Search Results (923)

Diabetes mellitus-related morbidity and mortality are primarily caused by long-term complications such as retinopathy, nephropathy, cardiomyopathy, and neuropathy. Diabetic neuropathy (DN) involves the progressive degeneration of axons and nerve fibers due to chronic exposure to hyperglycemia. This metabolic disturbance leads to excessive activation of the glycolytic pathway, inducing oxidative stress and mitochondrial dysfunction, ultimately resulting in nerve damage. There is no specific treatment for painful DN, and new approaches should aim not only to relieve pain but also to prevent oxidative stress and reduce inflammation. Given that existing therapies for painful DN are not effective for diabetic patients, mesenchymal stromal cells (MSCs)-based therapy shows promise for providing immunomodulatory and paracrine regulatory functions. MSCs from various sources can improve neuronal dysfunction associated with DN. Transplantation of MSCs has led to a reduction in hyperalgesia and allodynia, along with the recovery of nerve function in diabetic rats. While the pathogenesis of diabetic neuropathic pain is complex, clinical trials have demonstrated the importance of MSCs in modulating the immune response in diabetic patients. MSCs reduce the levels of inflammatory factors and increase anti-inflammatory cytokines, thereby interfering with the progression of DM. Further investigation is necessary to ensure the safety and efficacy of MSCs in preventing or treating neuropathic pain in diabetic patients. Full article

3D bioprinting has become a valuable tool for studying the biology of solid tumors, including glioblastoma multiforme (GBM). Our analysis of publicly available bulk RNA and single-cell sequencing data has allowed us to define the chemotactic profile of GBM tumors and identify the cell types that secrete particular chemokines in the GBM tumor microenvironment (TME). Our findings indicate that primary GBM tissues express multiple chemokines, whereas spherical monocultures of GBM cells significantly lose this diversity. Subsequently, the comparative analysis of GBM spherical monocultures vs. 3D-bioprinted multicultures of cells showed a restoration of chemokine profile diversity in 3D-bioprinted cultures. Furthermore, single-cell RNA-Seq analysis showed that cells of the perivascular niche (pericytes and endocytes) express multiple chemokines in the GBM TME. Next, we 3D-bioprinted cells from two glioblastoma cell lines, U-251 and DK-MG, alone and as co-cultures with mesenchymal stromal cells (representing cells of the perivascular niche) and assessed the chemokine secretome. The results clearly demonstrated that the interaction of tumors and mesenchymal cells leads to in a significant increase in the repertoire and levels of secreted chemokines under culture in 21% O₂ and 1% O₂. Our study indicates that cells of the perivascular niche may perform a substantial role in shaping the chemokine microenvironment in GBM tumors. Full article

Stem cell therapy in cat patients needs a high quantity of mesenchymal stromal/stem cells (MSCs) requiring in vitro propagation under culture conditions which may potentially impact cellular characteristics and genetic stability. This study aimed to assess the in vitro characteristics and cytogenetic stability of cat adipose tissue-derived MSCs (cAT-MSCs). For this purpose, morphological features, clonogenic potential, and proliferative capacity of cAT-MSCs were assessed at passages 2 (P₂), P₄, and P₆. Multipotency and immunophenotype were evaluated. Cytogenetic analyses were conducted up to P₆. The cAT-MSCs exhibited a spindle-shaped morphology in early passages. The doubling time increased from 2.5 days at P₂ to 9.4 at P₄ and 10.5 at P₆, accompanied by the observation of nuclear abnormalities such as cluster formation, karyorrhexis, karyolysis, and a decline in the mitotic index at P₄. Cells demonstrated multipotency capacity and were CD45−, CD90+, and CD44+. Metaphase analysis at P₂ and P₄ revealed some indications of structural instability such as gaps, breaks, deletions, duplications, and early chromatid segregation, but these alterations did not show an increase across passages. In conclusion, cAT-MSCs decreased their proliferative capacity after P₄, accompanied by morphological alterations and signs of structural instability. Full article

Although therapies based on mesenchymal stromal cells (MSCs) are being implemented in clinical settings, many aspects regarding these procedures require further optimization. Domestic dogs suffer from numerous immune-mediated diseases similar to those found in humans. This study aimed to assess the immunomodulatory activity of canine (c) Wharton jelly (WJ)-derived MSCs and refer them to human (h) MSCs isolated from the same tissue. Canine MSC(WJ)s appeared to be more prone to in vitro aging than their human counterparts. Both canine and human MSC(WJ)s significantly inhibited the activation as well as proliferation of CD4+ and CD8+ T cells. The treatment with IFNγ significantly upregulated indoleamine-2,3-dioxygenase 1 (IDO1) synthesis in human and canine MSC(WJ)s, and the addition of poly(I:C), TLR3 ligand, synergized this effect in cells from both species. Unstimulated human and canine MSC(WJ)s released TGFβ at the same level (p > 0.05). IFNγ significantly increased the secretion of TGFβ in cells from both species (p < 0.05); however, this response was significantly stronger in human cells than in canine cells. Although the properties of canine and human MSC(WJ)s differ in detail, cells from both species inhibit the proliferation of activated T cells to a very similar degree and respond to pro-inflammatory stimulation by enhancing their anti-inflammatory activity. These results suggest that testing MSC transplantation in naturally occurring immune-mediated diseases in dogs may have high translational value for human clinical trials. Full article

Hepatic fibrosis, arising from prolonged liver injury, entails the activation of hepatic stellate cells (HSCs) into myofibroblast-like cells expressing alpha-smooth muscle actin (α-SMA), thereby driving extracellular matrix deposition and fibrosis progression. Strategies targeting activated HSC reversal and hepatocyte regeneration show promise for fibrosis management. Previous studies suggest that extracellular vesicles (EVs) from mesenchymal stromal cells (MSCs) can suppress HSC activation, but ensuring EV purity is essential for clinical use. This study investigated the effects of MSC-derived EVs cultured in chemically defined conditions on liver spheroids and activated HSCs. Umbilical cord- and bone marrow-derived MSCs were expanded in chemically defined media, and EVs were isolated using filtration and differential ultracentrifugation. The impact of MSC-EVs was evaluated on liver spheroids generated in Sphericalplate 5D™ and on human HSCs, both activated by transforming growth factor beta 1 (TGF-β1). MSC-EVs effectively reduced the expression of profibrotic markers in liver spheroids and activated HSCs induced by TGF-β1 stimulation. These results highlight the potential of MSC-EVs collected under chemically defined conditions to mitigate the activated phenotype of HSCs and liver spheroids, suggesting MSC-EVs as a promising treatment for hepatic fibrosis. Full article

Mesenchymal stromal cells (MSCs) and their released factors (secretome) are intriguing options for regenerative medicine approaches based on the management of inflammation and tissue restoration, as in joint disorders like osteoarthritis (OA). Production strategy may modulate cells and secretome fingerprints, and for the latter, the effect of serum removal by starvation used in clinical-grade protocols has been underestimated. In this work, the effect of starvation on the molecular profile of interleukin 1 beta (IL1β)-primed adipose-derived MSCs (ASCs) was tested by assessing the expression level of 84 genes related to secreted factors and 84 genes involved in defining stemness potential. After validation at the protein level, the effect of starvation modulation in the secretomes was tested in a model of OA chondrocytes. IL1β priming in vitro led to an increase in inflammatory mediators’ release and reduced anti-inflammatory potential on chondrocytes, features reversed by subsequent starvation. Therefore, when applying serum removal-based clinical-grade protocols for ASCs’ secretome production, the effects of starvation must be carefully considered and investigated. Full article

Mesenchymal stromal cells (MSCs) can be isolated from various tissues of healthy or patient donors to be retransplanted in cell therapies. Because the number of MSCs obtained from biopsies is typically too low for direct clinical application, MSC expansion in cell culture is required. However, ex vivo amplification often reduces the desired MSC regenerative potential and enhances undesired traits, such as activation into fibrogenic myofibroblasts. Transiently activated myofibroblasts restore tissue integrity after organ injury by producing and contracting extracellular matrix into scar tissue. In contrast, persistent myofibroblasts cause excessive scarring—called fibrosis—that destroys organ function. In this review, we focus on the relevance and molecular mechanisms of myofibroblast activation upon contact with stiff cell culture plastic or recipient scar tissue, such as hypertrophic scars of large skin burns. We discuss cell mechanoperception mechanisms such as integrins and stretch-activated channels, mechanotransduction through the contractile actin cytoskeleton, and conversion of mechanical signals into transcriptional programs via mechanosensitive co-transcription factors, such as YAP, TAZ, and MRTF. We further elaborate how prolonged mechanical stress can create persistent myofibroblast memory by direct mechanotransduction to the nucleus that can evoke lasting epigenetic modifications at the DNA level, such as histone methylation and acetylation. We conclude by projecting how cell culture mechanics can be modulated to generate MSCs, which epigenetically protected against myofibroblast activation and transport desired regeneration potential to the recipient tissue environment in clinical therapies. Full article

Limbal mesenchymal stromal cells (LMSCs) reside in the limbal niche, supporting corneal integrity and facilitating regeneration. While mesenchymal stem/stromal cells (MSCs) are used in regenerative therapies, there is limited knowledge about LMSC subpopulations and their characteristics. This study characterized human LMSC subpopulations through the flow cytometric assessment of fifteen cell surface markers, including MSC, wound healing, immune regulation, ASC, endothelial, and differentiation markers. Primary LMSCs were established from remnant human corneal transplant specimens and passaged eight times to observe changes during subculture. The results showed the consistent expression of typical MSC markers and distinct subpopulations with the passage-dependent expression of wound healing, immune regulation, and differentiation markers. High CD166 and CD248 expressions indicated a crucial role in ocular surface repair. CD29 expression suggested an immunoregulatory role. Comparable pigment-epithelial-derived factor (PEDF) expression supported anti-inflammatory and anti-angiogenic roles. Sustained CD201 expression indicated maintained differentiation capability, while VEGFR2 expression suggested potential endothelial differentiation. LMSCs showed higher VEGF expression than fibroblasts and endothelial cells, suggesting a potential contribution to ocular surface regeneration through the modulation of angiogenesis and inflammation. These findings highlight the heterogeneity and multipotent potential of LMSC subpopulations during in vitro expansion, informing the development of standardized protocols for regenerative therapies and improving treatments for ocular surface disorders. Full article

Skin wounds often form scar tissue during healing. Early intervention with tissue-engineered materials and cell therapies may promote scar-free healing. Exosomes and extracellular vesicles (EV) secreted by mesenchymal stromal cells (MSC) are believed to have high regenerative capacity. EV bioactivity is preserved after lyophilization and storage to enable use in remote and typically resource-constrained environments. We developed a bioprinted bandage containing reconstituted EVs that can be fabricated at the point-of-need. An alginate/carboxymethyl cellulose (CMC) biomaterial ink was prepared, and printability and mechanical properties were assessed with rheology and compression testing. Three-dimensional printed constructs were evaluated for Young’s modulus relative to infill density and crosslinking to yield material with stiffness suitable for use as a wound dressing. We purified EVs from human MSC-conditioned media and characterized them with nanoparticle tracking analysis and mass spectroscopy, which gave a peak size of 118 nm and identification of known EV proteins. Fluorescently labeled EVs were mixed to form bio-ink and bioprinted to characterize EV release. EV bandages were bioprinted on both a commercial laboratory bioprinter and a custom ruggedized 3D printer with bioprinting capabilities, and lyophilized EVs, biomaterial ink, and thermoplastic filament were deployed to an austere Arctic environment and bioprinted. This work demonstrates that EVs can be bioprinted with an alginate/CMC hydrogel and released over time when in contact with a skin-like substitute. The technology is suitable for operational medical applications, notably in resource-limited locations, including large-scale natural disasters, humanitarian crises, and combat zones. Full article

The senescence of bone marrow mesenchymal stromal cells (MSCs) leads to the impairment of stemness and osteogenic differentiation capacity. In a previous study, we screened out stearoyl-CoA desaturase 2 (SCD2), the most evidently changed differential gene in lipid metabolism, using combined transcriptomic and metabolomic analyses, and verified that SCD2 could mitigate MSC senescence. However, the underlying molecular mechanism by which the rate-limiting enzyme of lipogenesis SCD2 manipulates MSC senescence has not been completely understood. In this study, we demonstrate that SCD2 over-expression alleviates MSC replicative senescence and ameliorates their osteogenic differentiation through the regulation of lipogenesis. Furthermore, SCD2 expression is reduced, whereas miR-200c-3p expression is elevated in replicative senescent MSCs. SCD2 is the direct target gene of miR-200c-3p, which can bind to the 3′-UTR of SCD2. MiR-200c-3p replenishment in young MSCs is able to diminish SCD2 expression levels due to epigenetic modulation. In addition, SCD2-rescued MSC senescence and enhanced osteogenic differentiation can be attenuated by miR-200c-3p repletion via suppressing lipogenesis. Taken together, we reveal the potential mechanism of SCD2 influencing MSC senescence from the perspective of lipid metabolism and epigenetics, which provides both an experimental basis for elucidating the mechanism of stem cell senescence and a novel target for delaying stem cell senescence. Full article

Human mesenchymal stromal cells (hMSCs), whether used alone or together with three-dimensional scaffolds, are the best-studied postnatal stem cells in regenerative medicine. In this study, innovative composite scaffolds consisting of a core–shell architecture were seeded with bone-marrow-derived hMSCs (BM-hMSCs) and tested for their biocompatibility and remarkable capacity to promote and support bone regeneration and mineralization. The scaffolds were prepared by grafting three different amounts of gelatin–chitosan (CH) hydrogel into a 3D-printed polylactic acid (PLA) core (PLA-CH), and the mechanical and degradation properties were analyzed. The BM-hMSCs were cultured in the scaffolds with the presence of growth medium (GM) or osteogenic medium (OM) with differentiation stimuli in combination with fetal bovine serum (FBS) or human platelet lysate (hPL). The primary objective was to determine the viability, proliferation, morphology, and spreading capacity of BM-hMSCs within the scaffolds, thereby confirming their biocompatibility. Secondly, the BM-hMSCs were shown to differentiate into osteoblasts and to facilitate scaffold mineralization. This was evinced by a positive Von Kossa result, the modulation of differentiation markers (osteocalcin and osteopontin), an expression of a marker of extracellular matrix remodeling (bone morphogenetic protein-2), and collagen I. The results of the energy-dispersive X-ray analysis (EDS) clearly demonstrate the presence of calcium and phosphorus in the samples that were incubated in OM, in the presence of FBS and hPL, but not in GM. The chemical distribution maps of calcium and phosphorus indicate that these elements are co-localized in the same areas of the sections, demonstrating the formation of hydroxyapatite. In conclusion, our findings show that the combination of BM-hMSCs and PLA-CH, regardless of the amount of hydrogel content, in the presence of differentiation stimuli, can provide a construct with enhanced osteogenicity for clinically relevant bone regeneration. Full article

Muscle-derived mesenchymal stromal cells (mdMSCs) hold great promise in regenerative medicine due to their immunomodulatory properties, multipotent differentiation capacity and ease of collection. However, traditional in vitro expansion methods use fetal bovine serum (FBS) and have numerous limitations including ethical concerns, batch-to-batch variability, immunogenicity, xenogenic contamination and regulatory compliance issues. This study investigates the use of 10% equine platelet lysate (ePL) obtained by plasmapheresis as a substitute for FBS in the culture of mdMSCs in innovative 2D and 3D models. Using muscle microbiopsies as the primary cell source in both models showed promising results. Initial investigations indicated that small variations in heparin concentration in 2D cultures strongly influenced medium coagulation with an optimal proliferation observed at final heparin concentrations of 1.44 IU/mL. The two novel models investigated showed that expansion of mdMSCs is achievable. At the end of expansion, the 3D model revealed a higher total number of cells harvested (64.60 ± 5.32 million) compared to the 2D culture (57.20 ± 7.66 million). Trilineage differentiation assays confirmed the multipotency (osteoblasts, chondroblasts and adipocytes) of the mdMSCs generated in both models with no significant difference observed. Immunophenotyping confirmed the expression of the mesenchymal stem cell (MSC) markers CD-90 and CD-44, with low expression of CD-45 and MHCII markers for mdMSCs derived from the two models. The generated mdMSCs also had great immunomodulatory properties. Specific immunological extraction followed by enzymatic detection (SIEFED) analysis demonstrated that mdMSCs from both models inhibited myeloperoxidase (MPO) activity in a strong dose-dependent manner. Moreover, they were also able to reduce reactive oxygen species (ROS) activity, with mdMSCs from the 3D model showing significantly higher dose-dependent inhibition compared to the 2D model. These results highlighted for the first time the feasibility and efficacy of using 10% ePL for mdMSC expansion in novel 2D and 3D approaches and also that mdMSCs have strong immunomodulatory properties that can be exploited to advance the field of regenerative medicine and cell therapy instead of using FBS with all its drawbacks. Full article

Head and neck cancer (HNC) entails a heterogenous neoplastic disease that arises from the mucosal epithelium of the upper respiratory system and the gastrointestinal tract. It is characterized by high morbidity and mortality, being the eighth most common cancer worldwide. It is believed that the mesenchymal/stem stromal cells (MSCs) present in the tumour milieu play a key role in the modulation of tumour initiation, development and patient outcomes; they also influence the resistance to cisplatin-based chemotherapy, the gold standard for advanced HNC. MSCs are multipotent, heterogeneous and mobile cells. Although no MSC-specific markers exist, they can be recognized based on several others, such as CD73, CD90 and CD105, while lacking the presence of CD45, CD34, CD14 or CD11b, CD79α, or CD19 and HLA-DR antigens; they share phenotypic similarity with stromal cells and their capacity to differentiate into other cell types. In the tumour niche, MSC populations are characterized by cell quiescence, self-renewal capacity, low reactive oxygen species production and the acquisition of epithelial-to-mesenchymal transition properties. They may play a key role in the process of acquiring drug resistance and thus in treatment failure. The present narrative review examines the links between MSCs and HNC, as well as the different mechanisms involved in the development of resistance to current chemo-radiotherapies in HNC. It also examines the possibilities of pharmacological targeting of stemness-related chemoresistance in HNSCC. It describes promising new strategies to optimize chemoradiotherapy, with the potential to personalize patient treatment approaches, and highlights future therapeutic perspectives in HNC. Full article

Our study explores the role of cancer-derived extracellular exosomes (EXs), particularly focusing on collagen alpha-3 (VI; COL6A3), in facilitating tumor dissemination and metastasis in epithelial ovarian cancer (EOC). We found that COL6A3 is expressed in aggressive ES2 derivatives, SKOV3 overexpressing COL6A3 (SKOV3/COL6A3), and mesenchymal-type ovarian carcinoma stromal progenitor cells (MSC-OCSPCs), as well as their EXs, but not in less aggressive SKOV3 cells or ES2 cells with COL6A3 knockdown (ES2/shCOL6A3). High COL6A3 expression correlates with worse overall survival among EOC patients, as evidenced by TCGA and GEO data analysis. In vitro experiments showed that EXs from MSC-OCSPCs or SKOV3/COL6A3 cells significantly enhance invasion ability in ES2 or SKOV3/COL6A3 cells, respectively (both, p <0.001). In contrast, ES2 cells with ES2/shCOL6A3 EXs exhibited reduced invasion ability (p < 0.001). In vivo, the average disseminated tumor numbers in the peritoneal cavity were significantly greater in mice receiving intraperitoneally injected SKOV3/COL6A3 cells than in SKOV3 cells (p < 0.001). Furthermore, mice intravenously (IV) injected with SKOV3/COL6A3 cells and SKOV3/COL6A3-EXs showed increased lung colonization compared to mice injected with SKOV3 cells and PBS (p = 0.007) or SKOV3/COL6A3 cells and PBS (p = 0.039). Knockdown of COL6A3 or treatment with EX inhibitor GW4869 or rapamycin-abolished COL6A3-EXs may suppress the aggressiveness of EOC. Full article

Research question: Theca interna cells (TICs) are an indispensable cell source for ovarian follicle development and steroidogenesis. Recent studies have identified theca stem cells (TSCs) in both humans and animals. Interestingly, TSCs express mesenchymal stem cell (MSC)-related markers and can differentiate into mesenchymal lineages. MSCs are promising for tissue engineering and regenerative medicine due to their self-renewal and differentiation abilities. Therefore, this study investigated the potential origin of TICs from MSCs. Design: Whole ovaries from postmenopausal organ donors were obtained, and their cortex was cryopreserved prior to the isolation of stromal cells. These isolated cells were differentiated in vitro to TICs using cell media enriched with various growth factors and hormones. Immunocytochemistry, an enzyme-linked immunosorbent assay, flow cytometry, and reverse transcription–quantitative polymerase chain were employed at different timepoints. Data were analyzed using one-way ANOVA. Results: Immunocytochemistry showed an increase in TIC markers from day 0 to day 8 and a significant rise in MSC-like markers on day 2. This corresponds with rising androstenedione levels from day 2 to day 13. Flow cytometry identified a decreasing MSC-like cell population from day 2 onwards. The CD13+ cell population and its gene expression increased significantly over time. NGFR and PDGFRA expression was induced on days 0 and 2, respectively, compared to day 13. Conclusions: This study offers insights into MSC-like cells as the potential origin of TICs. Differentiating TICs from these widely accessible MSCs holds potential significance for toxicity studies and investigating TIC-related disorders like polycystic ovary syndrome (PCOS). Full article