon J. Dean Trevor Sherwin Sanjay Marasini

Purpose: Preclinical studies report low-intensity ultraviolet C (UVC) light to be safe and effective in treating murine bacterial keratitis, however, limbal impacts of UVC have yet to be investigated directly. This study evaluated the depth and density of UVC-induced DNA damage in the porcine and human limbus following UVC exposures of varying supratherapeutic dose. Methods: The corneoscleral junction (limbus) of full-thickness porcine corneas was exposed to supratherapeutic doses of UVC light (265 nm, 1.93 mW/cm2) for 5, 10, 15, 30, or 60 min (exposure groups) or remained unexposed for the same durations (control groups), with a sample size of 6 per group. In parallel, human corneal tissue was exposed to UVC for 1 or 5 min and processed identically. Following exposure, all tissues were frozen, dissected, and analysed using immunohistochemistry to detect cyclobutane pyrimidine dimers (CPDs) as markers of DNA damage. CPD distribution, depth, and density were subsequently evaluated. Results: CPDs were localised predominantly in the superficial corneal epithelial layers, irrespective of the UVC dose. The mean ± SD thickness of the corneal epithelium in the UVC-exposed groups was 38.9 ± 18.9 µm, and the average depth of CPD formation was 13.3 ± 8.43 µm. The proportions of cells affected by CPDs within the corneal epithelium (mean ± SD) were 47.8 ± 25.6%, 58.5 ± 16.2%, 39.9 ± 26.4%, 41.3 ± 27.3%, and 38.9 ± 28.3% for exposure durations of 5, 10, 15, 30, and 60 min, respectively (p > 0.05). Human cornea showed similarly limited penetration, with no difference in CPD proportions between the 1 and 5 min UVC exposures (p = 0.70). Conclusions: UVC-induced DNA damage in both species was confined to the superficial cellular layers of the cornea, with no detectable damage observed in deeper tissues, including those where limbal stem cells reside, even after supratherapeutic doses of up to one hour of exposure.

Cells doi: 10.3390/cells15110966

Authors: Colin J. Hartman Petra Sergent Anna Barbara Emilia Zimmermann Olga R. Chávez-Alexander-Anderson Luis A. Perez Alonso Louise Lines Juan Carlos Pinto-Cárdenas Daniel Luna Dávalos Anna M. Schmoker Scott M. Palisoul Johannes vom Berg Xiaoxuan Ge Jay L. Rothstein Margaret E. Ackerman Steven Fiering Randolph J. Noelle Hugo Arias-Pulido

Cancer is a leading cause of death in dogs, and incidence rates in dogs exceed those in humans. Current therapeutic options for canine cancer patients remain limited, with most treatments focused on palliative care. Immune checkpoint inhibitors such as anti-PD-1, anti-PD-L1, and anti-CTLA-4 antibodies that have transformed cancer therapy and expanded the therapeutic options in humans could offer the same clinical benefit in canine cancer patients. This study details the engineering and functional characterization of mouse and chimeric mouse–canine anti-canine PD-1 (cPD-1) monoclonal antibodies. We demonstrate that anti-cPD-1 antibodies block the interaction between cPD-1 and its ligand cPD-L1, thereby inhibiting this immune signaling pathway. In a proof-of-concept study in seven companion canine cancer patients, intratumoral therapy with the lead anti-cPD-1 antibody (HugPetmab) was safe, well-tolerated, had no observed adverse events, and showed evidence of tumor control in a subset of injected tumors. These findings support the potential of HugPetmab antibody as an immunotherapeutic option for treating canine cancer patients.

Cells doi: 10.3390/cells15110965

Authors: Magda Mucha Alena Ryšavá Iwona Jarocka-Karpowicz Audrius Maruška Elżbieta Skrzydlewska Agnieszka Gęgotek

The aim of this study was to analyze the effect of concomitant use of cannabigerol (CBG) and 3-O-ethylascorbic acid (EAA) on changes in the proteome of UVA-irradiated skin melanocytes, with particular emphasis on adduct formation between lipid peroxidation products and metabolically important proteins. Proteomic analysis allowed the identification of 1248 proteins with statistically significantly changed expression following melanocytes irradiation and/or incubation with CBG/EAA. The top 25 proteins with the most strongly differentially abundant expression included proteins involved in cell protection/antioxidant response, as well as pro-inflammatory and proapoptotic signalization. Moreover, in melanocytes irradiated with UVA, the levels of lipid peroxidation product, 4-hydroxynonenal (4-HNE) and its protein adducts were increased, as well as significant changes in the profile of proteins modified by 4-HNE were observed. CBG and EAA, especially when used together, largely reverse these effects. This study for the first time demonstrated the combined effect of CBG and EAA on the proteome of melanocytes after their exposure to UVA radiation, which applies to both changes in protein expression and intracellular signaling based on proteins modified by 4-HNE. It can be suggested that CBG and EAA may provide melanocytes with effective protection against the effects of oxidative stress and perhaps even protect the skin from carcinogenesis.

Cells doi: 10.3390/cells15110964

Authors: Lukas Wenzel Leo Heinig Dongshi Wang Elise Vankriekelsvenne Nicole Wigger Annelie Zimmermann Johann Rößler Tim Clarner Markus Kipp

Reactive astrocytes are a hallmark of several neurological diseases in multiple sclerosis and experimental demyelination models. Their morphological alterations are commonly assessed by qualitative histopathology, yet quantitative tools are required to better capture astrocytic heterogeneity and to allow correlations with imaging-derived biomarkers. Here, we present a workflow for the quantitative analysis of Glial Fibrillary Acidic Protein (GFAP) network remodeling in astrocytes in the cuprizone model of demyelination. C57BL/6 mice were intoxicated with cuprizone for 3 or 5 weeks to induce progressive demyelination, microglial activation, and reactive astrogliosis. Brain sections were processed for anti-GFAP immunohistochemistry, and individual astrocytes from the stratum oriens of the hippocampus were digitally reconstructed. Diverse parameters of GFAP topology, including soma size, process length, branching order, convex hull area, and ramification index, were extracted using either the commercial Neurolucida® 360 software or the open-source Simple Neurite Tracer (SNT) plugin in ImageJ. Principal component analysis revealed clear differences between control astrocytes and astrocytes in cuprizone-intoxicated animals, with reactive astrocytes displaying increased numbers of primary processes, enhanced bifurcation, and process complexity. Comparative evaluation of Neurolucida® 360 and SNT demonstrated that both tools are suitable for astrocyte reconstruction, although Neurolucida® 360 enabled faster and more detailed tracing. This protocol provides a reproducible pipeline for the quantitative assessment of astrocyte morphology under control and pathological conditions, thereby supporting future efforts to link cellular remodeling to functional outcomes in neuroinflammatory disease models.

Cells doi: 10.3390/cells15110963

Authors: Valeria Fernandez Vallone Lina Hellwig Eddy Rijntjes Nicolai von Kügelgen Rajas Sane Robert Opitz Peter Kühnen Josef Köhrle Philipp Mergenthaler Harald Stachelscheid

Thyroid hormones (THs) are essential regulators of human brain development, and disrupted TH availability during pregnancy or early life is linked to adverse neurodevelopmental outcomes. Concerns that environmental chemicals interfere with TH signalling have increased the need for human-relevant in vitro systems to identify thyroid hormone system-disrupting chemicals (THSDCs) for risk assessment. Here, we compared two human-induced pluripotent stem cell (hiPSC)-derived brain organoid models for THSDC assessment: (i) human cortical organoids (COs) generated by unguided differentiation, offering higher architectural complexity but lower throughput; and (ii) neural stem cell-derived organoids (NSCOs), designed for scalability with reduced cellular diversity. Both models expressed key TH handling components, including the transporter SLC16A2 (MCT8) and the inactivating enzyme DIO3. Using LC–MS/MS, we show that exogenous T3 is depleted from culture media and metabolized to 3,3′-T2 and 3′-T1 in both models, alongside upregulation of T3-responsive genes (HR, KLF9, DIO3, SEMA3C). Pulse and chronic co-exposures to reference disruptors iopanoic acid (IA, deiodinase inhibitor) and silychristin (SC, MCT8 inhibitor) altered T3 metabolism and modulated T3-responsive transcriptional endpoints. In NSCOs, high-content imaging revealed treatment-associated changes in cell composition, with chronic T3 reducing the SOX2-positive progenitor pool and THSDCs blocking this effect. Together, these findings provide a framework for organoid qualification—linking TH handling, transcriptomic responsiveness, and scalable phenotypic readouts—as a necessary step toward model validation and implementation of brain organoids in THSDC risk assessment pipelines.

Cells doi: 10.3390/cells15110962

Authors: Muhammad Sohail Khan Imran Zafar Muhammad Noman Gabsik Yang Ki Sung Kang Jean C. Bopassa

Neurodegenerative disorders, including Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), and amyotrophic lateral sclerosis (ALS), arise from highly interconnected molecular and cellular abnormalities that progressively lead to neuronal dysfunction, synaptic failure, and cell death. This review provides a unified framework to understand the interrelated molecular mechanisms driving these diseases, with a focus on identifying key disease-specific intervention nodes. Core contributors include oxidative stress, mitochondrial dysfunction, protein aggregation, neuroinflammation, and emerging roles of peroxisomal dysfunction in redox imbalance, lipid dysregulation, and inflammatory amplification. Single-target therapies often show limited efficacy due to the complex, interconnected nature of these pathways. In contrast, polypharmacology, which targets multiple disease-relevant mechanisms simultaneously, offers a more promising therapeutic strategy. This review critically examines how pathway crosstalk drives neurodegenerative progression, with particular emphasis on mitochondrial–ROS–inflammatory signaling, aggregation–proteostasis failure, synaptic–neuroimmune dysfunction, and gut–brain communication. It evaluates various multi-node intervention strategies, including multi-target-directed ligands (MTDLs), molecular hybrids, natural products, drug repurposing, and nanocarrier-based delivery systems. Advances in network pharmacology, artificial intelligence (AI), bioinformatics, and multi-omics have enhanced the identification of actionable therapeutic nodes, candidate compounds, and brain-targeted delivery platforms. Notably, the NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome and cyclic GMP–AMP synthase (cGAS)—stimulator of interferon genes (STING) pathways—play distinct roles in neuroinflammation, amplifying neuronal damage by releasing inflammatory cytokines and inducing mitochondrial dysfunction. However, successful translation into clinical practice remains constrained by challenges such as blood–brain barrier penetration, patient heterogeneity, and biomarker limitations. The review advocates for a shift towards mechanism-informed, patient-stratified polypharmacological strategies to better address the network pathology of neurodegeneration, despite significant translational hurdles.

Cells doi: 10.3390/cells15110961

Authors: Maël Padelli Christophe Desterke Aurore Devocelle Denis Clay Agnès Bourillon Georges Uzan Antoinette Lemoine Julien Giron-Michel

Hepatocellular carcinoma (HCC) is a highly heterogeneous malignancy characterized by poor prognosis and limited therapeutic response. Cancer stem cells (CSCs) contribute to tumor progression, therapeutic resistance, and tumor recurrence. Among transcriptional regulators potentially involved in these processes, Specificity Protein 1 (SP1) has emerged as a candidate integrator of oncogenic and epigenetic signaling networks. However, its contribution to CSC-associated phenotypes and drug resistance in HCC remains incompletely defined. In this study, we combined transcriptomic analyses of TCGA datasets with functional experiments in HCC cell lines (Huh7 and HepG2). SP1-associated transcriptional programs were targeted pharmacologically using mithramycin A (MIT-A) and genetically using siRNA-mediated knockdown. The effects were assessed by RNA sequencing, RT-qPCR, Western blotting, flow cytometry, and functional assays evaluating proliferation, migration, CSC-associated properties, and response to sorafenib. MIT-A treatment markedly reduced the expression of stemness-associated transcription factors (NANOG, OCT4, SOX2) and CSC markers (CD133, CD24), impaired CSC-related functions including ALDH activity and the Side Population phenotype, and inhibited cell proliferation and migration. MIT-A also sensitized both parental and sorafenib-resistant HCC cells to sorafenib, associated with modulation of apoptotic regulators and reduced transporter-mediated efflux activity. SP1 knockdown partially reproduced several of these effects, supporting a contribution of SP1-dependent transcriptional programs to these phenotypes. Overall, these findings identify SP1-associated transcriptional networks as potential regulators of CSC features and therapeutic resistance in HCC and support targeting SP1-associated transcriptional programs as a strategy to enhance sorafenib efficacy.

Cells doi: 10.3390/cells15110959

Authors: Yimeng Chen Ding Zhang Jiajia Ma Huixin Li Jingrong Xu Cuixia Ma Yuqian Liu Zhenbing Zhao Garry P. Duffy Jun Ma Huixian Cui

Background: Rotenone is a widely used environmental pesticide, and epidemiological studies suggest that exposure is associated with an increased risk of Parkinson’s disease (PD); however, the molecular toxicological basis of this association remains incompletely defined. Ferroptosis is an iron-dependent, lipid peroxidation-driven form of regulated cell death that is relevant to PD and other neurodegenerative disorders. In this study, we provide disease-contextual functional evidence linking ferroptosis to rotenone-induced PD-like neurotoxicity. Methods: We combined network toxicology, human PD substantia nigra transcriptomic analysis using GSE7621, and SH-SY5Y cell-based validation. Rotenone-associated targets were predicted and analyzed for ferroptosis-related enrichment, PD transcriptomic signatures were used for disease-contextual candidate prioritization, and selected findings were validated using qPCR, CCK-8, Western blotting, C11-BODIPY lipid peroxidation staining, and transmission electron microscopy. Results: By further integrating a human PD substantia nigra transcriptomic dataset (GSE7621), we prioritized an 11-gene, PD-contextualized ferroptosis-associated candidate module (LIPF, FAM170A, MCHR1, IL17A, MYB, GFAP, ARMC3, GKN1, GATA3, IL17F, and TEKT1). In SH-SY5Y cells, rotenone exposure consistently upregulated this candidate transcriptional module, and this induction was broadly attenuated by the ferroptosis inhibitor ferrostatin-1 (Fer-1). In parallel, orthogonal functional assays supported an iron- and lipid peroxidation-driven injury state under rotenone exposure that was suppressible by ferroptosis inhibition and iron chelation. Finally, we further performed an exploratory drug–gene association screen to prioritize clinically available candidates, and a limited qPCR check suggested that several selected compounds partially attenuated representative hub-gene induction under rotenone exposure. Conclusions: Collectively, these findings provide disease-contextual and experimentally supported evidence linking rotenone exposure to ferroptosis-associated neurotoxicity, and identify a ferroptosis-responsive transcriptional module for future hypothesis-driven mechanistic investigation.

Cells doi: 10.3390/cells15110960

Authors: Sylwia Gul-Hinc Andrzej Szutowicz Anna Ronowska Agnieszka Jankowska-Kulawy

Glucose is the principal energy substrate for the brain. Hypo- and hyperglycemic episodes frequently occur in senescent people, contributing to functional and structural impairment of brain neurons and causing cognitive deficits in this population. In this study, we investigate whether long-term changes in the extracellular concentration of glucose affect viability and transmitter functions of septum-derived SN56 cholinergic neuronal cells through alterations in acetyl-CoA availability. Cells with low cholinergic expression (NCs) and cAMP/retinoic acid-induced high cholinergic expression (DCs) were investigated. Hypoglycemia brought about similar (approximately 20–30%) decreases in pyruvate dehydrogenase complex (PDHC) and ATP-citrate lyase (ACLY) activities and a 65% decline in lactate dehydrogenase (LDH) activity in NCs and DCs. Choline acetyltransferase (ChAT) and LDH activities in DCs were about 3–8 and 1.7–2.4 times higher than in NCs over the tested glucose concentration range, respectively. DCs appeared to be more resistant than NCs to hypoglycemia, as evidenced by lower glucose IC50 values for cell count and intracellular LDH activity. On the other hand, some of functional properties of DCs, such as the cholinergic phenotype and their plasma membrane functions (trypan blue exclusion, TB+), were found to be more sensitive to hypoglycemia than those of NCs, as demonstrated by the higher IC50 for glucose in DCs. Acetyl-CoA levels in DCs were 40% lower than in NCs, and decreased by about 25% with increasing hypoglycemia in both cell types. The cytotoxic effects of amyloid-β25–35 (Aβ) and sodium nitroprusside (SNP; NO generator) were also tested. In 25 mM glucose medium, these toxic compounds exerted greater detrimental effects on DCs than on NCs. In contrast, in 1 mM glucose, more evident cytotoxicity of SNP and Aβ was observed in NCs. These data suggest that the higher rate of glycolysis in differentiated cholinergic septal neurons may be a protective mechanism against hypoglycemia.

Cells doi: 10.3390/cells15110958

Authors: Veronica Bensa Hugo Lopes-Cardoso Martina Ardito Eleonora Ciampi Anastasiya Voronovska João Soares-Gonçalves Mirco Ponzoni Chiara Brignole João Nuno Moreira Fabio Pastorino

Background: Neuroblastoma (NB) causes about 15% of cancer deaths in childhood. Recently, we suggested cell-surface nucleolin (NCL) as a novel target for preclinical therapy against NB. Methods: Here, a broad range of human NB cell lines were evaluated for NCL expression. PEGylated liposomal nanoparticles, co-encapsulating C6- or C18-ceramides and doxorubicin (DXR) and functionalized with the F3 peptide (F3-lipo[C6-DXR] or F3-lipo[C18-DXR]), were tested against NCL-expressing NB cell lines, grown in monolayers (2D) and as multicellular tumor spheroids (3D). Untargeted liposomes were used as the control. Cytotoxicity and apoptotic/necrotic deaths were evaluated. Results: All NB cell lines expressed cell-surface NCL. Compared to untargeted formulations, F3-lipo[C6-DXR] and F3-lipo[C18-DXR] showed enhanced cellular association and antitumor effects against NB cells. Compared to F3-lipo[C18-DXR], F3-lipo[C6-DXR] was significantly more effective in reducing 2D and 3D NB cell lines’ viability (2D: IC50 range 313–995 nM and 239–629 nM, respectively; 3D: IC50 range 202–416.2 nM and 62.61–398.6 nM, respectively) and in inducing apoptotic cell death. F3-lipo[C6-DXR] also led to a greater cytotoxicity compared to liposomal DXR alone, highlighting the benefit of co-encapsulation. Conclusions: NCL is a promising target in NB, and F3-targeted liposomes enable the selective delivery of their cargo. F3-lipo[C6-DXR] showed superior antitumor activity, supporting ceramide–DXR co-encapsulation as a potential treatment strategy, which needs to be further validated.

Cells doi: 10.3390/cells15110957

Authors: Hadeel Edkaidek Divakar Dahiya Poonam Singh Nigam

Biological activity of diets consisting of dietary fibers, peptides and polyphenols is largely mediated by the gut microbiota, which converts these compounds into bioactive metabolites. This review examines the microbiota–epigenome axis, highlighting gut microbiota-derived metabolites, including short-chain fatty acids (SCFAs), urolithins, and phenolic acids, that modulate host gene expression through DNA methylation, histone modifications, and non-coding RNA regulation. Current evidence from molecular and microbiome studies indicates that these metabolites influence key metabolic and inflammatory pathways, including lipid absorption via CD36, SIRT1 activation, and one-carbon metabolism involving folate and S-adenosylmethionine (SAM). Inter-individual variability in metabolic responses is associated with differences in microbial composition and metabotypes, which determine the magnitude of epigenetic regulation. Furthermore, dietary polyphenols derived from pomegranate, berries, tea, cocoa, and grapes are shown to modulate gut microbiota composition and enhance epigenetic effects. A “butyrate–polyphenol synergy” model is proposed, in which combined microbial metabolites optimize host epigenetic programming. Overall, agri-food by-products are suggested to function as modulators of the host epigenetic landscape, providing a framework for microbiome-targeted dietary strategies to improve metabolic and inflammatory health.

Cells doi: 10.3390/cells15110956

Authors: Francesca Mancini Michela Cicchinelli Emanuela Teveroni Erica Pazzaglia Donatella Lucchetti Giulia Artemi Valentina Palmieri Federica Iavarone Domenico Milardi Andrea Urbani Tullio Ghi Annamaria Merola Fiorella Di Nicuolo

Human follicular development depends on coordinated communication between granulosa cells (GCs) and oocytes through endocrine cues, direct contacts, and extracellular vesicles (EVs). Exosomes are key EV mediators of intrafollicular signaling, but their cargo and functions in gonadotropin-stimulated GCs remain poorly defined. The human granulosa-like tumor cell line KGN was used to investigate exosome secretion and protein composition following human chorionic gonadotropin (hCG) stimulation. Exosomes were isolated by ultracentrifugation, characterized via nanoparticle tracking analysis (NTA), Scanning Electron Microscopy (SEM) and Western blotting, and analyzed using high-resolution mass spectrometry. Comparative proteomics integrating exosomal profiles with the whole secretome were performed, followed by bioinformatic analyses of protein networks, gene ontology, and pathway enrichment. hCG reshaped exosomal cargo, identifying 59 proteins enriched in exosomes, including Integrin α3 (ITGα3), Galectin-3-binding protein (LGALS3BP), tetraspanins (CD63, CD151), and proteasome subunits. Functional enrichment indicated roles in extracellular matrix remodeling, integrin signaling, proteostasis, and steroidogenesis. Comparison with the secretome revealed distinct protein distributions, supporting selective exosomal packaging. Western blot confirmed increased ITGα3 and LGALS3BP levels in exosomes upon hCG treatment. In conclusion, hCG modulates exosome cargo composition in granulosa cells, uncovering a novel mechanism of extracellular regulation.

Cells doi: 10.3390/cells15100954

Authors: Lihua Zhao Yanglin Chen Xiyun Guo Meng Zhou Tianxu Guo Junjun Ma Manling Zhang Linxin Cheng Jinbo Yu Yu Zhang Guang Yang Rongfeng Li Xihe Li

The mammalian early embryo possesses totipotency and can be captured as extended pluripotent stem cells (EPSCs). The first two cell differentiations result in epiblast, primitive endoderm, and trophectoderm, with the trilineage coexisting in a unified uterine microenvironment. Nevertheless, the in vitro counterparts—primed PSCs, trophoblast stem cells (TSCs), and extraembryonic endoderm (XEN) cells—require a distinct culture system. In this study, we successfully derived stable porcine EPSCs from fibroblasts at 35% efficiency, and interestingly observed that these EPSCs differentiated in parallel and gave rise to the transient coexistence (36–84 h) of trilineage cells when cultured in a single system (LCDM: hLIF, CHIR99021, DiM, and MiH). Then, XEN cells gradually predominated and eventually became the sole population in prolonged LCDM culture. However, TSCs and primed PSCs had to be differentiated from EPSCs under their respective culture system. EPSCs can differentiate into PGC-like cells independently of genetic modification and contribute to mouse neurula-stage embryos. Collectively, the trilineage coexistence phenomenon may provide novel insight into an early embryogenesis mechanism and strategy for porcine blastoid construction.

Cells doi: 10.3390/cells15100955

Authors: Sofoklis Stavros Angeliki Gerede Efthalia Moustakli Athanasios Zikopoulos Ioannis Tsakiridis Christina Messini Anastasios Potiris Ismini Anagnostaki Ioannis Arkoulis Spyridon Topis Themistoklis Dagklis Dimitrios Loutradis

In the human reproductive system, extracellular vesicles (EVs) have been recognized as playing a vital role in mediating cell–cell communication. They are considered critical for embryo development, implantation, gamete interaction, and fertilization. The various cargoes carried by EVs, depending on the physiological and pathological state of the cell, include proteins, lipids, nucleic acids, and mitochondrial components. EVs are recognized as critical carriers of redox-related signals and mitochondrial components, linking oxidative stress (OS) to reproductive failure and influencing gamete quality and embryo competence. Although considerable progress has been made, research remains poorly integrated, despite individual omics technologies providing valuable molecular insights. The use of multi-omics technologies, including transcriptomics, proteomics, metabolomics, and microbiome analysis, has been proposed as a global approach to understanding the complexities associated with EVs and discovering new biomarkers associated with infertility. ML and AI have been proposed to identify predictive signatures linked to ART effectiveness and reproductive outcomes, with a strong capacity to handle high-dimensional data. The review aims to provide an overview of current knowledge on EV-mediated redox–mitochondrial signaling in human reproduction, while highlighting the importance of emerging multi-omics and AI technologies for EV-mediated biomarker development. The review discusses the promise of EVs in the development of minimally invasive diagnostic approaches and therapeutic interventions, as well as the challenges in the standardization, integration, and clinical translation of EV-mediated research. In addition, the review proposes integrating computational approaches to better understand molecular pathways involved in the development of next-generation precision medicine in human reproduction.

Cells doi: 10.3390/cells15100953

Authors: Maria-del-Carmen Silva-Lucero Gustavo Lopez-Toledo Víctor-Adrián Cortés-Morales Juan-José Montesinos Raúl Sampieri-Cabrera David-E. García Juan-Ramon Padilla-Mendoza Obed-Ricardo Lora-Marin Jesus-Adrian Buendia-Meraz Fausto-Alejandro Jiménez-Orozco Israel López-Reyes Paul Mondragon-Teran Maria-del-Carmen Cardenas-Aguayo

Background: Human dental pulp stem cells (hDPSCs) are a promising source of multipotent mesenchymal stem cells (MSCs) for regenerative neurology because of their inherent neurogenic potential. However, robust and reproducible protocols for driving their terminal neuronal maturation in a fully defined, xeno-free environment are lacking. Methods: hDPSCs were isolated from a donor tooth and characterized for mesenchymal (CD105, CD90, CD73, CD13) and stemness-associated markers (SOX2, Oct3/4 and Nanog). Cells were differentiated in a novel, fully chemically defined medium 1% ITS medium (ITS: Insulin, Transferrin, Selenium) supplemented with glial cell line-derived neurotrophic factor (GDNF) or brain-derived neurotrophic factor (BDNF). Neuronal commitment and partial maturation were assessed via immunofluorescence, Western blot, and RT-PCR for markers such as NeuN (Neuronal nuclei) and NF-M (Neurofilament medium chain), and functionally by whole-cell patch-clamp electrophysiology. Results: Although undifferentiated hDPSCs expressed neural progenitor markers (βIII-tubulin and Nestin), only GDNF treatment in a chemically defined medium significantly upregulated mature neuronal markers (NeuN and NF-M) and downregulated mesenchymal markers. Importantly, GDNF-treated cells exhibited key functional changes, including hyperpolarized resting membrane potentials, increased membrane capacitance, and elevated input resistance, which are electrophysiological hallmarks of neural precursor or early neuronal maturation, compared to control cells cultured in medium containing fetal bovine serum (FBS). Although action potentials were not elicited, this represents a significant advancement toward achieving a functional neuronal state. Conclusion: This study demonstrates that a fully chemically defined medium enables GDNF to drive hDPSCs beyond the neural progenitor state towards a partially mature neuronal phenotype. This defined medium protocol eliminates serum variability, enhances reproducibility, and provides a critical step towards standardizing hDPSC-derived neuronal cells for disease modeling and cell-based therapy.

Cells doi: 10.3390/cells15100951

Authors: Mario Palumbo Luigi Della Corte Maria Rotonda Conte Giuseppe D’Angelo Mario Ascione Antonisia Pollio Pierluigi Giampaolino Giuseppe Bifulco

Background: Endometriosis is traditionally conceptualized as a localized gynecological disorder characterized by the presence of ectopic endometrial tissue. However, high recurrence rates following apparently complete surgical excision challenge this lesion-based paradigm and suggest the existence of underlying biological mechanisms that extend beyond residual disease. Increasing evidence indicates that endometriotic cells exhibit persistent molecular alterations, including dysregulated gene expression, epigenetic modifications, and immune dysfunction, which may contribute to disease maintenance and recurrence. Objective: This study aims to critically examine whether endometriosis can be considered a molecularly reprogrammed disease, characterized by persistent cellular and microenvironmental alterations that are not reversed by surgical removal of visible lesions. Methods: A narrative review of the literature was conducted using PubMed, Scopus, and Web of Science databases including studies published from January 2016 to March 2026. Studies investigating molecular, genetic, epigenetic, and immunological mechanisms of endometriosis persistence and recurrence were included. Particular attention was given to pathways involved in cellular survival, inflammation, hormone resistance, and epigenetic regulation. Results: Endometriotic cells demonstrate stable alterations in gene expression profiles, including pathways related to estrogen signaling, progesterone resistance, inflammation, and cellular proliferation. Epigenetic mechanisms, such as aberrant DNA methylation and histone modifications, appear to sustain these changes over time, contributing to a form of “molecular memory.” In parallel, the peritoneal microenvironment is characterized by chronic inflammation, immune tolerance, and impaired clearance of ectopic cells. These factors collectively support lesion persistence and may explain recurrence even after complete surgical excision. Emerging evidence also highlights the role of systemic factors, including endocrine–immune interactions and microbiome-related pathways, reinforcing the concept of endometriosis as a systemic rather than purely localized condition. Conclusions: Endometriosis may be more accurately defined as a persistent, molecularly reprogrammed disease driven by stable alterations in cellular behavior and the surrounding microenvironment. This paradigm shift has important clinical implications, suggesting that surgical treatment alone may be insufficient and that future therapeutic strategies should target the underlying molecular and immunological mechanisms responsible for disease persistence.

Cells doi: 10.3390/cells15100952

Authors: Jingya Yu Lulu Xin Ying Cui Chunxin Fan Yongheng Yang Xiaolu Zhang

Histone lactylation acts as a master regulator in tumor development, but its role in a noncoding RNA (ncRNA) network remains unclear. This study aims to reveal the interaction between H3K18la and the lncRNA-miRNA-mRNA regulatory network in hepatocellular carcinoma (HCC). Transcriptome sequencing and ChIP sequencing were performed in HCC and adjacent normal tissues. Cut&Run and qPCR were used to validate the H3K18la enrichment on LINC02732 and CD44 promoter. Dual luciferase reporter assay, qPCR and Western blotting were used to verify the LINC02732-miR-1291-ASF1B axis. Co-Immunoprecipitation was performed to validate ASF1B recruiting p300. CCK8 and mouse subcutaneous tumor formation were performed to demonstrate this axis promoting HCC. H3K18la enrichment on LINC02732 promoter elevates its expression in both HCC samples and cell lines, therefore enhancing ASF1B expression via sponging miR-1291. Moreover, ASF1B, a histone chaperone, promotes H3K18la by recruiting lactyltransferase p300, forming an ASF1B-H3K18la positive feedback loop. The axis upregulates CD44 expression and promotes HCC in vitro and in vivo. These findings demonstrated the influence of H3K18la on the LINC02732-miR-1291-ASF1B axis and the novel role of ASF1B in histone lactylation by recruiting p300, which together promoted HCC proliferation.

Cells doi: 10.3390/cells15100948

Authors: Nadeem Samee Lou Belz Nicolas Narboux-Nême Jean-Christophe Roux Nicolas Panayotis Giovanni Levi

Rett syndrome is a severe neurodevelopmental disorder caused predominantly by loss-of-function mutations in the X-linked gene MECP2. In addition to a vast array of neurological and physiological impairments, patients also frequently develop severe osteopenia with increased fracture risk; however, the mechanisms underlying these skeletal defects are not completely understood. Previous work in Mecp2-null mouse models has suggested that osteopenia is mainly due to impaired osteoblast function and reduced bone formation. Here, we examined bone mass, microarchitecture, and remodeling parameters in a Mecp2-null mouse model during postnatal development, with a particular focus on osteoclast involvement. Microcomputed tomography and histomorphometric analyses showed reduced bone mineral density and trabecular bone volume, which are associated with increased trabecular separation and cortical thinning. These structural alterations were accompanied by increased osteoclast number per bone surface, elevated urinary deoxypyridinoline, and higher expression of osteoclast-associated genes, including Cathepsin K. Furthermore, gene expression analysis revealed an age-dependent shift in bone remodeling. At postnatal day 35, mutant mice showed reduced expression of Dlx5 and Dlx6, consistent with low bone turnover. By postnatal day 55, Rankl and Cathepsin K were markedly upregulated, suggesting an increase in osteoclast resorptive activity, while key osteoblast markers and the RANKL/OPG ratio did not change significantly. A potential cell-autonomous contribution of Mecp2 to osteoclast maturation is also suggested by the analysis of public transcriptomic datasets on human osteoclast differentiation. Together, our findings identify increased osteoclast activity as a significant contributor to Rett-associated osteopenia and suggest that skeletal pathology in Mecp2 deficiency progresses from an early low-turnover state to a later phase of increased osteoclast resorption.

Cells doi: 10.3390/cells15100950

Authors: Elisa Farsetti Sarah Amato Monica Averna Elena Gatta Diego Guidolin Marco Pedrazzi Laura Lori Matilde Gnecco Guido Maura Luigi F. Agnati Manuela Marcoli Chiara Cervetto

Oxytocin’s capacity to affect the glial cell functions is increasingly recognized. We previously reported that oxytocin could cause both excitation and inhibition of Ca2+ signals and glutamate release in the processes of adult rodent astrocytes. Our purpose here was to investigate oxytocin receptor expression and oxytocin effects in astrocytes. In primary cortical astrocytes, we assessed the presence of oxytocin receptors by confocal imaging, and the effects of oxytocin receptor activation on intracellular Ca2+ signals and glutamate release. We found that oxytocin receptors are expressed in both the soma and processes of astrocytes; oxytocin at nanomolar concentrations could induce dual responses in astrocytes, namely facilitation and inhibition of Ca2+ signals and glutamate release; the oxytocin facilitatory and inhibitory effects were duplicated by the biased agonists carbetocin and atosiban, respectively; and the facilitatory and the inhibitory effect were dependent on activation of a Gq and a Gi pathway, respectively. It is concluded that oxytocin effects in astrocytes could duplicate the effects in processes prepared from astrocytes matured in neuron-astrocyte networks, substantiating the use of astrocytes to study astrocytic oxytocin molecular signaling.

Cells doi: 10.3390/cells15100949

Authors: Xiaoke Wang Chunyan Fang Shanyu Li Huakai Zeng Junyi Liu Xinwei Duan Xiaoyi Zhang Wenyan Jiang Xuejian Wang

Differentiation therapy holds significant potential for the treatment of multiple myeloma (MM). We previously identified that the aminopeptidase N (APN) inhibitor Bestatin promotes MM cell differentiation. Herein, we elucidate the underlying molecular mechanisms of this process. Utilizing MM1.S, U266, and RPMI-8226 cell lines, a combination of CCK-8 assays, flow cytometry, Wright–Giemsa staining, Western blotting, qRT-PCR, ELISA, APN enzymatic activity analysis, SA-β-gal staining, and bioinformatic analyses revealed elevated APN expression across all cell types. Bestatin treatment induced MM cell differentiation in a concentration-dependent manner, which was accompanied by the upregulation of the differentiation marker CD49e, increased immunoglobulin light chain secretion, elevated cellular senescence, and a concomitant suppression of cell proliferation and APN enzymatic activity. Mechanistically, Bestatin exerts its effects by downregulating the CD79B/BTK signaling pathway, thereby activating the downstream transcription factor STAT3. Consistent with this axis, direct inhibition of CD79B/BTK alone was sufficient to induce differentiation, while blockade of STAT3 completely abrogated the differentiation-promoting effect of Bestatin. The APN-neutralizing antibody (WM15) yielded consistent results with Bestatin, further validating this regulatory axis. Furthermore, both the CD79B/BTK inhibitor Ibrutinib and the STAT3 agonist GCDA potentiated the cytotoxicity of the clinical MM drug Ixazomib. Bestatin itself synergized with Ixazomib and enhanced the anti-proliferative effect of IL-6. In summary, our findings establish that the APN inhibitor Bestatin induces MM cell differentiation via the CD79B/BTK-STAT3 signaling axis. Targeting this pathway represents a promising strategy to enhance the efficacy of Ixazomib, providing a compelling rationale for novel combination therapies in MM.

Cells doi: 10.3390/cells15100947

Authors: Shuai Wang Xinyue Tu Haozhe Zhu Ce Gao Jianan Gao Jinsong Wei Hui Shi Jinrong Peng

Cell lineage relationship studies in developmental and regenerative biology have been greatly advanced using techniques such as fluorescent labeling driven by cell-type-specific promoters. Nevertheless, unbiased non-invasive tools for distinguishing cell lineages are inevitably desired. Mitochondrial DNA (mtDNA) exhibits wide-range single-nucleotide polymorphisms (SNPs) among individual cells. Here, we aim to distinguish cell types in organs/tissues of the same individual and in the regenerated liver based on the use of mtDNA SNPs. For this, two approaches—“Mitochondrial Alteration Enrichment and Sequencing” (MAESTER) and “mitochondrial single-cell assay for transposase-accessible chromatin with sequencing” (mtscATAC-seq)—were adopted to facilitate the detection of mtDNA SNPs in single cells. With MAESTER, we show that specific cell types in the liver and spleen of the same individual can be successfully defined using collective individual-specific markers composed of panels of unique mtDNA SNP combinations. For its application, we performed partial hepatectomy (PH) on a Krt19:DreERT2/+;R26:Rox-ZsGreen-Stop-Rox-tdTomato/+ mouse harboring tdTomato-labeled cholangiocytes following tamoxifen injection and demonstrated that utilizing panels of unique mtDNA SNP combinations detected by mtscATAC-seq in the pre-PH cholangiocytes as markers can faithfully trace the cell fate in the post-PH liver samples. Hence, this approach may serve as an unbiased tool for investigating cell lineage relationships in relevant research areas such as liver regeneration.

Cells doi: 10.3390/cells15100946

Authors: Marva Khalid Marvin L. Frommer Jeries Abu-Hanna Benjamin J. Langridge Clara Calero Pages Laura Awad Peter E. M. Butler

Adipose-derived stem cells (ADSCs) are widely used in regenerative medicine and are considered key effectors underlying the therapeutic efficacy of autologous fat grafting for scarring and skin fibrosis, yet clinical outcomes remain variable. This review examines how obesity alters the adipose microenvironment through chronic inflammation and metabolic dysfunction, resulting in epigenetic changes, mitochondrial impairment, oxidative stress, and premature cellular senescence in ADSCs. ADSCs from obese individuals exhibit reduced stemness, impaired differentiation, and a pro-inflammatory secretome with diminished regenerative capacity. While weight loss may partially reverse these effects, persistent epigenetic and functional memory limits full recovery. This review argues that donor metabolic status is a determinant of ADSC therapeutic potency and discusses key challenges and opportunities for improving regenerative outcomes.

Cells doi: 10.3390/cells15100945

Authors: René D. Verboket Lea Usov Isabell Bohl Jonas Neijhoft Marissa Penna-Martinez Ingo Marzi Dirk Henrich

Introduction: platelet-rich fibrin (PRF) is a second-generation platelet concentrate which is known for promoting cell migration, tissue repair, angiogenesis and bone formation. In contrast, the specific effects of trauma-activated PRF on mesenchymal stromal cells (MSC) are not yet fully understood. The present study investigates systemic effects of surgical trauma-activated PRF on MSCs in vitro, analyzing their metabolic activity, inflammatory responses, and regenerative capacity to optimize advanced treatment concepts for severe fractures and injuries. Material & Methods: PRF membranes (T-PRF from trauma patients, C-PRF from healthy controls) were generated. After co-incubation with MSC cells for 24, 72, and 120 h, further investigations of metabolic activity (MTT assay) and gene expression analyses were performed. Results: for MTT assay, results especially showed a significantly higher metabolic activity of T-PRF after 120 h. ELISA-results measuring cytokine levels (CXCL10, IL-6, VEGF, and IDO) exposed a frequent peak in T-PRF group at 72 h, declining slightly at 120 h. In the gene expression analyses, T-PRF exerted a comparatively stronger stimulating effect on MAPK14 and VEGFA after 24 h, while a decrease in gene expression for MAPK8, MAPK14, and RUNX2 was observed over time. Conclusion: surgical trauma-activated PRF seems to be a powerful inducer of early inflammatory and stress responses in MSCs with preserved angiogenic but limited osteogenic signaling. Therefore, a targeted balance between inflammatory activation and sustainable regeneration, as well as optimized preparation and possible combination with immunomodulatory approaches, appear to be crucial for the therapeutic success of PRF-based strategies.

Cells doi: 10.3390/cells15100944

Authors: Kamile Minkelyte Daqing Li Ying Li Ahmed Ibrahim

Olfactory ensheathing cell (OEC) transplantation has been widely shown to support axonal regeneration, remyelination, and functional recovery after central nervous system injury; however, autologous approaches are limited by low cell yields from patient biopsies, which may be insufficient for large spinal cord lesions. This study evaluated whether cryopreservation could provide a scalable alternative by preserving the therapeutic potential of mucosa-derived OECs. Using a rat dorsal root injury model, cryopreserved mucosa-derived OECs (CmOECs) were thawed and assessed for viability, phenotype, and efficacy following transplantation. Although total viable cell yield was reduced compared with primary cultures, the relative proportion of OECs remained stable, and cells retained characteristic morphology and marker expression in vitro. In vivo, transplantation of CmOECs resulted in significant functional recovery in climbing and forepaw fault tasks compared with injured controls, with outcomes comparable to primary mucosal OEC transplantation. Immunohistochemical analysis confirmed the survival and integration of transplanted cells at the dorsal root entry zone, alongside evidence of axonal regeneration and astrocytic remodeling. These findings demonstrate that mucosa-derived OECs retain therapeutic efficacy following cryopreservation and support the development of standardized OEC biobanks as a scalable strategy for spinal cord repair.

Cells doi: 10.3390/cells15100943

Authors: Yang Sun Wen Qin Yiting Gong Yinqiao Jian Fangling Jiang Rosa M. Rivero Ron Mittler Zhen Wu Rong Zhou

RNA N6-methyladenosine (m6A) is a prevalent epitranscriptomic modification that governs plant growth, development, and environmental adaptation. This review synthesizes recent advances in understanding the molecular mechanisms and biological functions of m6A in plants. The m6A landscape is dynamically regulated by methyltransferases (writers), demethylases (erasers), and m6A-binding proteins (readers), which collectively influence mRNA stability, translation efficiency, alternative polyadenylation (APA), and chromatin crosstalk. Functionally, m6A integrates diverse developmental processes—including embryogenesis, organogenesis, flowering, fruit ripening, and leaf senescence—with abiotic stress responses such as salt, drought, cold, and heat. Notably, m6A modification exhibits remarkable species-, cultivar-, and tissue-specific plasticity, enabling precise spatiotemporal gene regulation. Recent breakthroughs have revealed bidirectional crosstalk between m6A and histone modifications, forming a multi-layered regulatory network, while emerging concepts including phase separation, RNA structure dynamics, and stress memory further expand the functional repertoire of m6A. Despite significant progress, plant epitranscriptomics remains mechanistically underexplored, with critical gaps persisting in our understanding of translation initiation mechanisms, upstream regulatory signals controlling writers/erasers activities, and the functional significance of individual m6A sites. This review provided systematic insights into the complexity and specificity of m6A regulation in plants, offering a theoretical foundation for future efforts to decipher and ultimately manipulate this epitranscriptional layer for crop improvement.

Cells doi: 10.3390/cells15100942

Authors: Łukasz Pietras Marta Migocka-Patrzałek Bartosz Budziak Dariusz Rakus Agnieszka Gizak

Fructose 1,6-bisphosphatase 2 (FBP2) is a multifunctional protein whose cellular functions depend on its oligomeric state. Forced FBP2 tetramerization has been linked to microtubule disruption and impaired mitochondrial trafficking, accompanied by abnormal mitochondrial morphology. Here, we identify MIC60 (mitofilin), a core element of the mitochondrial contact site and cristae organizing system (MICOS), as a potential mediator of these effects. Using proximity ligation assay, protein crosslinking combined with mass spectrometry, and ultrastructural analysis, we demonstrate that FBP2 is in close proximity to MIC60 under basal conditions and this proximity is reduced upon FBP2 tetramerization or partial FBP2 depletion. Loss of this proximity coincides with marked remodeling of inner-membrane ultrastructure. These findings are consistent with a working model in which dimeric FBP2 contributes to the coordination of microtubule-dependent mitochondrial positioning with MICOS-linked intramitochondrial organization, providing a plausible mechanistic bridge between metabolic cues (AMP/NAD+) and mitochondrial structural integrity.

Cells doi: 10.3390/cells15100940

Authors: Maxime Roes Claude Libert Jolien Vandewalle

Peroxisome proliferator-activated receptor alpha (PPARα) is a key transcriptional regulator of lipid metabolism, highly expressed in metabolically active organs such as the heart. In cardiomyocytes, where approximately 70% of energy is derived from fatty acid oxidation, PPARα plays a central role in maintaining metabolic homeostasis. Moreover, the transcription factor is implicated in postnatal maturation of the heart and immune modulation. Dysregulation of PPARα signaling has profound consequences for cardiac energy balance, particularly under stress conditions. Accordingly, its role has been extensively investigated in cardiovascular diseases, including ischemia/reperfusion, diabetic cardiomyopathy and sepsis-induced cardiomyopathy. Upon ischemia/reperfusion and sepsis, cardiac PPARα expression is typically downregulated, contributing to impaired fatty acid breakdown and reduced metabolic flexibility. In contrast, diabetic cardiomyopathy is characterized by sustained PPARα activation, promoting excessive fatty acid oxidation, lipid accumulation and lipotoxicity. These context-dependent effects highlight a complex role of PPARα in cardiac diseases. PPARα has emerged as a promising therapeutic target, as its modulation can alleviate cardiac injury in preclinical models. However, further research is required to validate its efficacy in human disease, improve cardiomyocyte-specific targeting strategies to minimize systemic side effects, and better define optimal timing of intervention, as inappropriate or prolonged modulation may lead to detrimental outcomes.

Cells doi: 10.3390/cells15100941

Authors: Mohamed Abdelhakim Toshio Miyata

Plasminogen activator inhibitor-1 (PAI-1), encoded by SERPINE1, is the principal physiological inhibitor of tissue-type and urokinase-type plasminogen activators and a central regulator of fibrinolysis. Beyond its canonical hemostatic role, PAI-1 has emerged as a pleiotropic mediator of tissue remodeling, fibrosis, metabolic dysfunction, cancer progression, cellular senescence, and age-associated immune dysregulation. A central argument of this review is that PAI-1 should be understood not only as a downstream biomarker of aging-associated pathology, but also as an active effector linking senescence-associated secretory phenotype (SASP) signaling, chronic low-grade inflammation, impaired immune surveillance, fibrotic extracellular matrix remodeling, and a prothrombotic state. In this framework, PAI-1 may function as an immune-aging checkpoint: a molecular node through which senescent, stromal, malignant, and inflammatory cells reinforce immune evasion and tissue dysfunction. Structure-guided drug discovery has enabled the development of small-molecule PAI-1 inhibitors, including TM5275, TM5441, TM5509, and TM5614. Among these, TM5614 is an orally available investigational compound that has progressed to clinical evaluation. Preclinical studies support anti-thrombotic, anti-fibrotic, anti-inflammatory, anti-senescent, and tumor-microenvironment-modulating effects of PAI-1 inhibition, while early clinical studies have evaluated TM5614 in chronic myeloid leukemia, immune-checkpoint-refractory malignant melanoma, non-small-cell lung cancer, and COVID-19-associated pneumonia. This review summarizes the biology of PAI-1, expands the discussion of immunoaging, reviews representative preclinical and clinical data, compares available PAI-1 inhibitors, and discusses the translational opportunities and safety considerations for TM5614 and related compounds.

Cells doi: 10.3390/cells15100939

Authors: Magalie Tardif Manon Saby Stéphanie Forté Thomas Pincez

Sickle cell disease (SCD) is a monogenic disorder responsible for recurrent vaso-occlusive crises, progressive organ damage, and shortened life expectancy. For decades, allogeneic hematopoietic stem cell transplantation from a matched sibling donor has been the only established cure, but its reach remains limited by donor availability and transplant-related toxicity. The approval of two autologous gene therapy products in 2023, exagamglogene autotemcel (exa-cel) and lovotibeglogene autotemcel (lovo-cel), marked a turning point for the SCD population and the gene therapy field in general. This review proposes a molecular rationale for fetal hemoglobin reactivation and β-globin gene addition, describes the engineering of lentiviral and CRISPR-based platforms, and highlights the clinical evidence accumulated to date that demonstrated durable disease modification with acceptable short-term toxicity. We then assess the clinical positioning of gene therapy within the broader spectrum of curative options compared to current available treatments and address the financial, ethical and psychosocial barriers that limit access to gene therapy both within high-income countries and globally. Critical research priorities include long-term safety surveillance, comparative effectiveness studies, pediatric trials below 12 years, and validated patient-reported outcome instruments. Base editing, non-genotoxic conditioning, and in vivo delivery represent the most promising avenues to broaden access and reduce treatment burden.

Cells doi: 10.3390/cells15100938

Authors: Joen-Rong Sheu Kuan-Hung Lin Ray-Jade Chen Hao-Ping Chia Ting-Yu Chen Thanasekaran Jayakumar Hsueh-Hsiao Wang Hsien-Yu Peng Jiun-Yi Li Wan-Jung Lu

Background/Objectives: Circulating platelets mediate physiological hemostasis and are implicated in pathological thrombosis, which can cause vascular occlusion, leading to heart attacks or strokes. Costunolide is a sesquiterpene lactone extracted from Saussurea lappa. Although this lactone has multiple biological effects, including anti-inflammatory and antioxidant effects, that help slow the progression of atherosclerosis, its influence on platelet activation remains unclear. In this study, we examined the potential antiplatelet and antithrombotic effects of costunolide. Methods: We used platelet aggregation, flow cytometry, and Western blot analysis to examine its in vitro antiplatelet effects. Results: Our results indicated that costunolide inhibited platelet aggregation induced by collagen, but not by thrombin or the thromboxane A2 analog U46619, suggesting that costunolide selectively inhibits collagen-induced platelet activation. Additionally, costunolide blocked collagen-mediated granule release, calcium mobilization, and glycoprotein IIb/IIIa (GPIIb/IIIa) activation. Costunolide also inhibited phospholipase Cγ2 (PLCγ2), pleckstrin (a downstream target of protein kinase C), Akt, and mitogen-activated protein kinase. Moreover, it prevented collagen/epinephrine-induced pulmonary thrombosis and increased the survival rate of mice. Furthermore, costunolide delayed thrombus formation in the mesenteric vessels while it did not significantly affect hemostasis, suggesting it exhibits antithrombotic activity without bleeding tendency. These findings indicate that costunolide can block PLCγ2-PKC, Akt, and MAPK signaling pathways and subsequent granule release, calcium mobilization, and GPIIb/IIIa activation, eventually impeding platelet activation, platelet aggregation, and thrombus formation. Conclusions: In conclusion, besides its multiple biological activities that are beneficial for slowing the progression of atherosclerosis, we also demonstrated the antiplatelet and antithrombotic activities of costunolide. These effects highlight the therapeutic potential of costunolide in the treatment of patients with cardiovascular disease, particularly stroke and heart attack.

Cells doi: 10.3390/cells15100937

Authors: Krishna Sharma Puttur Santhoshkumar Tenzin Tender

α-Crystallin, the predominant protein of the eye lens, possesses molecular chaperone activity and antioxidative properties, both of which are essential for maintaining lens transparency. Its chaperone function prevents the formation of light-scattering protein aggregates, while its antioxidative activity mitigates oxidative stress through both direct and indirect mechanisms. However, with aging, α-crystallin undergoes cumulative post-translational modifications and oxidative damage, leading to protein crosslinking and a decline in chaperone efficacy. Notably, α-crystallin exhibits free radical-scavenging activity comparable to that of serum albumin, a well-characterized antioxidant protein. In addition, its ability to bind redox-active metal ions and convert them into redox-inactive forms significantly reduces reactive oxygen species (ROS) generation in vivo. α-Crystallin also interacts with key proteins and signaling pathways involved in oxidative stress responses, further enhancing its multifunctional protective role. This review summarizes current evidence on the antioxidative properties of α-crystallin and their relationship to its chaperone function, highlighting its importance in lens homeostasis and age-related cataract formation.

Cells doi: 10.3390/cells15100935

Authors: Changshun Li Xin Zhang Peiyu Tang Mengying Li Weijian Hu Meng Zhou Jiabin Xu

Icariin (ICA), a prenylated flavonoid glycoside from Epimedium (Yin Yang Huo), exhibits multi-organ pharmacological effects and has emerged as a promising candidate for osteoporosis therapy and bone tissue regeneration because of its capacity to modulate diverse osteogenic, anti-inflammatory, and angiogenic signaling pathways. Preclinical studies in osteoporotic models suggest that ICA improves trabecular microarchitecture and increases bone mineral density. Mechanistically, ICA modulates bone remodeling bidirectionally: it promotes osteoblast differentiation and extracellular matrix mineralization via activation of pro-osteogenic pathways, including Wnt/β-catenin and PI3K/Akt signaling, while simultaneously inhibiting osteoclastogenesis and bone resorption by suppressing RANKL-mediated NF-κB activation, thus reestablishing remodeling equilibrium. Despite these benefits, clinical advancement is hindered by the suboptimal oral bioavailability of ICA, stemming from poor intestinal absorption and extensive first-pass metabolism. To address this, innovative delivery systems have been engineered to enhance localized bioavailability and sustain therapeutic efficacy, such as hydrogel depots, nanoparticle formulations, and 3D-printed scaffolds enabling precise, controlled release. In bone tissue engineering applications, ICA-incorporated biomaterials—either standalone or in combination with osteogenic factors or exosomes—foster a regenerative niche by mitigating inflammation and oxidative stress, while synergistically promoting osteogenesis and angiogenesis, thereby expediting bone defect healing and osseointegration. Overall, these mechanistic elucidations and delivery advancements underscore ICA’s potential as a translational candidate for osteoporosis treatment and bone regenerative therapies. This review aims to critically and systematically synthesize current evidence on ICA-mediated bone repair and regeneration, with a particular emphasis on the molecular regulation of osteogenic signaling, the restoration of bone-remodeling homeostasis, and delivery-system-enabled strategies that may facilitate translational application.

Cells doi: 10.3390/cells15100936

Authors: Norifumi Takeda Hiroki Yagi Takayuki Fujiwara Hitomi Aono-Setoguchi Ryo Inuzuka Issei Komuro

Heritable thoracic aortic diseases (HTAD) are inherited conditions that increase the risk of thoracic aortic aneurysms, dissections, and premature aortic rupture. Advances in human genetics and experimental models have transformed our understanding of these disorders from a phenotype-based classification system to a mechanism-based view involving extracellular matrix (ECM) architecture, transforming growth factor-β (TGFβ) signaling, and vascular smooth muscle cell contractility. Marfan syndrome, Loeys–Dietz syndrome, and nonsyndromic HTAD demonstrate how genetic mutations can disrupt the components that stabilize the aortic wall. These pathogenic mechanisms influence matrix organization, intracellular signaling, and the contractile machinery within the mechanically stressed proximal aorta. In this review, we summarize current mechanistic insights into the major forms of HTAD and discuss how new molecular and cellular concepts could influence surveillance, genetic counseling, and genotype-guided therapeutic strategies.

Cells doi: 10.3390/cells15100934

Authors: Hua Teng Chao Li Mingjin Wang Jing Zhang Yi Zhang Michael Chopp Zheng Gang Zhang

Cerebrolysin has a salutary effect on impaired cerebral endothelial cell (CEC) permeability. Using an in vitro endothelial permeability assay, the present study tested the hypothesis that exosomes released by Cerebrolysin-treated CECs (Cerebro-Exos) have a robust therapeutic effect on dysfunctional CECs. Stoichiometric analysis showed marked differences in cargo profiles between Cerebro-Exos and exosomes derived from CECs without Cerebrolysin treatment (Naïve-Exos), in which Cerebro-Exos were highly enriched with metabolic and tight junction related proteins compared to Naïve-Exos. Cerebro-Exos had a superior effect compared to Naïve-Exos on restoring CEC integrity impaired by fibrin and tissue plasminogen activator (tPA). Treatment of fibrin- and tPA-challenged CECs with Cerebro-Exos robustly reduced fibrin- and tPA-augmented proteins involved in inflammation and coagulation and substantially increased fibrin- and tPA-decreased proteins that are related to tight junctions and metabolism. Collectively, these data indicate that Cerebro-Exos have a broad effect on improvement of dysfunctional CECs, which is likely achieved by the alteration of CEC proteins.

Cells doi: 10.3390/cells15100933

Authors: Matías Alfonso Rubio Eduardo Velásquez Sofia Antonucci María José Sánchez Carmen Romero

Epithelial ovarian cancer (EOC) remains a lethal malignancy requiring novel therapeutic strategies due to high recurrence and chemoresistance. This study evaluated the combined antitumor effect of metformin and the co-transfection of tumor-suppressor microRNAs miR-145 and miR-23b in A2780 and OV90 EOC cell lines using both 2D and 3D models. In monolayer cultures, our approach significantly reduced the expression of proliferation markers Ki-67 and c-MYC, and decreased cell migration and invasion in both cell lines compared to controls. In 3D spheroid models, the treatment reduced VEGF secretion and relative spheroid area in A2780 cells, significantly increasing cytotoxicity; however, OV90 spheroids exhibited marked resistance. Fluorescent miRNA tracking revealed that this resistance occurs despite successful intracellular delivery, indicating an intrinsic biological resistance conferred by the 3D microenvironment. Overall, these findings suggest that the combined administration of metformin and miRs effectively limits tumor progression, but also strongly underscore the importance of using complex 3D models to accurately evaluate therapeutic efficacy and intrinsic resistance mechanisms.

Cells doi: 10.3390/cells15100932

Authors: Jianan Wu Xia Yi Lanlan Wang Kaixuan Yang Minghuan Liu Jiawei Song Zenghui Yue

Atherosclerosis (AS) is a chronic inflammatory disease of large arteries. It underlies many cardiovascular disorders, including coronary artery disease, myocardial infarction, stroke, and peripheral arterial disease. Current therapies have improved outcomes, especially lipid-lowering, antithrombotic, and anti-inflammatory treatments. Yet residual cardiovascular risk remains, and new molecular targets are still needed. Leucine-rich α-2-glycoprotein 1 (LRG1) is an inflammation-inducible secreted glycoprotein. It has drawn attention because it is linked to pathological angiogenesis, vascular dysfunction, tissue remodeling, and fibrosis. Recent studies indicate that LRG1 is related to AS at several levels. These include circulating clinical associations, plaque localization, and experimental models. In AS, LRG1 may not simply act as a biomarker. It may promote macrophage pro-inflammatory polarization, disturb endothelial homeostasis, support abnormal angiogenesis, and influence extracellular matrix remodeling and plaque structural change. This review examines the biological features of LRG1 and the current evidence connecting it with AS. It also discusses possible mechanisms, therapeutic feasibility, and current limitations. Overall, LRG1 appears to be a promising but still incompletely validated candidate target in AS.

Cells doi: 10.3390/cells15100931

Authors: Atefeh Namipashaki Hanchen Yu Mark A. Bellgrove Ziarih Hawi

Attention deficit hyperactivity disorder (ADHD) is a common and highly heterogeneous neurodevelopmental condition with complex biological underpinnings. Despite substantial progress in identifying genetic and neurobiological correlates, the cellular mechanisms linking genetic variation to functional brain alterations remain poorly understood. Human induced pluripotent stem cell (iPSC) technology provides a powerful platform to investigate these mechanisms by enabling the generation of patient-specific neural cell types and the direct interrogation of molecular, cellular, and network-level phenotypes. In this review, we summarise the current understanding of the neurobiological mechanisms underlying ADHD, including dopaminergic dysregulation, delayed neurodevelopmental maturation, and excitatory/inhibitory imbalance. We then discuss how iPSC-based models, combined with genome engineering and advanced functional assays, can be used to dissect gene-specific effects, study neural circuit development, and establish scalable platforms for therapeutic discovery. Finally, we outline key methodological considerations for designing robust iPSC-based models of ADHD. Together, these approaches provide new opportunities to bridge genetic risk with cellular function and accelerate the development of mechanistically informed therapeutic strategies.

Cells doi: 10.3390/cells15100930

Authors: Haseeb Ahmad Faizan Masood Uzair Iqbal Mohamed Shaltout Yunus Yukselten Richard E. Sutton

Protein–protein interactions (PPIs) are fundamental to viral replication, regulating processes such as assembly, genome packaging, and virion maturation. Despite their biological importance, these interactions remain challenging to study and are relatively underexploited as therapeutic targets. Split reporter systems, based on protein-fragment complementation, provide quantitative platforms to measure PPIs by reconstituting reporter activity when interacting protein partners are brought into proximity. These systems can be applied in vitro and in live cells which enables detection of dynamic and multimeric interactions in physiologically relevant contexts. Major classes of split reporter systems include β-lactamase, alkaline phosphatase, luciferase-based platforms, green fluorescent protein, and horseradish peroxidase. Assay performance depends on factors such as fusion protein stability, expression levels, and reporter kinetics, which influence sensitivity, dynamic range, and reliability. These approaches have been applied to study viral protein interactions across diverse systems, including HIV-1 matrix and nucleocapsid proteins, flaviviral capsid proteins, hepatitis B virus core protein, and chikungunya virus capsid. Split reporter assays also enable high-throughput screening for small-molecule inhibitors that disrupt viral PPIs and multimerization. This provides a functional readout linked to viral replication. Despite the challenges that exist in assay optimization and protein stability, the sensitivity and versatility of these systems provide a framework to interrogate viral protein interactions and support the development of antiviral therapeutics.:

Cells doi: 10.3390/cells15100924

Authors: Fernando Riback da Silva Pedro Rafael Firmino Dias Isadora Carolina Betim Pavan Andressa Peres de Oliveira Fernanda Luisa Basei Leticia Ester dos Santos Lizandra Maia de Sousa Sílvio Roberto Consonni André Gustavo de Oliveira Leonardo Reis Silveira Jörg Kobarg

The family of Nek kinases has 11 human members that are conserved in their kinase domains but diverse in their regulatory domains. Functionally, they can be associated with diverse aspects of cell cycle regulation, from mitosis and primary cilia function to centrosome disjunction in the G2 phase and checkpoints of the DNA damage response. However, novel functional contexts have emerged in recent years, including regulatory roles of Neks 1, 4, 5, and 10 in mitochondrial metabolic and morphological homeostasis. We recently generated, by CRISPR-Cas9 technology, a DU-145 prostate cancer cell line, with an NEK6 gene knockout. Here, we focus on a detailed characterization of changes in this cell line, in mitochondrial respiration function and morphology. DU-145 NEK6 knockout cells exhibited reduced mitochondrial respiration and a fragmented phenotype in electron microscopy, with reduced mitochondrial cristae numbers. Alterations in mitochondrial architecture and respiration were correlated with increased expression of anaerobic glycolytic proteins (HK2, PFKP, and LDHA) and decreased expression of PDH, an enzyme of aerobic glycolysis. Molecular analysis by Western blot revealed decreased levels of mitochondrial mass and biogenesis protein markers (TOM20, TFAM), without alterations in other markers such as VDAC1/3 or mtDNA copy number in the NEK6 knockout cells. Furthermore, the regulators of mitochondrial fusion/fission are altered in the knockout cells (decrease in the Long-OPA1:Short-OPA1 ratio and DRP1 total level), which is associated with an increase in endoplasmic reticulum–mitochondria contact at ≤20 nm observed in transmission electron microscopy (TEM) image analysis. Using analysis of TEM micrographs, we found an increase in the autophagic structures (autophagosome, amphisome, and autolysosome), with mitochondria as cargo in some structures, which was correlated with a decrease in LC3A/B and an increase in the BECLIN1 total level, and with an increase in acidic vesicles approximation, suggesting that reduction in TOM20 and TFAM without alterations in VDAC1/3 and mtDNA copy number might be related to mitochondrial degradation through autophagy. Together, our data suggest a new role for NEK6 in regulating mitochondrial homeostasis, where its loss alters mitochondrial morphology and respiration, and could be associated with an increase in the degradation of the dysfunctional mitochondria through autophagy.

Cells doi: 10.3390/cells15100929

Authors: Xinyi Peng Genglin Guo Songyu Li Songyao Sun Cong Ouyang Jiajun Cui

Inflammatory bowel diseases, particularly ulcerative colitis (UC), are chronic relapsing inflammatory disorders with limited therapeutic options. The zinc-finger transcription factor ZBTB4 has been implicated in the initiation and progression of cancer, but its role in UC remains unknown. Here, we found that ZBTB4 deficiency exacerbates dextran sulfate sodium (DSS)-induced colitis in C57BL/6J male mice. Compared with the wild type, ZBTB4 deficiency increases weight loss, colon shortening and proinflammatory cytokine production. RNA-seq analysis revealed that ZBTB4 deficiency enhances Serpine1 expression and activates the NF-κB pathway. NF-κB inhibition by JSH-23 alleviated the effect of ZBTB4 deficiency on DSS-induced colitis. These results imply the protective role of ZBTB4 in UC. Through an integrated drug screening, we identified a natural sesquiterpene lactone, handelin, as a potential compound to enhance ZBTB4 expression in NCM460 cells. Handelin administration relieved colitis in wild-type mice but produced no effect in ZBTB4 knockout mice, demonstrating that its anti-colitic effect depends on ZBTB4 expression. Collectively, our results indicate the key role of ZBTB4 in UC and ZBTB4 agonists may serve as a novel approach for UC treatments.

Cells doi: 10.3390/cells15100928

Authors: Masood Sepehrimanesh Sarah Victoria Melen Fatima Yeasmin Victor Adeleke Ojo Francisca Walden Humaira Urmee Jenna Etheridge Aruna Kumari Nasu

Neurodegenerative diseases, including Alzheimer’s disease (AD), Parkinson’s disease (PD), and amyotrophic lateral sclerosis (ALS; Lou Gehrig’s disease), represent a growing global health burden characterized by progressive neuronal loss and functional decline. Despite decades of intensive research, effective disease-modifying therapies remain limited, underscoring the urgent need for innovative therapeutic strategies. This review highlights recent advances in the understanding of disease etiology and emerging treatment approaches, with a particular focus on modalities with translational potential. We discussed novel disease-modifying interventions, including gene and cell therapies, RNA-targeting strategies, and immunotherapies aimed at clearing misfolded proteins such as amyloid-β, tau, and α-synuclein. In parallel, we examined the evolving recognition of neuroinflammation and mitochondrial dysfunction as actionable therapeutic targets, alongside progress in precision medicine and biomarker-guided approaches that enable early diagnosis and individualized treatment. Additionally, we summarized developments in repurposed pharmacological agents, neuroprotective compounds, and lifestyle interventions, emphasizing the importance of integrative, multimodal strategies. Across AD, PD, and ALS, convergent molecular mechanisms, including protein misfolding, oxidative stress, and disrupted proteostasis, present opportunities for cross-disease therapeutic targeting. Finally, we addressed key challenges and future directions, including translating preclinical efficacy into clinical success, optimizing CNS-targeted delivery systems, and navigating ethical considerations surrounding gene editing and stem cell therapies.

Cells doi: 10.3390/cells15100927

Authors: Arianna S. Gholami Ciara N. Walsh Jean McBryan

Pleiotrophin (PTN), a heparin-binding growth factor with potent mitogenic and angiogenic activity, has emerged as a key regulator of mammary gland biology and a potential driver of breast cancer progression. This review integrates current evidence on PTN’s roles from normal mammary development, where it can delay ductal outgrowth, to triple negative breast cancer, where it promotes lung metastasis and correlates with poor survival. Though frequently reported as being overexpressed in breast cancer, the published data indicates that PTN transcription is reduced in cancer relative to normal breast cells. By contrast, serum PTN protein levels have been shown by multiple studies to be elevated in breast cancer patients relative to healthy controls. We examine the expression and function of PTN at a cellular level and explore the interplay between PTN and the tumour microenvironment. We evaluate preclinical models, clinical correlations, and emerging biomarker data that position PTN as a candidate prognostic indicator and therapeutic target. Despite growing interest, significant gaps remain regarding context-specific signalling. By integrating developmental and oncogenic perspectives, this review highlights PTN as a pivotal but underexplored factor in mammary gland physiology and breast cancer and outlines future research directions needed to translate PTN-targeted strategies into clinical benefit.

Cells doi: 10.3390/cells15100926

Authors: Lara Atzgerstorfer Magdalena Prantl Andrea Waldhauser Isabella Derler Marc Fahrner

Store-operated Calcium (Ca2+) entry (SOCE), mediated by stromal interaction molecule 1 (STIM1) and Orai1, is a central pathway controlling intracellular Ca2+ homeostasis. Gain-of-function (GoF) mutations in STIM1 cause a spectrum of clinically overlapping disorders historically classified as Stormorken Syndrome (STK), tubular aggregate myopathy (TAM), and York Platelet Syndrome (YPS). However, increasing evidence indicates that these entities could represent a shared disease spectrum rather than distinct conditions. At the molecular level, STIM1 activation is governed by a series of autoinhibitory checkpoints that maintain the protein in a tightly controlled resting state. GoF mutations disrupt these regulatory constraints, leading to dysregulated SOCE activity that is frequently, but not uniformly, associated with constitutive channel activation depending on the specific mutation and cellular context. While many disease-associated variants localize to the EF hand, a highly conserved helix–loop–helix Ca2+ binding motif, and the CC1 (coiled-coil 1) domain involved in molecular regulation of STIM1 activation, an increasing number of mutations in the C-terminal region further expands the mechanistic and clinical spectrum. In this review, we summarize current concepts of molecular STIM1 activation and discuss how distinct mutations perturb specific regulatory elements of the protein. By systematically integrating published case reports into a comprehensive overview, including a mutation–phenotype correlation table, we highlight the remarkable variability in and incomplete penetrance of clinical manifestations.

Cells doi: 10.3390/cells15100925

Authors: Yosuk Min Donghyeon Kim Hong-Beom Park Hae-Seul Choi Sohyun Hwang Kwang-Hyun Baek

Epithelial ovarian cancer (EOC) poses a challenge owing to its high rate of recurrence and drug resistance, resulting in a 5-year survival rate of 30% in advanced stages. To elucidate the molecular mechanisms underlying EOC recurrence, we analyzed transcriptome data from patients with EOC and identified elevated USP19, a deubiquitinating enzyme, as being elevated in patients without recurrence in our previous study. Single-cell RNA sequencing analysis revealed that increased USP19 expression in epithelial cells is associated with activation of apoptotic pathways, suggesting that USP19 may inhibit EOC recurrence through deubiquitination of its binding proteins. Using the protein–protein interaction database, we identified DnaJC7 as a binding partner of USP19 and confirmed their interaction experimentally. USP19-mediated deubiquitination of DnaJC7 increases its protein stability. Notably, upregulation of USP19 and DnaJC7 disrupted the interaction between p53 and MDM2, and knockdown of USP19 and DnaJC7 resulted in decreased p53 expression following cisplatin treatment. These findings highlight the therapeutic potential of enhancing the USP19-DnaJC7 axis to stabilize p53 and improve cisplatin efficacy. Promoting USP19-mediated deubiquitination by stabilizing DnaJC7 may offer a novel combination strategy to enhance the efficacy of cisplatin-based cancer therapy.

Cells doi: 10.3390/cells15100922

Authors: Jemima Sani Bin Yi Yaguang Xi

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with limited effective therapies. Two-dimensional (2D) in vitro models poorly recapitulate tumor microenvironment (TME) interactions, impeding the translational relevance of TNBC immunotherapy research. Three-dimensional patient-derived tumor organoids (3D PDTOs) have emerged as advanced preclinical models that better mimic tumor–immune interactions. The objective of this systematic review was to assess the landscape of 3D PDTO adoption in TNBC research and evaluate their application in addressing key bottlenecks in TNBC immunotherapy. We retrieved 394 studies published between 2015 and 2025 from the PubMed and ClinicalTrials.gov databases. Of those, 153 studies were included in the review. Fifty-eight (58) TNBC-specific studies met the inclusion criteria, including explicit mention of 3D PDTOs in the title or abstract, with confirmation in the Methods and Results sections. Studies were excluded if they used non-patient-derived tumor organoids or referred to other 3D models as 3D PDTOs. Data were collected from January 2025 through December 2025. Eligible studies were screened in three (3) tiers, grouped by relevant themes and graphed in Excel. We present an overview of the adoption of 3D PDTOs in TNBC research, highlighting the most common application trends within this scope. We also discuss the potential impact of artificial intelligence (AI) and regulatory guidance from the United States Food and Drug Administration (FDA) and the European Medicines Agency (EMA) pertinent to the adoption of organoids as human-relevant models to improve translational outcomes. Overall, this review provides actionable insights for leveraging 3D PDTOs to advance translational TNBC research and precision oncology.

Cells doi: 10.3390/cells15100923

Authors: Saad A. Farooqui Maryline Santerre Natalia Shcherbik Bassel E. Sawaya

Viral infections of the central nervous system produce memory impairment through mechanisms that extend beyond acute neuronal injury. Herpes simplex virus type 1, human immunodeficiency virus, varicella zoster virus, cytomegalovirus, Epstein–Barr virus, influenza, SARS-CoV-2, West Nile virus, and Zika virus each enter or engage the brain through distinct routes, yet converge on four shared molecular pathways that selectively damage hippocampal circuits: mitochondria-associated membrane (MAM) dysfunction, chronic neuroinflammation, blood–brain barrier (BBB) disruption, and impaired CREB-BDNF signaling. These pathways specifically compromise the dentate gyrus, CA3, and CA1 subfields, producing predictable deficits in pattern separation, associative retrieval, and temporal memory binding. Antiretroviral and antiviral therapies suppress viral replication but fail to reverse organelle-level dysfunction, leaving most hippocampal injury unaddressed. Emerging plasma biomarkers, p-tau217, neurofilament light chain, and GFAP, combined with hippocampal subfield MRI, now enable mechanistic stratification before irreversible circuit loss occurs. This review proposes, as a unifying hypothesis, that virus-associated memory impairment represents a convergent hippocampal syndrome driven by shared downstream pathways, and that combination therapies targeting these pathways simultaneously offer greater therapeutic promise than pathogen-specific approaches alone. The evidentiary basis for this framework varies across pathogens and conditions; direct mechanistic evidence, mechanistic analogy, and preclinical data are distinguished throughout.

Cells doi: 10.3390/cells15100921

Authors: Ludivine Renaud Samantha Kotz Aravind Menon Carol Feghali-Bostwick

Background: Systemic sclerosis (SSc) is a systemic autoimmune disease leading to extensive fibrosis of the skin and many visceral organs, including the lungs. The need for effective treatments is urgent, as none exists today that can stop or reverse the progression of fibrosis. We examined the effect of restoring KLF4 levels in primary human lung fibroblasts of SSc patients with pulmonary fibrosis as a potential therapeutic strategy. Methods: We restored KLF4 levels in SSc lung fibroblasts by adenoviral infection and extracted total RNA for RNA sequencing. Genes and systems level analyses were performed, and selected genes of interest and markers of alveolar, inflammatory, and fibrotic fibroblasts were further validated at the mRNA and protein levels. Results: Our results showed that restoring KLF4 levels in SSc lung fibroblasts initiated dedifferentiation of αSMA and CTHRC1, expressing myofibroblasts by repressing markers of inflammatory and fibrotic fibroblasts while boosting markers of alveolar fibroblasts. Our data also revealed that restoring KLF4 levels prevented TGFβ1-induced fibrogenesis in normal lung fibroblasts, and reduced fibrosis in explanted human lungs in organ culture. Conclusions: Our results in human primary SSc lung fibroblasts showed that restoring KLF4 levels initiated dedifferentiation of fibrotic and inflammatory fibroblasts towards the phenotype of alveolar fibroblasts, their lineage precursors, highlighting the potential of KLF4 as a therapy to stop and reverse fibrosis.

Cells doi: 10.3390/cells15100920

Authors: Qun Chen Zachary Kiernan Gina Labate Oluwatoyin Akande Edward J. Lesnefsky Mohammed Quader

Donation after circulatory death (DCD) involves unavoidable ischemia–reperfusion injury (IRI). Mitochondrial permeability transition pore (MPTP) opening plays a critical role in DCD heart injury. Activation of ubiquitous calpains, including calpain-1 and calpain-2 (CPN1/2), increases MPTP opening in DCD hearts. Mitofilin, a mitochondrial inner membrane protein that regulates cristae morphology, is also involved in MPTP opening during ischemia–reperfusion. However, it remains unclear whether CPN1/2 activation contributes to mitofilin-mediated IRI in DCD hearts. We first incubated a mitofilin peptide with exogenous CPN1 in vitro to investigate the link between CPN1 activation and mitofilin degradation. Next, we tested whether CPN1/2 inhibition reduces cardiac injury in DCD hearts by preserving mitofilin and limiting MPTP opening. Sprague-Dawley (SD) rat hearts were subjected to 25 min of in vivo ischemia followed by ex vivo perfusion with or without the CPN1/2 inhibitor MDL-28170 (10 µM). In vitro incubation with CPN1 led to mitofilin degradation, confirming mitofilin as a CPN1 substrate. CPN1/2 inhibition significantly reduced infarct size compared with untreated DCD hearts, preserved mitofilin expression, and decreased MPTP opening. These findings indicate that CPN1/2 activation promotes MPTP opening in DCD hearts through mitofilin degradation. Timely inhibition of CPN1/2 represents a promising strategy to reduce cardiac injury and improve DCD heart function.

Cells doi: 10.3390/cells15100919

Authors: Mafalda Meunier Ana Valério-Bolas Armanda Rodrigues Flávia Fróis-Martins Rui Ferreira Inês Cardoso Marta Monteiro Joana Palma-Marques Manuela Carvalheiro Telmo Nunes Wilson T. Antunes Graça Alexandre-Pires Isabel Pereira da Fonseca Gabriela Santos-Gomes

Leishmaniasis caused by Leishmania infantum is a zoonotic disease endemic in many regions worldwide. The antigen-presenting dendritic cells (DCs) bridge the innate and adaptive immune response by activating T lymphocytes. Therefore, the present study examines whether T lymphocyte activation can be directed by autologous DCs primed by extracellular vesicles (EVs) derived from L. infantum. For this, lymphocytes were co-cultured with monocyte-derived DCs (moDCs) that were primed by EVs. moDC signaling and activation were examined by gene expression of toll-like receptors and cytokines. The antigen-presentation ability was analyzed through major histocompatibility complex molecules, and T cell subpopulations were explored by immunophenotyping. In co-cultures, EV-primed moDCs upregulated TLR2, TLR4, and TLR9, along with overexpression of MHC molecules. Co-cultures involving moDCs primed by EVs promoted the upregulation of both pro-inflammatory and regulatory cytokines associated with the expansion of non-conventional regulatory and central memory T cell subsets within the CD8+ T cell subpopulation. These findings suggest that activated moDCs can modulate cytotoxic lymphocytes, thereby promoting a balanced inflammatory microenvironment counterbalanced by a concurrent regulatory immune response. Thus, cell-based immune strategies using moDCs loaded with Leishmania-derived EVs represent a potential first step toward the development of innovative and personalized immune prophylactic and therapeutic approaches for leishmaniasis.

Cells doi: 10.3390/cells15100918

Authors: Mingxi Jia Jingjing Sun Xiuli Yang Yue Cui Zixuan He Haixia Han

Natural bioactive compounds present promising avenues for the prevention and therapeutic intervention of cancer. Octacosanol has garnered significant attention for its distinctive biological properties, yet its specific antitumor effects and underlying mechanisms remain unclear. This study systematically evaluated its antitumor effects and elucidated the associated molecular mechanisms. We confirmed that it dose-dependently inhibited A549 cell proliferation in vitro. It also remarkably suppressed cell invasion and migration by downregulating MMP2 and MMP9 expression, an effect that was associated with reduced phosphorylation of JAK3/STAT3 and PI3K/AKT, suggesting a potential regulatory role of these signalling cascades. Meanwhile, it significantly inhibited tumor cell VEGF secretion and VEGF-mediated neoangiogenesis by modulating the PI3K/AKT signaling axis. Mouse experiments demonstrated that octacosanol significantly reduced tumor p-AKT, MMP2, and MMP9 levels, indicating its in vivo anti-metastatic effect. It also remarkably decreased tumor microvessel density, alongside reduced VEGF and vascular endothelial marker CD31 expression, further verifying its potent anti-angiogenic activity. This work provides evidence of octacosanol’s dual anti-metastatic and anti-angiogenic effects in lung cancer and offers novel mechanistic insights into its activity against this highly prevalent malignancy. These findings establish a solid foundation for further exploration and development of octacosanol as a promising adjuvant for clinical antitumor therapy.

Cells doi: 10.3390/cells15100917

Authors: Kylee A. Hunter Anne-Marie C. Overstreet Bryon Benjamin Koff Hridai Dharan Steven Overend Jeannette S. Messer

Inflammatory bowel diseases (IBDs) are diseases of chronic inflammation and intestinal epithelial cell (IEC) death that affect an estimated 7 million people worldwide. Intestinal barrier restoration is the most important determinant of remission in IBD, yet there are very few existing therapies that protect IECs from damage or support epithelial repair. The goal of this study was to develop a model system and tools that can be used to identify therapeutics that promote IEC survival in IBD. We developed a Beclin-1 cleavage reporter (BICR) that detects calpain-mediated Beclin-1 cleavage and the switch from autophagy to programmed cell death. We modified BICR with the HIV Tat peptide (BICR-Tat) and tested it in a model of live bacterial stress using commensal E. coli and IEC. BICR sensitively and specifically detected calpain activity in cell-free assays, and BICR-Tat successfully detected Beclin-1 cleavage and autophagy failure in IEC. Achieving IEC survival in the microbe-challenged IBD gut would be an important advance toward intestinal barrier restoration in this intractable disease. The BICR-Tat reporter coupled with the model of microbial stress developed in this study could enable high-throughput screening approaches to identify therapeutics with the potential to achieve barrier healing and sustained remission in IBD.

Cells doi: 10.3390/cells15100916

Authors: Georgia R. Layton Riyaz Somani Giovanni Mariscalco Farooq Donoo G. André Ng Ibrahim Antoun Mustafa Zakkar

Coronary artery bypass grafting (CABG) using the autologous saphenous vein (SV) remains widely performed for obstructive atherosclerosis; however, vein graft disease drives recurrent ischaemia through early thrombosis and progressive intimal hyperplasia, and accelerated atherosclerosis developing within the grafts. Extracellular vesicles (EVs) are membrane-bound particles that transfer proteins, lipids, and microRNAs between cells. They modulate endothelial dysfunction, vascular smooth muscle cell phenotypic switching, inflammation, and coagulation, which are core processes in vein graft remodelling. Arterialisation exposes the vein to abrupt rises in shear stress, cyclic stretch, and intraluminal pressure. These forces increase EV release and reshape EV cargo in experimental systems, suggesting a potential mechanism for amplifying early graft injury which warrants direct investigation in vein tissue. This review synthesises current evidence for cell-specific EV contributions from ECs, vascular smooth muscle cells, platelets, and macrophages, and appraises EV-associated microRNAs with biomarker potential relevant to graft failure pathways. We also review therapeutic strategies that may modulate EV signalling including antiplatelet therapy, statins, KCa3.1 inhibition, and pro-reparative mesenchymal stromal cell-derived EVs. No published clinical studies evaluate EV-based biomarkers specifically for saphenous vein graft patency, and none prospectively predict saphenous graft failure. CABG provides a well-defined time zero event that enables longitudinal sampling and risk stratification. Prospective studies linking EV phenotypes and miRNA signatures to imaging-defined graft outcomes are needed to support clinical translation.

Cells doi: 10.3390/cells15100914

Authors: Kai Quan Hongxia Yang Guangtian Tang Ziwei Li Hailin Zou Jing Ma Xinsheng Yao

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by immune dysregulation and multi-organ damage. Abnormal B cell activation and autoantibody production constitute the core pathological mechanism of SLE. However, the proportion, BCR pairing types, clonal evolution patterns, and transcriptomic features of dual BCR B cells in SLE remain incompletely elucidated. In this study, we employed single-cell RNA sequencing (scRNA-seq) combined with single-cell B cell receptor repertoire sequencing (scBCR-seq) to preliminarily analyze the proportion and characteristics of dual BCR B cells in SLE model mice (MRL/Lpr and SLE.Yaa) as well as in peripheral blood from SLE patients. The results showed: (1) Compared with control groups, the proportion of dual BCR B cells in SLE model mice and patients exhibited a decreasing trend, whereas the diversity of the CDR3 repertoire decreased and clonality increased. Increased clonal sharing was observed between single BCR B cells and dual BCR B cells. The main pairing types of dual BCR B cells were H + κ1 + κ2, H1 + H2 + κ, and H1 + H2 + κ + λ, with preferential utilization of autoimmunity-associated V gene families such as IGHV4-34, and high expression of IGHG subtypes. (2) Tracking analysis of B cell receptor clonality and effector molecule expression revealed that in SLE, dual BCR B cells tend to enrich in IFN-α/γ responses, TNF-NFκB inflammation, and complement pathways, and highly express interferon-related genes such as Ly6a, Isg15, MX1, and IFI6. (3) In both single BCR B and dual BCR B cells from SLE patients, the proportion of the naïve B cell subset decreased, whereas the proportions of plasma and Breg subsets increased and exhibited clonal expansion. SLE dual BCR Breg cells highly expressed IL10, HSPA1A, and others. This study is the first to reveal, at the high-throughput single-B-cell level, that the proportion, subset origin distribution, CDR3 repertoire composition, and effector molecule expression of dual BCR B cells display unique characteristics in SLE model mice and patients, providing baseline comparative data and novel research perspectives for further investigation into B cell effector functions and mechanisms in SLE patients.

Cells doi: 10.3390/cells15100915

Authors: Ştefan Agoston Alina Grama Alexia Onaciu Alexandra Mititelu Gabriel Benţa Tudor Lucian Pop

Autoimmune hepatitis (AIH) is a progressive inflammatory liver disease characterized by hypergammaglobulinemia, circulating antibodies, and distinctive histological features, with a higher prevalence in females. Immune responses targeting hepatic antigens are considered the main mechanism behind AIH. Many cytokines are involved in the inflammatory response typical of this disease. Interleukin 17 (IL-17) is a powerful pro-inflammatory protein that serves as a key link between the innate and adaptive immune systems. It plays an important role in regulating the inflammatory response in various tissues, including the liver. Several studies have shown that increased IL-17 levels are associated with the severity and progression of AIH. This review explores IL-17’s role in the AIH inflammatory pathway and summarizes existing evidence linking it to liver damage. We also highlight the potential of future therapies targeting this cytokine.

Cells doi: 10.3390/cells15100913

Authors: Anna Makowska Udo Kontny

Natural killer (NK) cells are increasingly recognized as a complementary platform to T-cell-based cancer immunotherapies. Their innate, MHC-unrestricted recognition, capacity to mediate antibody-dependent cellular cytotoxicity (ADCC) and comparatively favorable toxicity profile have given rise to a broad therapeutic pipeline that includes cytokine-supported regimens, adoptive NK products, bispecific and trispecific NK engagers, and chimeric antigen receptor (CAR)-engineered NK cells. Clinical data, particularly in hematologic malignancies, show that NK-cell-based strategies can be safe and biologically active, although limited persistence, suboptimal trafficking and immune escape remain key challenges. Nasopharyngeal carcinoma (NPC), an Epstein–Barr virus (EBV)-driven epithelial cancer, illustrates how a tumor microenvironment (TME) can simultaneously impair NK function and create specific vulnerabilities that NK-focused therapies can exploit. This review summarizes NK biology and current therapeutic platforms, analyzes major limitations, highlights the specific context of NK-cell-based strategies in NPC and compares NK- and T-cell-based therapies with an emphasis on clinical translation.

Cells doi: 10.3390/cells15100910

Authors: Nakyung Oh Van Ngu Trinh

Cancer stem cells (CSCs) are a distinct subpopulation within a tumor that play an important role in tumor initiation, metastasis, therapeutic resistance, and cancer relapse. Their persistence is strongly influenced by the tumor microenvironment (TME), which provides a range of biological signals that maintain stemness, promote immune evasion, and resistance to cancer treatment. Therefore, effective targeting of CSCs is essential to improve therapeutic efficacy. In this review, we summarize the key characteristics of CSCs and their niche within the TME, emphasizing their interactions with immune cells, stromal components, and secreted factors. We also discuss the major challenges in targeting CSCs, including immune evasion, metabolic constraints, and intratumoral heterogeneity. We further highlight current and emerging immunotherapeutic strategies targeting CSCs, including immune checkpoint inhibitors, cancer vaccines, monoclonal antibodies, nanobodies, bispecific antibodies, antibody-drug conjugates (ADCs), CAR-T and CAR-NK cell therapies, oncolytic viruses, as well as innovative approaches such as targeted protein degradation. Finally, we emphasize the importance of a combinatorial approach that integrates CSCs targeting with modulation of the TME. Together, these strategies may lead to more durable responses, enhance therapy efficacy and reduce the risk of tumor recurrence.

Cells doi: 10.3390/cells15100912

Authors: Yunwei Shi Hong Wang Nur Salihah Lau Amanda Tan Ying Xin Caiping Wang Feng Ru Tang

Prenatal exposure to ionizing radiation is a known risk factor for neurodevelopmental deficits; however, the molecular mechanisms linking chronic embryonic insult to abnormal brain development remain poorly understood. This study investigated the long-term consequences of chronic prenatal gamma irradiation throughout gestation in C57BL/6 mice. Behavioural analysis of adult offspring revealed a specific increase in depression-like behaviours, with no significant alterations in anxiety or general exploratory activity. Immunohistochemical assessment demonstrated a significant reduction in adult hippocampal neurogenesis, marked by decreased doublecortin (DCX)-positive newborn neurons in the subgranular zone and fewer NeuN-positive mature neurons in the dentate gyrus hilus. Integrated RNA-seq, qPCR, and Western blot analyses implicated the upregulation of the Mef2c/Egr1 signalling pathway in this neurogenic deficit. Furthermore, miRNA sequencing identified a pronounced decrease in miR-1843a-3p, which was subsequently validated to directly target Mef2c. Collectively, these findings suggest that prenatal gamma irradiation disrupts neurogenic processes and adult brain function, leading to specific behavioral abnormalities. This long-term impairment is associated with, and may be at least partially mediated by, dysregulation of the miR-1843a-3p/Mef2c/Egr1 pathway.

Cells doi: 10.3390/cells15100911

Authors: Rikuto Fujita Takashi Ishino Takashi Oda Tomohiro Kawasumi Manabu Nishida Yuichiro Horibe Nobuyuki Chikuie Takayuki Taruya Takao Hamamoto Tsutomu Ueda Sachio Takeno

Background: Eosinophilic chronic rhinosinusitis (ECRS) is characterized by refractory nasal polyps and severely impaired mucociliary clearance (MCC). The molecular mechanisms underlying the modulation of mucociliogenesis following IL-4/13 blockade with dupilumab remain poorly understood, notwithstanding its proven clinical efficacy. Methods: Bulk RNA Barcoding and sequencing (BRB-seq) was performed on nasal polyp tissues collected from healthy controls (n = 6), patients with non-ECRS (n = 8), and patients with ECRS both before and four weeks after dupilumab treatment (n = 9) to identify the early molecular drivers underlying ciliary regeneration. Comprehensive gene-set scoring systems were developed to evaluate multiciliogenesis master regulators, master regulators of core/ciliary planar cell polarity (PCP) and PCP components. Interaction scores for epithelial-derived cytokines—thymic stromal lymphopoietin (TSLP), IL-25, and IL-33—were calculated based on ligand and cognate receptor subunit expression. Results: The ciliary master regulatory hierarchy (e.g., FOXJ1, RFX2/3), PCP components (CELSR1 and the ciliogenesis and planar polarity effector (CPLANE) module: FUZ, INTU, WDPCP), and structural ciliogenesis pathways were robustly restored following IL-4/13 blockade. The TSLP interaction score correlated with global mucosal damage, serving as a trigger for compensatory multiciliogenesis. The pre-treatment IL-33 interaction score emerged as a significant predictor of transcriptomic ciliary recovery (p < 0.05). DNASE1L3—the primary endonuclease for degrading eosinophilic extracellular traps (EETs)—remained persistently downregulated post-treatment. Conclusions: IL-4/13 blockade successfully restores the structural and directional “hardware” of the respiratory epithelium but fails to rectify the enzymatic “software” required for mucus degradation. This “residual molecular scar” may explain the persistent mucus hyperviscosity observed in some ECRS patients even after clinical polyp resolution.

Cells doi: 10.3390/cells15100909

Authors: Patrícia A. António Joana R. Lérias Carolina M. Gorgulho Karina Balan Vitaly Balan Markus J. Maeurer

Adoptive cell therapy (ACT) using tumor-infiltrating lymphocytes (TILs) has achieved clinically and biologically relevant responses in patients with solid cancer. Clinical efficacy has been increasingly linked to a specific T-cell phenotype, particularly CD8+ TILs exhibiting a progenitor stem-cell-like profile (CD39− CD69−). This review explores the critical role of the sphingosine-1-phosphate (S1P) axis in orchestrating these responses. We detail the biological antagonism between the activation marker CD69 and S1P receptor 1 (S1PR1), where mutual exclusivity dictates thymic selection, if T-cells are retained in tissues or allowed to recirculate and maintain long-term immune surveillance. The S1PR1:S1P axis is further recognized as a critical regulator of mitochondrial fitness, sustaining the high energetic demands of precursor T-cells. We examine the “double-edged sword” nature of S1P in the tumor microenvironment (TME), where it can drive pro-tumorigenic processes like angiogenesis and vascular mimicry (VM), be hijacked by cancer cells to create immune-excluded environments, or S1P can increase T-cell fitness. We summarize the current landscape of clinical trials (as of January 2026) that target S1P production or signaling to modulate anti-tumor responses or use S1P as a biologically relevant marker of treatment outcome.

Cells doi: 10.3390/cells15100908

Authors: Ceng-Ceng Ge He-Xin He Ming-Yu Pei Shu-Qin Tang Wei He Man-Man Bian Ming Pan Si-Yang Huang

Toxoplasma gondii (T. gondi) is a widespread zoonotic parasite that poses a significant threat to global public health, yet effective therapeutic options remain limited. In this study, we found that the flavonoid compound myricetin (MYR) can significantly inhibit the proliferation of T. gondii. This effect is associated with the inhibition of dihydroorotase (TgDHO) activity in the de novo pyrimidine biosynthesis pathway, and this inhibition can be partially reversed by exogenous supplementation with uracil. Further studies revealed that MYR treatment can induce cell cycle arrest in tachyzoites and impair bradyzoite proliferation, concurrently disrupting the UDP-GlcNAc glycosylation of the cyst wall. In mouse models, MYR demonstrated significant efficacy, achieving an 80% survival rate in acute infection and inducing morphological abnormalities in intracerebral cysts during chronic infection. Collectively, these findings elucidate the anti-Toxoplasma activity and multifaceted mechanisms of MYR, providing valuable insights for developing novel therapeutics against toxoplasmosis.

Cells doi: 10.3390/cells15100907

Authors: Estera Bakinowska Wojciech Jerzy Biniek Kajetan Kiełbowski Kamil Dyrka Konrad Szewczyk Hanna Ostałowska Zuzanna Leciej Andrzej Pawlik

Diabetes mellitus is a chronic and progressive metabolic disorder associated with abnormal blood glucose levels. The term involves several diseases with different pathophysiology mechanisms and treatment strategies. Stem cell-based treatments represent an emerging strategy for patients with diabetes mellitus with severe pancreatic insufficiency and poor glycemic control. Over the last 20 years, researchers have investigated mesenchymal stem cell infusion and the transplantation of stem cell-derived β cells and islet tissues. This review aims to comprehensively discuss the latest advances in the field of stem cell use in diabetes, including clinical studies and preclinical experiments aiming at improving the efficacy and safety of stem cell use.

Cells doi: 10.3390/cells15100906

Authors: Ae Ra Kim Julia Jamka William F. Jackson Emma D. Flood Jonathon L. McClain Brian D. Gulbransen

Anticontractile factors secreted by perivascular adipose tissue (PVAT) play an important role in regulating vascular tone. This process is driven by the neurotransmitter norepinephrine (NE), but recent data show that adrenergic innervation in PVAT is sparse. How limited innervation might initiate broad responses through PVAT depots remains unknown. Here, we used Ca2+ imaging with genetically encoded sensors, selective drugs, immunolabeling and a conditional ablation model to test the hypothesis that gap junction coupling among PVAT adipocytes contributes to how signals initiated by NE are distributed through PVAT depots. Despite exhibiting differing sensitivities to NE, adipocytes in aortic and mesenteric PVAT and in white adipose tissue displayed robust expression of the gap junction protein connexin-43 (Cx43). Blocking gap junction coupling with the drug carbenoxolone (Cbx) limited NE-evoked Ca2+ responses among adipocytes, while blocking Cx43 hemichannels with the mimetic peptide 43Gap26 had no significant effect. Fluorescence recovery after photobleaching (FRAP) in mPVAT was decreased in the presence of Cbx, suggesting impaired gap junction communication. Wire myography recordings of mesenteric arteries showed that the EC50 for NE was higher in samples with intact PVAT than those without; however, this effect was not significantly different in samples from mice that lacked Cx43 in adipocytes. Analysis of multiple connexins showed that adipocytes upregulate Cx26 gene expression when Cx43 is deleted. These observations support the conclusion that Cx43-mediated gap junction coupling among PVAT adipocytes contributes to distributing signals initiated by NE; however, how this mechanism contributes to regulating vessel constriction remains unclear. This, and how potential compensatory mechanisms are enacted in adipocytes lacking Cx43, should be addressed in future work.

Cells doi: 10.3390/cells15100905

Authors: Dagmara Szkolnicka Lydia González del Barrio Carlos D. Quintana Calderón Justyna M. Kowal Shruthi Sampath Giles Dudley Joakim Sørensen Allan E. Karlsen David C. Hay

Annually, there are more than two million deaths from liver disease. This is driven by organ inflammation and scarring, leading to a decline in function and regeneration. Frequently, this can develop into decompensated liver disease, resulting in the loss of physiological balance and toxin build-up within the body, with an increased risk of patient mortality. Currently, there are no approved medicines for the long-term treatment of liver cirrhosis. The only successful treatment option for end-stage liver disease patients is donor organ transplantation. However, patient requirement outstrips the number of donated organs. To address this bottleneck, researchers around the world have developed cell-based prototype systems to restore failing liver function, with some in clinical trials. Although significant progress has been made, no mainstream commercial liver assist products are available for routine clinical use. In this study we developed a stem cell-derived vascularized liver tissue implant prototype from pluripotent cells. The liver tissue was produced from a stem cell line that is banked at clinical grade, and displayed stable and mature liver function over a 6-week period in vitro. This included decreasing levels of the fetal marker, alpha-fetoprotein, when the serum albumin increased. This was further supported by stable alpha-1-antitrypsin secretion and cytochrome P450 function. Following the establishment of stable liver tissue, it was delivered as a cell product or attached to an electrospun polycaprolactone scaffold, to form a tissue implant. Next, cellular material was quality-controlled, and subsequently shipped to a contract research organization for external in vivo validation. The transplanted liver tissue functioned when implanted into the kidney capsule and subcutaneously, remaining functional for up to two weeks in vivo.

Cells doi: 10.3390/cells15100904

Authors: Maria Latiano Maria De Angelis Anna Latiano Orazio Palmieri Tiziana Pia Latiano Marco Donatello Delcuratolo Matteo Tardio Francesca Bazzocchi Marco Gentile Fulvia Terracciano Grazia Anna Niro Francesca Tavano

Pancreatic cancer (PC) remains one of the most aggressive and lethal malignancies worldwide, largely due to late diagnosis, aggressive biology, limited therapeutic options and responsiveness. Conventional diagnostic and monitoring strategies, including imaging and serum biomarkers such as CA 19-9, provide limited sensitivity for early detection and suboptimal accuracy for the dynamic assessment of treatment response and disease evolution. These limitations highlight the urgent need for innovative, minimally invasive approaches capable of improving patient stratification and guiding personalized management. In this context, liquid biopsy has emerged as a promising, minimally invasive approach able to capture tumor-derived molecular information through the analysis of circulating cell-free nucleic acids, including circulating cell-free DNA (cfDNA) and circulating cell-free RNA (cfRNA). Released into the bloodstream by tumor cells, these analytes offer a real-time and comprehensive snapshot of tumor biology, capturing genetic, epigenetic, and transcriptional alterations through a simple blood draw. Liquid biopsy-based analyses hold significant potential for early detection, prognostic assessment, therapeutic decision-making, monitoring of minimal residual disease, and identification of resistance mechanisms. This review discusses the current state of research on circulating cell-free nucleic acids in PC, highlighting their biological basis, methodological approaches, clinical potential, and the challenges limiting their widespread implementation. By underscoring their translational relevance, we aim to outline how integrated liquid biopsy strategies, alongside the need for standardization and cross-study harmonization, may contribute to a more precise and dynamic approach to PC management.

Cells doi: 10.3390/cells15100903

Authors: Hua-Yun Huang Yi Kong Chun-Miao Li Yu-Le Sui Qian-Bao Wang Zhen-Hua Zhao Ling-Lin Kong Zhao-Lin Wu Wei Han

The fat mass and obesity-associated gene (FTO) has been shown to play a critical role in fat deposition in both humans and livestock. However, its involvement in subcutaneous and intramuscular fat deposition in chickens remains underexplored. In this study, we investigated the regulatory effects and pathways of FTO on subcutaneous and intramuscular fat deposition in chickens through functional gene verification and bioinformatics analysis. Our results demonstrated that, compared to the control group, exogenous transfection of an FTO lentiviral overexpression vector significantly inhibited cell proliferation and increased lipid accumulation in both subcutaneous and intramuscular adipocytes (p < 0.05). Furthermore, transfection of FTO siRNA markedly increased cell proliferation and reduced lipid accumulation in both subcutaneous and intramuscular adipocytes. A total of 413 and 164 differentially expressed genes were regulated by FTO in subcutaneous and intramuscular adipocytes, respectively. Pathway analysis revealed that the regulation of the actin cytoskeleton was a key process involved in FTO-mediated fat deposition in both subcutaneous and intramuscular adipocytes. Additionally, NRAS and ITGAV (subcutaneous fat), as well as FGF9, PIK3R2, FGF16, and RHOA (intramuscular fat), were identified as key genes enriched in this pathway. In conclusion, FTO differentially regulates fat deposition in chicken subcutaneous and intramuscular adipocytes by targeting distinct functional genes within the actin cytoskeleton pathway.

Cells doi: 10.3390/cells15100902

Authors: Jiao Lu Christopher Ward Sichong Qian Lilei Zhang Jiang Chang Zheng Sun

Histone deacetylase (HDAC) inhibitors are approved for cancer treatment and are being investigated for a wide range of other diseases. Despite their therapeutic promise, clinical studies have reported cardiac side effects, particularly electrocardiogram (EKG) abnormalities, with QT interval prolongation being one of the most consistently reported findings. The mechanisms underlying these cardiac effects remain unclear. In this study, we investigated the role of HDAC3 in cardiac electrophysiology. We found that postnatal depletion of cardiac HDAC3 in mice caused QT interval prolongation, recapitulating the EKG abnormalities reported with HDAC inhibitor use. Adult-onset inducible depletion of cardiac HDAC3 induced additional EKG abnormalities, including T-wave flattening, inversion, and biphasic T waves, which are also observed clinically. Loss of HDAC3 deacetylase activity, without affecting HDAC3 protein levels, was sufficient to induce QT prolongation. Disruption of HDAC3 function altered the expression of ion channel genes, including the downregulation of potassium channel genes such as Kcnh2, Kcne1, and Kcnip2. Moreover, a single dose of HDAC inhibitors, romidepsin or mocetinostat, caused reversible QT prolongation in mice. Consistent with these findings, HDAC inhibitor treatment altered the expression of potassium channel genes, with a predominant downregulation of multiple Kcn family members, including Kcnq1, Kcnh2, and Kcnip2. These findings establish HDAC3 enzymatic activity as a key regulator of cardiac repolarization and provide mechanistic insight into HDAC inhibitor-associated cardiotoxicity.

Cells doi: 10.3390/cells15100899

Authors: Maciej Pudełek Maksym Pudełek Julia Przeniosło Sylwia Kędracka-Krok Zbigniew Madeja Jarosław Czyż

Chemotherapy-induced metabolic reprogramming of glioblastoma multiforme (GBM) cells increases intracellular levels of reductive and energetic carriers, thereby fueling drug-relocation and retention systems and enhancing GBM drug-resistance. We have previously shown the role of this process in the adaptation of poly(morpho)nuclear “giant” cells (PGCs) in T98G populations to doxorubicin (DOX)-induced stress. Here, we addressed the role of a “resistance triad”, which coordinates metabolic T98G reprogramming with the activation of the drug-relocation and drug-retention axis, in the recovery of GBM populations from chemotherapeutic stress. A combination of proteomic analyses with metabolic and phenotypic profiling of pulse DOX-treated T98G cells revealed the significance of mitochondrial dynamics for the efficiency of the T98G “resistance triad”. DOX-induced mobilization of ATP-generating systems and ATP-dependent anabolic pathways was accompanied by the formation of DOX-negative, “mosaic” mitochondrial networks and the upregulation of mitofusin-2 (MFN2) in T98G PGCs. Transient MFN2 down-regulation correlated with the respiratory capacity of T98G cells, while impairing cell welfare in the absence and presence of DOX. However, minute fractions of PGCs, which withstood combined MFN2 down-regulation and pulse DOX treatment, retained mitochondrial networks and displayed efficient ABC transporter-/V-type channel-dependent lysosomal DOX retention. Collectively, a “triad” of mitochondrial activation, ABC transporter-dependent perinuclear redistribution and V-type channel-mediated lysosomal DOX compartmentalization determines DOX resistance of T98G cells. Whereas MFN2-dependent mitochondrial rearrangements may contribute to these processes, complementary adaptative mechanisms can compensate MFN2 dysfunction, limiting its potential as a therapeutic target.

Cells doi: 10.3390/cells15100901

Authors: Huma Shahzad Mohammed S. Razzaque

Phosphate (Pi) and calcium (Ca2+) are essential mineral ions that play coordinated roles in maintaining normal cellular functions. While various steps of calcium signaling are well characterized, emerging evidence suggests the critical role of both intracellular and extra cellular phosphate in regulating intracellular Ca2+. In the cytoplasm, phosphate influences ATP production and organelle calcium buffering and influences the activity of calcium pumps, such as sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA) and the plasma membrane Ca2+-ATPase (PMCA). Extracellular phosphate, taken up via sodium-dependent phosphate transporters, triggers signaling cascades that affect the processes of calcium influx, storage, and release. Additionally, high extracellular phosphate levels can disrupt calcium homeostasis through the systemic interactions of hormones such as fibroblast growth factor 23 (FGF23), vitamin D and parathyroid hormone (PTH), especially under pathological conditions such as chronic kidney disease (CKD). This article briefly summarizes the current understanding of the bidirectional influence of intra- and extracellular phosphate on calcium dynamics at the cellular level, with a focus on the underlying mechanisms.

Cells doi: 10.3390/cells15100900

Authors: Simona Di Stefani Maura Cimino Rosaria Tinnirello Martina Maria Cocco Cinzia Maria Chinnici Giandomenico Amico Valentina Di Felice Filippo Macaluso Bruno Douradinha Paolo Di Nardo Gioacchin Iannolo

Cardiovascular diseases remain the leading cause of mortality in developed countries. Among these conditions, acute myocardial infarction (AMI) is associated with particularly high rates of cardiac morbidity and mortality. Cardiac development in mammals is primarily dependent on cardiomyocyte (CM) proliferation during embryonic and early postnatal stages. However, following birth, the proliferative capacity of CMs declines markedly, with only limited cellular renewal occurring during adult life in response to pathological injury. Consequently, the irreversible loss of functional cardiomyocytes and the subsequent formation of fibrotic scar tissue frequently lead to persistent cardiac dysfunction and progressive impairment of cardiac physiology. Cardiomyocyte self-renewal is a tightly regulated process involving multiple molecular pathways. Among factors implicated in this regulation, microRNAs (miRNAs) have emerged as key modulators coordinating both cardiac development and tissue repair mechanisms. In this context, extracellular vesicles (EVs) have attracted considerable interest as potential modulators of these regenerative processes. In particular, mesenchymal stromal cells (MSCs) represent a promising therapeutic platform due to their immunomodulatory and anti-fibrotic properties demonstrated across multiple in vitro and in vivo models. Furthermore, the therapeutic potential of MSC-derived EVs can be enhanced through bioengineering approaches aimed at improving targeted molecular delivery. In this review, we summarize recent advances in the development and application of EV-based therapeutic strategies, with particular emphasis on their potential use as advanced therapy medicinal products (ATMPs) for cardiovascular regeneration and repair.

Cells doi: 10.3390/cells15100896

Authors: Benedict C. Albensi Aida Adlimoghaddam

Alzheimer’s disease (AD) is characterized by the accumulation of amyloid beta (Aβ) and neurofibrillary tangles in brain tissue; however, AD is multifactorial, and different etiopathogenic mechanisms involve factors that can affect mitochondrial function, which are associated with AD. While high-dose lithium is a well-established mood stabilizer, accumulating evidence suggests that low-dose lithium provides significant neuroprotection by reversing AD pathology, cognitive impairment, and inflammation. Despite these findings, there is limited information on how lithium affects brain energy metabolism. In the current study, we investigated the effect of lithium (0, 0.1, 1, and 10 mM) on mitochondrial function in AD neurons. Neuronal cells were isolated from the hippocampi of embryonic day 14–17 (E15–E17) control (C57BL/6) mice and 3xTg-AD mice. Mitochondrial oxygen consumption rate (OCR), mitochondrial Cytochrome C Oxidase (COX) activity, total ATP activity, and the expression of mitochondrial complex protein involved in oxidative phosphorylation (OXPHOS) were measured in control vs. 3xTg-AD in the presence and absence of lithium treatment. In the present study, lithium treatment significantly increased (p < 0.05) mitochondrial OCR, COX, total ATP, and levels of mitochondrial complex protein subunits (Complex I–V) in 3xTg-AD neurons. However, lithium had no effect on energy metabolism in control neurons. Together, these data indicate that lithium improves mitochondrial function under pathological states. Overall, these results have important implications for the treatment of disorders in which brain energy regulation is compromised, including AD. Particularly, our results highlight a role for lithium in regulating bioenergetics in early-stage AD and suggest that neuronal cells may be a crucial therapeutic target for preventing AD.

Cells doi: 10.3390/cells15100897

Authors: Jinmeng Chu Qingzhi Zhao Hui Ye Meixu Li Yizhen Wang Tiantian Xu Yong Cai Jingji Jin

Cyclin-dependent kinase 1 (CDK1) is frequently upregulated in multiple cancers and plays a central role in cell cycle progression and tumorigenesis. However, whether CDK1 directly regulates the histone acetyltransferase KAT8 (also known as MOF) in non-small cell lung cancer (NSCLC) remains unclear. Here, we identify CDK1 as a kinase that directly interacts with and phosphorylates KAT8 at serine 348 (S348) and threonine 418 (T418). Mechanistically, CDK1-mediated phosphorylation, particularly at S348, enhances the interaction between KAT8 and MSL1, thereby stabilizing the MSL complex and promoting KAT8-dependent acetylation of histone H4 at lysine 16 (H4K16). Functionally, the phosphorylation-deficient mutant KAT8-S348A exhibits impaired MSL complex assembly, reduced H4K16 acetylation, and decreased NSCLC cell proliferation both in vitro and in vivo. Pharmacological inhibition of CDK1 using RO-3306 suppresses KAT8 phosphorylation and H4K16 acetylation, leading to significant tumor growth inhibition. Notably, this effect is partially rescued by re-expression of wild-type KAT8 but not by the S348A mutant, supporting a phosphorylation-dependent mechanism. Collectively, these findings define a CDK1–KAT8 signaling axis that promotes NSCLC proliferation through epigenetic regulation and suggest that targeting CDK1-dependent KAT8 phosphorylation may represent a potential therapeutic strategy for lung cancer.

Cells doi: 10.3390/cells15100898

Authors: Luke Weaver Mikhail F. Alexeyev

Transmitochondrial cybrid technology is a key approach for elucidating the effects of mitochondrial DNA (mtDNA) mutations in defined nuclear genetic backgrounds and for studying nuclear–mitochondrial interactions. However, its application is limited by the availability of suitable recipient cell lines and by technically demanding enucleation procedures. We report three advances in cybrid technology: (1) enucleation using mitomycin C, a widely used agent for generating feeder layers in stem cell culture, which does not depend on cell attachment and provides a gentler alternative to actinomycin D; (2) selection of cybrids using mitochondrial uncouplers, which can reduce background survival of non-cybrid cells; and (3) cryopreservation of enucleated donor cells in liquid nitrogen, preserving fusion competence and increasing experimental flexibility. Additionally, we validate newly developed mtDNA-free (ρ0) derivatives of HCT116, HT1080, and U2OS cell lines as recipients for cybrid generation. These advances facilitated donor cell preparation, improved cybrid selection, and enhanced experimental flexibility, including the demonstration of preserved fusion competence of enucleated HeLa cells after 10 years of cryostorage. The ρ0 derivatives of HCT116, HT1080, and U2OS cells were confirmed as effective recipients. Together, these improvements enhance the efficiency and accessibility of transmitochondrial cybrid technology and are expected to facilitate its broader application.

Cells doi: 10.3390/cells15100895

Authors: Mingwei Wang Qiaohui Ying Qing Li Xia Lou Shuchang Dai Zhong Liu

Background: Macrophages were long regarded as passive executors of erythrophagocytosis responsible for systemic iron recycling. However, increasing evidence has reframed them as immunometabolic hubs that sense diverse environmental cues to modulate systemic iron homeostasis. Main body: This review examines the molecular architecture underlying macrophage iron metabolism and outlines how iron metabolic processes are dynamically regulated across spatial and temporal scales through the integration of mechanotransductive, mitochondrial, and epigenetic signaling pathways. Across disease contexts, macrophage iron handling displays marked heterogeneity, exemplified by contact-dependent iron transfer in tumors and ferroptosis-driven instability in cardiovascular disease. In cardiovascular pathologies, iron overload is associated with enhanced ferroptosis-related cascades that contribute to atherosclerotic plaque instability. Furthermore, at mucosal interfaces, host–pathogen competition over nutritional immunity highlights epigenetic strategies by which pathogens perturb host iron machinery. Conclusions: Linking these mechanistic insights to clinical translation, emerging therapeutic strategies are discussed that move beyond non-specific systemic iron chelation toward more targeted interventions. These include engineering macrophages for targeted drug delivery, exploiting nanomedicine-based redox modulation to influence macrophage phenotypes, and non-invasive regulation via the gut microbiota–epigenetic axis. Collectively, elucidating macrophage iron metabolic networks provides a conceptual framework for the development of precision approaches to inflammatory, metabolic, and malignant diseases.

Cells doi: 10.3390/cells15100894

Authors: Zhaoyuan Xu Yinzhi Xu Lidan You Hiroki Yokota

In the original publication [...]

Cells doi: 10.3390/cells15100893

Authors: Jianchun Xiao

Toxoplasma gondii is an obligate intracellular parasite that infects virtually all warm-blooded animals, progressing through acute and chronic stages. The Akt/mTOR signaling axis plays critical roles in cell survival, proliferation, and metabolism, making it a key target for intracellular pathogens. This study investigated how T. gondii infection modulates this pathway during both infections. Outbred CD-1 mice were infected intraperitoneally with the virulent GT1 strain of T. gondii. Mice for acute studies were sacrificed five days post-infection, while those for chronic studies were treated with sulfadiazine and sacrificed five months post-infection. Phosphoprotein expression of eight Akt/mTOR pathway components was measured in liver tissues using a multiplexed bead-based immunoassay. Acute T. gondii infection caused broad suppression of Akt/mTOR signaling, with 6 of 8 markers significantly downregulated, including pS6RPSer235/236, pAKTS473, pBADSer136, pIRS1S636/639, pPTENSer380, and pGSK-3α/βSer21/9. In contrast, chronic infection related to cyst burden selectively activates specific nodes of the pathway, including pBADSer136, pmTORSer2448, and pGSK-3α/βSer21/9. Infection induced strong correlations between inter-components, which reflect coherent and coordinated pathway-level reprogramming rather than random perturbation. These findings show that acute and chronic T. gondii infections have opposing effects on host Akt/mTOR signaling for their own benefit, which may present new therapeutic targets.

Cells doi: 10.3390/cells15100892

Authors: Evalds Viguls Anita Ilze Gulbe Simons Svirskis Valerija Groma Sandra Skuja

Astrocytes are key regulators of neuronal, metabolic, and vascular homeostasis, yet their morphological diversity and involvement in alcohol-related brain pathology remain incompletely characterized. In this study, we investigated astrocytic morphology in the human striatum of control individuals and subjects with short- and long-term alcohol exposure using immunohistochemistry combined with Sholl-based morphometric analysis, and ultrastructural assessment. GFAP immunohistochemistry was used to identify astrocytes, assess their morphology, and manually quantify GFAP+ cells in gray and white matter, followed by Sholl-based morphometric analysis to characterize astrocytic branching architecture and spatial organization. The number of GFAP+ astrocytes differed between tissue compartments, with a significant increase in white matter in alcohol-exposed individuals and no detectable change in gray matter. Morphometric analysis revealed pronounced astrocytic heterogeneity across all study groups. Sholl-derived metrics supported the distinction of six recurrent astrocytic morphometric profiles in the human striatum, distinguished by soma size, branching complexity, process length, and cell territory size. These profiles were present across gray and white matter, indicating intrinsic astrocytic structural diversity. Ultrastructural analysis further revealed alcohol-associated alterations at the astrocyte–vascular interface, including swelling of perivascular astrocytic endfeet, accumulation of intermediate filaments, and focal reductions in vascular wall coverage. Together, these findings demonstrate substantial astrocytic structural diversity in the human striatum accompanied by alcohol-related gliovascular remodeling.

Cells doi: 10.3390/cells15100891

Authors: Robert Lee Alexus Acton Madeline Holliday Nicholas J. Lennemann William J. Placzek

Hexokinase 2 (HK2) catalyzes the first committed step of glucose metabolism—the conversion of glucose to glucose-6-phosphate—directing carbon flux into an array of metabolic pathways such as glycolysis, pentose phosphate pathways, amino acid biosynthesis, and others. Given its prominent role in glucose metabolism, it is critical we understand the regulation of HK2 to appreciate its role in normal physiological function as well as in disease states like cancers. In this study we sought to establish the ability of myeloid cell leukemia 1 (MCL1) to bind and regulate HK2 via its reverse Bcl-2 homology (rBH3) motifs. We employed a combination of biochemical and metabolic analysis in non-small cell lung cancer (NSCLC) cell models (H1299, A549, and NCI-H23) to establish a fundamental link between apoptosis and metabolic regulation. This demonstrates that MCL1 directly binds and enhances HK2 enzymatic activity through interactions with rBH3 on HK2. Consequently, we observe significant reductions in glucose-derived metabolites and impaired cellular metabolic plasticity with the disruption of the HK2-MCL1 interaction. These findings establish a novel mechanism by which anti-apoptotic proteins can directly regulate glucose metabolism.

Cells doi: 10.3390/cells15100890

Authors: Ayhan Bilir Berna Yıldırım Mete Hakan Karalök

Mitochondrial stress has emerged as a key regulator of tumor–immune interactions, extending beyond its classical bioenergetic role to coordinate metabolic adaptation and immune regulation. Rather than merely accompanying tumor progression, mitochondrial dysfunction contributes to immune evasion and resistance to immunotherapy. Here, we propose that mitochondrial stress functions as a unifying axis governing three key determinants of anti-tumor immunity: immune visibility, immune cell fitness, and the metabolic architecture of the tumor microenvironment. Mechanistically, mitochondrial reactive oxygen species, mitochondrial DNA release, and mitophagy modulate antigen presentation and T cell function. We further highlight emerging experimental platforms, including 3D spheroid and organoid systems, that enable physiologically relevant investigation of mitochondria-driven tumor–immune interactions. Together, this perspective provides a mechanistic framework for understanding and targeting resistance to immune checkpoint blockade.

Cells doi: 10.3390/cells15100887

Authors: Dohyun Lee Jongsu Jeon Gyuwon Huh Seoyeong Baek Daehun Kim Hyeon-Ji Kang Hoyul Lee In-Kyu Lee Jae-Han Jeon Hoe-Yune Jung

Amino acids are key regulators of metabolism, their coordinated effects on skeletal muscle signaling and metabolic remodeling under physiological conditions remain incompletely understood. Here, we investigated whether 14 weeks of combined histidine and proline supplementation (HISPRO; 700 mg/kg) enhances skeletal muscle function through metabolic reprogramming in normal ICR mice. HISPRO significantly improved muscle performance compared with the control group, including grip strength, rota-rod, and treadmill. Histological and biochemical analyses revealed a shift toward oxidative muscle phenotype compared with the control group, with larger muscle fibers and succinate dehydrogenase-positive fibers. Consistently, HISPRO promoted mitochondrial biogenesis and oxidative metabolism compared with the control group, as evidenced by upregulation of mitochondrial regulatory genes, mitochondrial DNA copy number, citrate synthase activity, and oxidative phosphorylation (OXPHOS) complex levels in skeletal muscles. Mechanistically, HISPRO was associated with activation of the SIRT1-PGC1α-AMPK signaling axis compared with the control group, as evidenced by increased Nampt and Nmnat1 expression, an elevated NAD+/NADH ratio, and enhanced AMPK phosphorylation. SIRT1 inhibition markedly attenuated HISPRO-induced increases in mitochondrial biogenesis markers but did not fully suppress OXPHOS protein expression, suggesting the involvement of both SIRT1-dependent and -independent mechanisms. Notably, HISPRO also improved muscle function in dexamethasone-induced muscle atrophy model. It restored mitochondrial biogenesis and function, and suppressed atrophy-related markers compared with the dexamethasone-treated group. HISPRO may contribute to improving muscle quality through coordinated metabolic regulation and could represent a complementary nutritional for supporting muscle metabolic health.

Cells doi: 10.3390/cells15100889

Authors: Michael Joseph Martina Gabrielli Elisa Tonoli Gareth W. V. Cave Elisabetta A. M. Verderio

Mesenchymal stromal cells (MSCs), also known as multipotent stromal cells or mesenchymal stromal cells, support cell growth and viability through the secretion of trophic factors and immunomodulatory molecules. Their secretome exerts cytoprotective effects in the brain, although the mechanisms underlying MSC-mediated neurological recovery remain poorly understood. A substantial portion of the MSC secretome is delivered via extracellular vesicles (EVs), membrane-bound particles that facilitate intercellular communication. EVs derived from MSCs of various origins exhibit therapeutic potential, and numerous studies are examining the miRNA and protein cargo contained within MSC-EVs. Despite these efforts, methodological differences across the literature and the inherent variability associated with MSC sources have limited data interpretation and identification of EV-factors which may be responsible for neuroprotection. In this study, we have reviewed proteomic, transcriptomic and lipidomic datasets from a selection of recent MSC-EV studies, to identify shared cargo components that may contribute to promoting cell repair and plasticity in brain, counteracting neurodegeneration.

Cells doi: 10.3390/cells15100888

Authors: Srdjan Aleksandric Barry Uretsky Ana Djordjevic-Dikic Dejan Orlic Nebojsa Antonijevic Milorad Tesic Stefan Juricic Marko Banovic Vojislav Giga Nikola Boskovic Zlatko Mehmedbegovic Ivana Jovanovic Dejan Simeunovic Sinisa Stojkovic Vladan Vukcevic Goran Stankovic Branko Beleslin

Myocardial bridging (MB) is a congenital coronary anomaly characterized by systolic compression of the intramyocardial arterial segment and delayed early diastolic artery relaxation, resulting in reduced vessel luminal diameter in diastole. Current evidence suggests that MB, particularly in the left anterior descending artery, may cause anginal symptoms and/or myocardial ischemia through several different pathophysiological and cellular mechanisms acting independently or synergistically: (1) delayed early diastolic relaxation of intramyocardial arterial segment; (2) impaired endothelial-dependent vasodilation with vessel smooth muscle cell hyperactivity in the coronary artery with MB, especially within the bridged segment; (3) focal (septal) ischemia due to “septal steal” phenomenon; and (4) development and progression of an atherosclerotic lesion in the coronary artery segment proximal to MB. Patients with isolated-MB may also experience anginal pain and/or myocardial ischemia due to concomitant structural and/or functional abnormalities of the coronary microcirculation. Both MB and coronary microvascular dysfunction refer to a subgroup of patients with angina and/or ischemia with non-obstructive coronary arteries (ANOCA/INOCA). Therefore, it may be challenging to determine whether MB is causing anginal pain and/or ischemia, particularly since both phenomena have also been reported without MB’s existence. Therefore, comprehensive coronary physiology testing should be encouraged in patients with this coronary anomaly to identify the underlying cause of anginal pain and/or myocardial ischemia, enabling optimal therapeutic strategies in these patients. This review is focused on different pathophysiological and cellular mechanisms of MB-related angina and/or ischemia and future perspectives in the functional assessment of MB severity, bearing in mind the complexity of coronary physiology in the presence of this anomaly.

Cells doi: 10.3390/cells15100886

Authors: André Vinicius Saueressig Kruel Mariângela Ferreira Daiane Agostini Cristiano Valter Diesel Marcelo Queiroz Carlos Roberto Galia Guilherme Liberato da Silva Stephany Huber Fernanda Majolo

Introduction: Orthobiologics such as Platelet-Rich Plasma (PRP) and Injectable Platelet-Rich Fibrin (i-PRF) have emerged as promising tools in regenerative medicine. However, the lack of methodological standardization and the still limited comparative characterization between these products represent significant barriers to their optimized clinical application. This comparative laboratory study aimed to characterize and differentiate PRP and i-PRF, focusing on their cellular composition, obtained volume, and total Platelet-Derived Growth Factor (PDGF-BB) content. Materials and Methods: This study was conducted with 34 individuals meeting standard blood donation criteria. Peripheral blood samples were collected from all participants. PRP was obtained using a modified double-spin centrifugation protocol, whereas i-PRF was prepared using a modified low-speed centrifugation technique. Cellularity (platelet and leukocyte counts), final produced volume, and total PDGF-BB content were assessed using complete blood count analysis and an enzyme-linked immunosorbent assay (ELISA), respectively. Statistical analysis was performed using Linear Mixed Models (LMMs). Results: Both protocols resulted in significant increases in platelet and leukocyte concentrations compared to baseline values. PRP showed significantly higher platelet and leukocyte concentrations compared with i-PRF, as well as markedly higher PDGF-BB levels. In contrast, i-PRF yielded a substantially greater final volume and enabled a higher absolute delivery of total leukocytes, whereas PRP delivered a greater absolute number of platelets. In exploratory analyses, female sex, the presence of comorbidities, and increased abdominal circumference were associated with variations in product volume and cellular composition. Discussion: These findings indicate that PRP and i-PRF exhibit distinct biological profiles in terms of cellularity, volume, and total PDGF-BB content. Whether these laboratory differences translate into distinct clinical outcomes remains unknown. The results should therefore be viewed as hypothesis-generating: they suggest that PRP and i-PRF may not be interchangeable, and that future randomized clinical trials are needed to define product-specific indications based on the target tissue and desired biological mechanism.

Cells doi: 10.3390/cells15100885

Authors: Hsin-Ju Chiang Chang-Chun Hsiao Steve Leu

Background: Aging, especially after menopause, reduces the quantity and function of adult stem cells. Estrogen deficiency impairs proliferation, differentiation, and regenerative capacity. This study evaluated whether estrogen enhances endothelial differentiation of adipose-derived stromal cells (ADSCs) and improves therapeutic efficacy in critical limb ischemia (CLI). Methods: Male-derived ADSCs were assessed in vitro for endothelial differentiation using flow cytometry, biochemical assays, and angiogenesis analyses. Therapeutic effects were tested in a rat CLI model using endothelial-differentiated ADSCs (ED-ADSCs) with or without 17β-estradiol (E2). An ovariectomized (OVX) model examined estrogen deficiency and supplementation in vivo. Results: E2 promoted endothelial differentiation, increasing ERα/ERβ expression and activating PI3K/Akt/eNOS and MAPK signaling. This led to elevated VEGF expression, enhanced tube formation, and increased CD34+, KDR+, and CD31+ cell populations. In vivo, E2-pretreated ED-ADSCs significantly improved blood flow recovery. Estrogen deficiency reduced endothelial progenitor populations, which were restored by E2 supplementation. Conclusions: Estrogen modulates endothelial-associated characteristics and angiogenesis-related responses of ADSCs via ER-associated signaling, and may contribute to improved functional outcomes in ischemic conditions. E2 preconditioning may represent a potential strategy for stem cell-based therapy in estrogen-deficient settings.

Cells doi: 10.3390/cells15100884

Authors: Chie Masaki Norihito Inoue Tomohiro Chiba

Signal transducers and activators of transcription (STAT) proteins, which operate via canonical and non-canonical mechanisms, are critically implicated in thyroid tumorigenesis. This review integrates their multifaceted roles in thyroid cancer. STAT3 acts as a “double-edged sword”: hyperactive STAT3 drives metastasis and BRAF inhibitor resistance in advanced carcinomas, yet paradoxically acts as a tumor suppressor by restraining the Warburg effect via non-canonical mitochondrial localization. Clinically, preserved nuclear STAT3 independently predicts a favorable prognosis and is inversely correlated with TERT promoter mutations, offering a biological modifier for clinical risk stratification. Furthermore, STAT1 regulates differentiation via the IGF2BP2-m6A axis, STAT5 drives proliferation upon release from TRβ suppression, and STAT6 confers chemoresistance. While novel direct STAT3 inhibitors (e.g., TTI-101) and rational combinations with immune checkpoint inhibitors or STING agonists show promise in overcoming refractory disease, the intricate dual functionality of STAT family proteins demands rigorous biomarker-guided precision medicine approaches.

Cells doi: 10.3390/cells15100883

Authors: Shuangrui Chen Jin Yan Xiaochun Peng

Immunotherapy has emerged as an established clinical approach for lung cancer; however, both intrinsic and adaptive resistance mechanisms substantially constrain its therapeutic efficacy. Within the tumor microenvironment (TME), cancer-associated fibroblasts (CAFs) serve as pivotal stromal mediators of this resistance. These fibroblasts manifest their immunomodulatory effects, in part, through the secretion of small extracellular vesicles (sEVs). While multiple lines of evidence strongly implicate CAF-sEVs in immunotherapy resistance, establishing a direct causal link remains an active area of investigation. Clinically, elevated levels of specific CAF-sEV cargoes correlate with poor response in lung cancer patients. Functionally, CAF-sEVs can directly suppress T-cell activity and drive pro-resistance phenotypes via defined molecular pathways, and pharmacological inhibition of sEV secretion has been shown to attenuate resistance in preclinical models. However, the extent to which these effects are independent of other CAF-derived factors remains to be fully elucidated in vivo. This review comprehensively synthesizes the biophysical properties of CAF-derived sEVs, delineates the molecular mechanisms underpinning their role in immunotherapy resistance, critically evaluates the existing causal evidence and its limitations, and assesses their translational potential as diagnostic biomarkers and therapeutic targets—ultimately providing a conceptual framework to overcome resistance barriers in lung cancer immunotherapy.

Cells doi: 10.3390/cells15100882

Authors: Sonali Singh Sabine Ring Xuan Lei Mohamad Alabdullah Yutaka Inaba Alexander Enk Karsten Mahnke

The surface receptor DEC205 (alias CD205, NLDC-145, Ly75) has long been a marker for murine dendritic cells and Langerhans cells and functions as receptor to facilitate antigen uptake. We revisited the tissue expression pattern of DEC205 in mice and revealed expression by endothelial cells (ECs) in the central nervous system. Using the murine brain-derived EC line bEnd.3, we could show that DEC205 serves as an endocytic receptor that directs potential ligands to lysosomal compartments. As for its function in ECs, upon engagement by anti-DEC205 antibodies, DEC205 seems to guide Claudin 5, a component of endothelial junctions, away from the surface of ECs to a LAMP-1+, degradative compartment. This opens endothelial junctions, eventually allowing leukocytes to migrate from blood to tissue sites. In aggregate, by regulating the blood–brain barrier, DEC205 in ECs may contribute to the regulation of inflammatory processes in the central nervous system.

Cells doi: 10.3390/cells15100881

Authors: Gerhild Euler Hannah Disch Maximilian Trautmann Anne Bernhardt Jennifer Krechmeier Rainer Schulz Jacqueline Heger

MAO-B-specific inhibition, either in knockout (KO) mice or pharmacologically, preserves left ventricular function and reduces cardiac fibrosis after myocardial infarction or pressure overload. We investigated whether stimulation of MAO-B in cardiac fibroblasts provokes ROS production and myofibroblast development. Fibroblast-specific MAO-B knockdown (KD) mice were created by crossing Col1a2CreERT mice with MAO-Bfl/fl mice. The KD was induced by tamoxifen injection. Fibroblasts of KD mice and wild types (WTs) were isolated and reduced MAO-B expression in KD fibroblasts was confirmed. In isolated mitochondria from the left ventricle of these mice, ROS production was reduced under stimulation with the specific MAO-B substrate β-phenylethylamine (PEA). Mitochondrial ROS production in fibroblasts, detected by MitoSox Red staining, increased under PEA (1000 µM) stimulation only in WT fibroblasts. mRNA of the marker genes for myofibroblast differentiation, Col1a1 and periostin, increased 2- or 3-fold, respectively, in WT but not in MAO-B KD fibroblasts. The enhanced migration potential under PEA was reduced in MAO-B KD fibroblasts. In conclusion, stimulation of MAO-B in cardiac fibroblasts leads to the formation of mitochondrial ROS, enhancement of myofibroblast marker gene expression and migration of the cells. Excessive fibrosis caused by elevated MAO-B activity in myocardial infarction can therefore contribute to cardiac dysfunction.

Cells doi: 10.3390/cells15100880

Authors: Yinglin Liu Liling Xu Yanmei Li Cheng Lu Zepei Fan Jun Ling Yingwei Wang Zheng Wu

Limbal stem cell (LSC) transplantation is an important treatment for limbal stem cell deficiency (LSCD), but low efficacy in maintaining LSC stemness during in vitro expansion greatly affects its wider application. The primary contributing factors include a low proportion of stem cells and the lack of a stable, supportive microenvironment over prolonged culture. Rabbit corneal tissues preserved under deep cryogenic conditions for more than six months retain viable limbal stem cells (LSCs), and primary LSCs isolated from these tissues exhibit robust stem cell characteristics. It is noteworthy that the NLRP3/Caspase-1/IL-1β signaling axis was activated in corneal epithelial cells, and outer limbal layers preserved for one or two years. Based on these findings, a combined strategy integrating deep cryopreservation with IL-1β induction was established for the processing of limbal tissues. The combined cryogenic and IL-1β preconditioning yielded primary LSCs with maintained p63+ cell proportions, a reduction in K3+ differentiated cells from approximately 80% to 60%, and a 6.25-fold increase in colony-forming efficiency. In addition, an increased proportion of cells in the G2/M phase and enhanced proliferative capacity were observed. The enriched LSC population also exhibited improved stratified epithelial reconstruction potential. These findings identify an effective strategy for preserving and enriching LSCs from limbal tissue, providing a practical and efficient approach for LSC preparation prior to transplantation. Further in vivo studies will be important to validate the functional performance of these cells in ocular surface reconstruction.

Cells doi: 10.3390/cells15100878

Authors: Emilia Magdalena Łukasik Klaudia Aneta Marcinkowska Agnieszka Śmieszek

Canine osteosarcoma (OSA) is a highly aggressive primary bone tumor and a valuable model in comparative oncology. Nevertheless, commonly used canine in vitro models remain incompletely and inconsistently characterized, while exhibiting substantial biological heterogeneity affecting experimental outcomes. This study aimed to comparatively characterize three canine osteosarcoma cell lines (OSCA8, OSCA29, and D17) in reference to canine hTERT fibroblasts, and with a focus on functional properties and selected molecular features, namely including miR-27b-3p and IGF2BP3 expression. The cytophysiological profile of the cells was evaluated in relation proliferation and migratory capacity. In turn, gene expression was determined with RT-qPCR, and proteins detected with Western blotting. The D17 cell line showed the highest metabolic activity and the largest fraction of S-phase cells, whereas OSCA8 cells demonstrated the greatest clonogenic potential and the highest migratory activity in the wound healing assay. OSCA29 cells displayed an intermediate functional profile, while all OSA cell lines exhibited comparable migratory capacity in transwell assay. At the molecular level, miR-27b-3p expression was significantly higher in OSCA8 and D17 cells than in OSCA29 cells. In turn, IGF2BP3 transcript levels were lower in OSCA29 cells, whereas protein analysis revealed distinct immunoreactive forms. Together, these findings highlight the functional heterogeneity of commonly used canine osteosarcoma cell lines and broaden their current characterization.

Cells doi: 10.3390/cells15100879

Authors: Elizabeth R. Dufficy Amalia Goula Emma Melia Abigail Bellamy Jason L. Parsons

The use of proton beam therapy (PBT), as a more precision-targeted radiotherapy technique, is increasing in the treatment of head and neck squamous cell carcinoma (HNSCC). PBT benefits from the precise delivery of the radiation dose to the tumour via the Bragg peak. However, challenges still remain in the treatment of HNSCC with radiotherapy, particularly with tumour radioresistance and recurrence, requiring strategies leading to radiosensitisation. There are added complexities with the use of PBT given the increase in linear energy transfer (LET) at and around the Bragg peak, which can cause an altered cellular response compared to low-LET radiation. Nevertheless, targeting the cellular DNA damage response is considered an important strategy to enhance tumour cell killing caused by radiotherapy. Therefore, using specific inhibitors against the protein kinases ataxia telangiectasia mutated (ATM), ataxia telangiectasia and Rad3-related (ATR) and the DNA-dependent protein kinase catalytic subunit (DNA-Pkcs), we investigated their impact in radiosensitising HPV-negative HNSCC cells to PBT of increasing LET. We demonstrate that inhibitors against ATR (AZD6738), and particularly ATM (AZD1390) and DNA-Pkcs (AZD7648), could significantly decrease clonogenic survival of HNSCC cell lines following PBT at both low and relatively high LET (~2 keV/µm and ~8 keV/µm, respectively). We confirmed that the inhibitors in combination with PBT led to DSB persistence through neutral comet assays and monitoring γH2AX/53BP1 foci. We also show that this strategy can enhance the sensitivity of patient-derived organoids of HNSCC to PBT of both low and high LET, highlighting this as a strategy which should be exploited further.

Cells doi: 10.3390/cells15100877

Authors: Kyungmin Ji Chenjun Shi Jitao Zhang Raymond R. Mattingly

Plexiform neurofibromas (pNF1s) are benign peripheral nerve sheath tumors caused by NF1 loss, leading to dysregulated RAS/mitogen-activated protein kinase (MAPK) signaling. While the mitogen-activated protein kinase kinase (MEK) inhibitors, selumetinib and mirdametinib, can reduce tumor volume, surgical resection remains the primary treatment for immediate debulking and symptom relief. Complete removal is often limited by tumor infiltration along nerve plexuses, and residual tumors may undergo postsurgical tissue remodeling, producing localized regions of stiffened extracellular matrix (ECM). The impact of ECM stiffness on pNF1 growth and drug responses remains unclear. Using immortalized patient-derived pNF1 tumor cell lines cultured in 3D hydrogels with defined stiffness (1.5 kPa, soft; 7 kPa, stiff), we found that stiff ECM promoted spread morphology, increased growth, and progressive intracellular softening. Stiff ECM also reduced lysyl oxidase (LOX) expression, suggesting mechanoadaptive ECM remodeling, and increased P-glycoprotein expression. Under the same conditions, stiff ECM was associated with reduced sensitivity to selumetinib. These results provide the first evidence that ECM stiffening, including that plausibly associated with postsurgical remodeling, may contribute to pNF1 growth and reduced sensitivity to selumetinib in this 3D pNF1 culture model. Our findings highlight mechanobiology as a key regulator of tumor behavior and support further investigation of ECM-targeted strategies to improve outcomes in neurofibromatosis type 1 (NF1).

Cells doi: 10.3390/cells15100875

Authors: Adriana Ancer-Arellano Yareth Gopar-Cuevas María-de-Lourdes Chávez-Briones Ivett Miranda-Maldonado Sofia A. Córdova-Zúñiga Jesús Ancer-Rodríguez Marta Ortega-Martínez Gilberto Jaramillo-Rangel

Aging is the primary biological driver of progressive cellular dysfunction and a major risk factor for disease development. The lungs and kidneys are highly vulnerable to cellular damage during aging due to their continuous exposure to environmental and metabolic stressors. Increasing evidence supports the existence of a bidirectional communication axis between the lungs and kidneys. In this review, we propose an integrative mechanistic framework that links alterations in cell turnover along this axis during aging. Based on the literature reviewed, we found that age-related cellular changes induce cellular senescence. Senescent cells undergo irreversible cell cycle arrest; furthermore, telomere shortening limits cell proliferation and promotes resistance to apoptosis. However, apoptosis can increase when a critical damage threshold is reached. In this context, senescent cells acquire a senescence-associated secretory phenotype (SASP) and release circulating mediators that can transmit damage signals between the lungs and kidneys. Taken together, these processes promote a pathological feedback loop in which age-related changes in one organ can exacerbate dysfunction in another, reinforcing a bidirectional axis of damage that increases susceptibility to developing lung and kidney diseases.

Cells doi: 10.3390/cells15100876

Authors: Karina Köpke Inge S. Zuhorn Frank A. E. Kruyt

Glioblastomas (GB) are highly aggressive brain tumors with poor patient prognosis and low survival rates. To identify novel therapeutic targets, the tumor microenvironment (TME) is increasingly examined, with a particular focus on biomechanical changes in the extracellular matrix (ECM) that contribute to GB aggressiveness. In GB, the ECM stiffens, regulating cell behavior through mechanotransduction. Preclinical in vitro and ex vivo studies generally report increased stiffness in GB relative to healthy brain tissue, whereas clinical in vivo measurements often report decreased stiffness. This review examines potential causes for this discrepancy, highlighting both biological and technical factors. Preclinical measurements are frequently performed using atomic force microscopy (AFM), while clinical stiffness is assessed via magnetic resonance elastography (MRE). Differences in methodology, including sample preparation, measurement modalities, and spatial scale, partly explain divergent stiffness values. Biological factors such as necrosis, edema, and physical confinement by the skull, which are preserved only in vivo, also contribute to these differences. To reconcile these findings, future research should employ physiologically relevant in vitro models that better replicate in vivo GB biomechanics, together with high-throughput and accurate animal models. Integrating these approaches may clarify the biomechanical landscape of GB and result in more effective therapeutic strategies.