Background/Objectives: Metabolic dysfunction-associated steatohepatitis (MASH), characterized by liver inflammation, fibrosis, and fat accumulation, can develop into cirrhosis and liver cancer. Despite its increasing prevalence worldwide, there are few established therapies for advanced MASH. We previously demonstrated that stem cells from human exfoliated deciduous teeth-conditioned media (SHED-CM) exerted therapeutic effects in a MASH mouse model. The gut-liver axis is thought to be associated with liver disease progression, and soluble Siglec-9 (sSiglec-9), an immunoinhibitory receptor, is a key protein in SHED-CM that induces anti-inflammatory macrophages and has intestinal epithelial protective effects. Therefore, we evaluated sSiglec-9's role in intestinal barrier protection in MASH mice. Methods: We evaluated sSiglec-9 effects on intestinal barrier function using in vitro Caco-2 cell monolayers injured by TNF-α and IFN-γ. For the MASH mouse model, male C57BL/6J mice were given a Western diet and high-sugar solution orally; to induce liver injury, CCl4 was intraperitoneally administered for 12 weeks. Mice were treated weekly with 10 ng/g sSiglec-9 or vehicle. Intestinal permeability was assessed by blood 4 kDa FITC-dextran concentration, and intestinal transcriptomes and liver histology were analyzed. Results: sSiglec-9 decreased intestinal permeability and liver inflammation in MASH mice. sSiglec-9 and SHED-CM reduced 4 kDa FITC-dextran permeability in injured Caco-2 cells, and sSiglec-9 significantly reduced intestinal permeability and modulated expression of 34 intestinal genes. The NAFLD Activity Score indicated significantly reduced inflammation following sSiglec-9 treatment. Conclusions: sSiglec-9 may protect intestinal barrier function by mitigating mucosal inflammation. sSiglec-9 treatment may represent a novel therapeutic approach for MASH via gut-liver axis modulation.

Nanotechnology is transforming contemporary medicine by providing cutting-edge tools for the treatment and diagnosis of complex disorders. Advanced techniques such as bioimaging and photodynamic therapy (PDT) combine early diagnosis and targeted therapy, offering a more precise approach than conventional treatments. However, a significant obstacle for PDT is the need to selectively deliver photosensitizers to disease sites while minimizing systemic side effects. In this context, mesenchymal stem cells have emerged as promising biological carriers due to their natural tropism towards tumors, low immunogenicity, and their ability to overcome biological barriers. In this study, two push-pull compounds, NPI-PTZ and BTZ-PTZ, phenothiazine derivatives featuring aggregation-induced emission (AIE) abilities, were analyzed. These molecules proved to be excellent fluorescent probes and photosensitizing agents. When administered to human bone marrow-derived multipotent stromal cells (hBM-MSCs) and human adipose multipotent stem cells (hASCs), the compounds were efficiently internalized, maintained a stable fluorescent emission for several days, and showed phototoxicity after irradiation, without inducing major cytotoxic effects under normal conditions. These results highlight the potential of NPI-PTZ and BTZ-PTZ combined with mesenchymal stem cells as theranostic tools, bridging bioimaging and PDT, and suggest new possibilities for advanced therapeutic approaches in clinical applications.

Gliomas are the most common central nervous system tumors and account for 30% of all primary brain tumors, 80% of all malignant ones, and the vast majority of deaths that are caused by brain tumors. Among them, glioblastoma multiforme has the most aggressive and invasive course. Due to its heterogeneity, it is difficult to treat, and one of the reasons for this are glioma stem cells (GSCs). Therapies such as radiotherapy and chemotherapy are used to treat gliomas but do not bring the expected results. Therefore, treatments targeting glioma stem cells are emerging. A promising strategy is to target GSCs with natural compounds. This review aims to describe the problem of glioma stem cells, the treatment of gliomas, and therapies based on natural compounds, which are promising for the future.

Breast cancer (BC) frequently metastasizes to bone, leading to poor patient prognosis. The infiltration of cancer cells in bone impairs its homeostasis, triggering a pathological interaction between tumors and resident cells. Preclinical models able to mimic the bone microenvironment are needed to advance translational findings on BC mechanisms and treatments. We designed strontium-doped calcium phosphate cement to be employed for culturing cancer and bone cells and developed an in vitro bone metastasis model. The platform was established step by step, starting with the monoculture of cancer cells, mature osteoblasts (OBs) differentiated from mesenchymal stem cells, and mature osteoclasts (OCs) differentiated from Peripheral Blood Mononuclear Cells. The model was implemented with the co-culture of cancer cells with OBs or OCs, or the co-culture of OBs and OCs, allowing us to discriminate the interaction between the actors of the bone metastatic niche. The biomimetic material was further challenged with bone metastasis patient-derived material, showing good versatility and biocompatibility, suggesting its potential use as bone substitute. Overall, we developed a bone-mimicking model able to reproduce reciprocal interactions between cancer and bone cells in a biomimetic environment suitable for studying the biomolecular determinants of bone metastasis and, in the future, as a drug efficacy platform.

Electrospun collagen-based fibrous mats are of increasing interest for cell culture, regenerative medicine, and tissue engineering. The focus of this investigation is on the assessment of the electrospinning ability of bovine split hide collagen (BSHC), the effect of glycosaminoglycan (GAG) incorporation on the mats' structural morphology, and the impact on the adhesion and proliferation of human adipose-derived mesenchymal stem cells (hAD-MSCs). Electrospun mats were prepared using benign and fluoroalcohol solutions of BSHC and BSHC/GAGs under varied operation conditions. SEM observations and analysis were employed to characterize the structural morphology of the mats. Several parameters were used to evaluate the hAD-MSC behavior: cytotoxicity, cell morphology, cell number and spreading area, cytoskeleton, focal adhesion contacts, and cell proliferation. Electrospinning using benign solvents was impossible. However, fiber mats were successfully prepared from hexafluoropropanol (HFP) solutions. Different structural morphologies and fiber diameters of the electrospun mats were observed depending on the composition and concentration of the electrospinning solutions. Both BSHC and BSHC/GAG mats supported the in vitro adhesion, growth, and differentiation of hAD-MSCs, with some variations based on their composition and structural morphology. The absence of cytotoxicity and the good hAD-MSC adhesiveness make them promising substrates for cell adhesion, proliferation, and further stem cell differentiation.

The epigenetic regulation of adipogenic differentiation in dental stem cells (DSCs) remains poorly understood, as research has prioritized osteogenic differentiation for dental applications. However, elucidating these mechanisms could enable novel regenerative strategies for soft tissue engineering. Periodontal ligament stem cells (PDLSCs) exhibit notable adipogenic potential, possibly linked to histone 3 acetylation at lysine 9 (H3K9ac); however, the mechanistic role of this modification remains unclear.

Protein-coated ultra-high viscosity (UHV)-alginate hydrogels are essential to mimic the physiological in vivo environment of humans in several in vitro applications. This work presents an optimized bioreactor-integrated freeze-drying process for MatrigelTM-coated UHV-alginate microcarriers in the context of human induced pluripotent stem cell (hiPSC) expansion. The impact of freeze-drying on the UHV-alginate microcarriers using trehalose 100 mg/mL in 0.9% NaCl as a lyoprotective agent, as well as the stem cell response using hiPSCs, was analyzed using microscopy-based screenings. First observations of the process showed that the integrity of the cake was preserved in the samples with a maximum vapor exchanging rate. Following rehydration, the UHV-alginate microcarriers retained their original morphology. Upon the addition of Poloxamer 188, stickiness and bubble formation were reduced. The expansion of hiPSCs in a suspension bioreactor resulted in a 5-7-fold increase in total cell count, yielding at least 1.3 × 107 cells with viability exceeding 80% after seven days of cultivation. In flow cytometry analysis, the pluripotency factors OCT3/4 and SSEA4 resulted in positive signals in over 98% of cells, while the differentiation factor SSEA1 was positive in fewer than 10% of cells. Supported by preceding in silico predictions of drying time, this study presents, for the first time, basic steps toward a "ready-to-use" bioreactor-integrated freeze-drying process for UHV-alginate microcarriers in the iPSC context.

Diabetic foot ulcers (DFU) represent a major complication of diabetes mellitus, affecting millions of patients worldwide and leading to high morbidity and amputation risks. The impaired healing process in DFU is driven by vascular insufficiency, neuropathy, chronic inflammation, and infections. Conventional treatments, including blood sugar control, wound debridement, and standard dressings, have shown limited efficacy in achieving complete healing. Recent advancements have introduced novel therapeutic approaches such as stem cell therapy, exosome-based treatments, and bioengineered scaffolds to accelerate wound healing and tissue regeneration. Mesenchymal stem cells (MSCs), particularly adipose-derived stem cells (ASCs), exhibit anti-inflammatory, pro-angiogenic, and immunomodulatory properties, enhancing wound repair. Additionally, exosomes derived from ASCs have demonstrated the ability to promote fibroblast proliferation, regulate inflammation, and stimulate angiogenesis. The integration of bioengineered scaffolds, including hydrogels, hyaluronic acid (HA), or micro-fragmented adipose tissue (MFAT), offers improved drug delivery mechanisms and a controlled healing environment. These scaffolds have been successfully utilized to deliver stem cells, growth factors, antioxidants, anti-glycation end products, anti-inflammatory and anti-diabetic drugs, or antimicrobial agents, further improving DFU outcomes. This review highlights the potential of combining novel 3D scaffolds with anti-diabetic drugs to enhance DFU treatment, reduce amputation rates, and improve patients' quality of life. While promising, further clinical research is required to validate these emerging therapies and optimize their clinical application.

Delayed or non-healing of bone defects in an aging, multi-morbid population is still a medical challenge. Current replacement materials, like autografts, are limited. Thus, artificial substitutes from biodegradable polymers and bioactive glasses (BGs) are promising alternatives. Here, novel cerium-doped mesoporous BG microparticles (Ce-MBGs) with different cerium content were included in photocrosslinkable, methacrylated gelatin (GelMA) for promoting cellular functions of human mesenchymal stem cells (hBMSCs). The composites were studied for intrinsic morphology and Ce-MBGs distribution by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDX). They were gravimetrically analyzed for swelling and stability, compressive modulus via Microsquisher® and bioactivity by Fluitest® calcium assay and inductively coupled plasma-optical emission spectrometry (ICP-OES), also determining silicon and cerium ion release. Finally, seeding, proliferation, and differentiation of hBMSCs was investigated. Ce-MBGs were evenly distributed within composites. The latter displayed a concentration-dependent but cerium-independent decrease in swelling, while mechanical properties were comparable. A MBG type-dependent bioactivity was shown, while an enhanced osteogenic differentiation of hBMSCs was achieved for Ce-MBG-composites and related to different ion release profiles. These findings show their strong potential in promoting bone regeneration. Still, future work is required, e.g., analyzing the expression of osteogenic genes, providing further evidence for the composites' osteogenic effect.

Endocannabinoid signaling plays a significant role in neurogenesis and nervous system physiology, but its roles in the development of other tissues are just beginning to be appreciated. Previous reports have shown the presence of the key endocannabinoid receptor Cannabinoid receptor 1 (CB1 or Cnr1) in multiciliated (MCC) tissues and its upregulation in kidney diseases, yet the relationship between Cnr1 and renal MCC development is unknown. Here, we report that Cnr1 is essential for cilia development across tissues and regulates renal MCCs via cyclic AMP (cAMP) signaling during zebrafish embryogenesis. Using a combination of genetic and pharmacological studies, we found that the loss of function, agonism and antagonism of cnr1 all lead to reduced mature renal MCC populations. cnr1 deficiency also led to reduced cilia development across tissues, including the pronephros, ear, Kupffer's vesicle (KV), and nasal placode. Interestingly, treatment with the cAMP activator Forskolin (FSK) restored renal MCC defects in agonist-treated embryos, suggesting that cnr1 mediates cAMP signaling in renal MCC development. Meanwhile, treatment with the cAMP inhibitor SQ-22536 alone or with cnr1 deficiency led to reduced MCC populations, suggesting that cnr1 also mediates renal MCC development independently of cAMP signaling. Our findings indicate that cnr1 has a critical role in controlling renal MCC development both via cAMP signaling and an independent pathway, further revealing implications for ciliopathies and renal diseases.

Previously, we reported that mammalian cells, specifically human dermal fibroblasts (HDFs), could be transdifferentiated by lactic acid bacteria (LAB). Later, we observed that HDFs incorporated LAB-derived ribosomes, forming the ribosome-induced cell clusters (RICs) and transdifferentiating into cells derived from all three germ layers. Based on this insight, we hypothesized that incorporating ribosomes into non-mammalian cells could reveal the universality of this mechanism and open the door to commercial applications. Our current study demonstrates that ribosome incorporation can transdifferentiate chick primary muscle-derived cells (CMCs) into adipocytes, osteoblasts, and chondrocytes. Furthermore, the culture medium supernatant from ribosome-incorporated CMCs was found to significantly enhance CMC's proliferation. RNA-seq analysis revealed that RICs-CMC exhibit increased expression of genes related to multi-lineage cell growth. In addition, we developed a novel technological shift in meat production-the "CulNet System"-which replicates organ interactions within mechanical systems for cell-cultured meat production. While significant efforts are still required to implement this technology in a cost-effective manner, we believe that combining the "CulNet System" with ribosome-incorporated multipotent cells that have prolonged culture capability could substantially improve the scalability and cost-effectiveness of cultured chicken meat production. This report highlights a promising approach for cell-culture-based meat production, offering a sustainable alternative to traditional methods.

Inflammatory bowel disease (IBD) results from a breakdown in the symbiotic relationship between the intestinal commensal microflora and the mucosal immune system. Non-toxigenic Bacteroides fragilis, a common human colon symbiote, has been shown to alleviate colitis. However, the underlying mechanisms of this alleviation remain incompletely understood. Herein, it is demonstrated that promoting the secretion of B. fragilis outer membrane vesicles (BfOMVs+) enhances its ability to alleviate dextran sodium sulfate (DSS)-induced colitis, while inhibiting B. fragilis OMV secretion (BfOMVs-) reduces this effect. BfOMVs+ alleviates colitis by inhibiting neutrophil recruitment and neutrophil extracellular trap (NET) formation. Further, B. fragilis OMVs (Bf-OMVs) are isolated and extracted, then administered them intraperitoneally to DSS-induced colitis mice, observing that Bf-OMVs can target intestinal tissues, the spleen, and bone marrow, and they are internalized by neutrophils to inhibit NET formation, thereby alleviating colitis. The expression profile of miRNAs in Bf-OMVs is assessed, revealing that Bf-OMVs are enriched with mmu-miR-like sRNA, miR-5119, which targets and inhibits PD-L1, leading to the suppression of GSDMD-mediated NET release and promoting the proliferation of intestinal stem cells (ISCs), culminating in the alleviation of colitis. These findings provide new insights into the role of B. fragilis OMVs in the pathogenesis and treatment of IBD.

Mesenchymal stem cells (MSCs) have a broad clinical potential, but their selection and expansion on plastic cause unknown purity and phenotypic alterations, reducing therapy efficiency. Furthermore, their behavior in non-adherent conditions during systemic transplantation remains poorly understood. The sphere formation from single cells is commonly used to assess stemness, but MSCs lack this ability, raising questions about their anchorage dependence for proliferation. We investigated whether bone marrow-derived MSCs can complete cytokinesis in non-adherent environments. Primary human and mouse bone marrow-derived MSCs were synchronized in early mitosis using nocodazole and were cultured on soft, rigid, or non-adherent surfaces. Both human and mouse MSCs displayed an ALIX (abscission licensor) recruitment to the midbody 40-90 min post-nocodazole release, regardless of the substrate adherence. Cells maintained for 4hr in the suspension remained viable, and daughter cells rapidly migrated apart upon the re-adhesion to fibronectin-coated surfaces, demonstrating cytokinesis completion in suspension. These findings distinguish MSCs from fibroblasts (which require adhesion for division), provide a more general stemness feature, and suggest that adhesion-independent cytokinesis is a trait relevant to the post-transplantation survival and tissue homing. This property may offer strategies to expand MSCs with an improved purity and functionality and to enhance engraftment by leveraging cell cycle manipulation to promote an early extracellular matrix deposition at target sites.

Human-induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells (HLCs) have been shown to be useful for the development of cell-based regenerative strategies and for modelling drug discovery. However, stem cell-derived HLCs are not identical in nature to primary human hepatocytes (PHHs), which could affect the cell phenotype and, potentially, model reliability. Therefore, we employed the in-depth gene expression profiling of HLCs and other important and relevant cell types, which led to the identification of clear similarities and differences between them at the transcriptional level. Through gene set enrichment analysis, we identified that genes that are critical for immune signalling pathways become downregulated upon HLC differentiation. Our analysis also found that TAV.HLCs exhibit a mild gene signature characteristic of acute lymphoblastic leukaemia, but not other selected cancers. Importantly, HLCs present significant similarity to PHHs, making them genuinely valuable for modelling human liver biology in vitro and for the development of prototype cell-based therapies for pre-clinical testing.

The central nervous system (CNS) is highly susceptible to damage due to its limited ability to regenerate. Injuries to the CNS, whether from trauma, ischemia, or neurodegenerative diseases, disrupt both cellular and vascular structures, leading to immediate (primary) and subsequent (secondary) damage. Primary damage involves the physical disruption of cells and blood vessels, weakening the blood-brain barrier (BBB) and triggering excitotoxicity and calcium overload. Secondary damage develops over hours to days and is marked by ionic imbalance, mitochondrial dysfunction, oxidative stress, and chronic inflammation, which further aggravates tissue damage. Inflammation plays a dual role: acute inflammation helps in repair, while chronic inflammation accelerates neurodegeneration. Microglia and astrocytes play key roles in this inflammatory response, with M1-like microglia promoting pro-inflammatory responses and M2-like microglia supporting anti-inflammatory and repair processes. Neurodegenerative diseases are characterized by the accumulation of misfolded proteins such as Tau, amyloid-beta, TDP-43, and α-synuclein, which impair cellular function and lead to neuronal loss. Neurodegenerative diseases are characterized by the accumulation of misfolded proteins and influenced by genetic risk factors (e.g., APOE4, TARDBP). Despite the CNS's limited regenerative abilities, processes like synaptogenesis, neurogenesis, axonal regeneration, and remyelination offer potential for recovery. Therapeutic approaches aim to target inflammatory pathways, enhance repair mechanisms, and develop neuroprotective treatments to counter excitotoxicity, oxidative stress, and apoptosis. Advances in stem cell therapy, gene therapy, and personalized medicine hold promise for improving outcomes. Future research should focus on combining strategies, utilizing advanced technologies, and conducting translational studies to bridge the gap between preclinical research and clinical application. By better understanding and leveraging the complex processes of CNS injury and repair, researchers hope to develop effective therapies to restore function and enhance the quality of life for individuals with CNS disorders.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder affecting 1-3% of the population globally. Owing to its multifactorial origin, complex genetics, and heterogeneity in clinical phenotypes, it is difficult to faithfully model ASD. In essence, ASD is an umbrella term for a group of individually rare disorders, each risk gene accounting for <1% of cases, threaded by a set of overlapping behavioral or molecular phenotypes. Validated behavioral tests are considered a gold standard for ASD diagnosis, and several animal models (rodents, pigs, and non-human primates) have traditionally been used to study its molecular basis. These models recapitulate the human phenotype to a varying degree and have been indispensable to preclinical research, but they cannot be used to study human-specific features such as protracted neuronal maturation and cell-intrinsic attributes, posing serious limitations to translatability. Human stem cell-based models, both as monolayer 2D cultures and 3D organoids and assembloids, can circumvent these limitations. Generated from a patient's own reprogrammed cells, these can be used for testing therapeutic interventions that are more condition and patient relevant, targeting developmental windows where the intervention would be most effective. We discuss some of these advancements by comparing traditional and recent models of ASD.

Male reproductive aging proceeds gradually and involves complex alterations across germ cells, somatic cells, and the testicular niche. Multi-omics analyses highlight shifts in spermatogonial stem cell dynamics, diminished sperm quantity and quality, and reconfigured support from Sertoli and Leydig cells. These somatic cells show numerical declines and exhibit senescence-associated changes that amplify inflammatory signals and compromise blood-testis barrier integrity. Concurrently, fibrosis and heightened immune cell infiltration disrupt intercellular communication, contributing to further deterioration of spermatogenesis. Epigenetic remodeling-including DNA methylation drift, histone modification imbalances, and altered small non-coding RNA profiles-adds another dimension, reducing sperm integrity and potentially exerting transgenerational effects on offspring health. Observed hormonal changes, such as reduced testosterone and INSL3 production by aging Leydig cells, reflect the additional weakening of testicular function. These multifactorial processes collectively underlie the drop in male fertility and the increased incidence of adverse outcomes, such as miscarriages and developmental anomalies in the offspring of older fathers. Research into mitigation strategies, including interventions targeting senescent cells, oxidative stress, and inflammatory pathways, may slow or reverse key mechanisms of testicular aging. These findings underscore the importance of understanding the molecular hallmarks of male reproductive aging for preserving fertility and safeguarding offspring well-being.

Recent studies on myogenic satellite cells (SCs) in Duchenne muscular dystrophy (DMD) documented altered division capacities and impaired regeneration potential of SCs in DMD patients and animal models. It remains unknown, however, if SC-intrinsic effects trigger these deficiencies at pre-contractile stages of myogenesis rather than resulting from the pathologic environment. In this study, we isolated SCs from a porcine DMD model and age-matched wild-type (WT) piglets for comprehensive analysis. Using immunofluorescence, differentiation assays, traction force microscopy (TFM), RNA-seq, and label-free proteomic measurements, SCs behavior was characterized, and molecular changes were investigated. TFM revealed significantly higher average traction forces in DMD than WT SCs (90.4 ± 10.5 Pa vs. 66.9 ± 8.9 Pa; p = 0.0018). We identified 1390 differentially expressed genes and 1261 proteins with altered abundance in DMD vs. WT SCs. Dysregulated pathways uncovered by gene ontology (GO) enrichment analysis included sarcomere organization, focal adhesion, and response to hypoxia. Multi-omics factor analysis (MOFA) integrating transcriptomic and proteomic data, identified five factors accounting for the observed variance with an overall higher contribution of the transcriptomic data. Our findings suggest that SC impairments result from their inherent genetic abnormality rather than from environmental influences. The observed biological changes are intrinsic and not reactive to the pathological surrounding of DMD muscle.

Arsenic, in the form of inorganic arsenite, is toxic to the kidney and can cause acute kidney injury, manifesting as destruction of proximal tubule cells. Nephron repair is possible through the proliferation of resident tubular progenitor cells expressing CD133 and CD24 surface markers. We simulated regenerative repair in the continued presence of i-As (III) using a cell culture model of a renal progenitor cell line expressing CD133 (PROM1) and CD24. Continued exposure and subculturing of progenitor cells to i-As (III) led to a reduction in the expression of PROM1 and CD24, as well as a decrease in the ability to differentiate into tubule-like structures. Cessation of i-As (III) and recovery for up to three passages resulted in continued repression of PROM1 and reduced ability to differentiate. Chronically exposed cells exhibited an ability to form colonies in soft agar, suggesting neoplastic transformation. Chronically exposed cells also exhibited an induction of CD44, a cell surface marker commonly found in renal cell carcinoma, as well as in tubular repair in chronic renal injury such as chronic kidney disease. These results demonstrate potential adverse outcomes of renal progenitor cells chronically exposed to a nephrotoxicant, as well as in environmental exposure to arsenic.

The adult human heart has a limited ability to regenerate after injury, leading to the formation of fibrotic scars and a subsequent loss of function. In fish, mice, and humans, cardiac remodeling after myocardial injury involves the activation of epicardial and endocardial cells, pericytes, stem cells, and fibroblasts. The heart's extracellular matrix (ECM) plays a significant role in the regeneration and recovery process. The epicardium, endocardium, and pericytes reactivate the embryonic program in response to ECM stimulation, which leads to epithelial-mesenchymal transition, cell migration, and differentiation. This review analyzes the role of ECM in guiding the differentiation or dedifferentiation and proliferation of heart components by comparing significant findings in a zebrafish model with those of mammals.