STEM CELLS最新文献

Mesenchymal stem cells (MSCs) have been widely studied for treating immune-mediated diseases due to their immunomodulatory abilities. Recent studies have shown that priming MSCs with inflammatory cytokines can enhance these functions, yet the optimal priming conditions for canine MSCs remain poorly defined. In this study, we investigated the effects of priming canine adipose tissue-derived MSCs (cAMSCs) with inflammatory cytokines IFN-γ, TNF-α, and IL-17 at various concentrations (10, 20, and 50 ng/mL) to evaluate their immunomodulatory and migratory capacities. Of the 3 cytokines evaluated, only IFN-γ priming significantly enhanced the expression of immunosuppressive genes IDO and PD-L1, and robustly suppressed T-cell proliferation across all concentrations compared to naïve cAMSCs in both direct co-culture and indirect (conditioned medium) assays. TNF-α priming significantly increased HGF expression and promoted cell cycle progression, while IL-17 priming upregulated COX2 and TGF-β expression; however, both exhibited limited immunomodulatory effects compared to IFN-γ. In addition, IFN-γ induced strong expression of adhesion and migration-related genes, including E-CADHERIN, ICAM1, and VCAM1, and promoted cAMSCs migration in a wound healing assay. Despite increasing MHC II, IFN-γ did not induce CD80, preserving the low immunogenic profile of cAMSCs. These findings support IFN-γ priming as the most effective strategy to enhance the immunomodulatory and migratory functions of cAMSCs, offering substantial potential for MSC-based therapies in veterinary medicine.

The maintenance of corneal epithelial homeostasis relies on limbal stem cells (LSCs) located at the limbus. Although short-term cultured LSC transplantation effectively treats LSC deficiency, prolonged culture leads to stemness loss and abortive colony formation, and the mechanisms remain elusive. In this study, we employed single-cell transcriptomics to investigate LSC population dynamics and changes in gene expression during extended serial culture. Transcriptomic data from 22 708 cells revealed 19 clusters, identifying 3 distinct limbal progenitor populations (Progenitors 1-3) with unique transcriptional profiles and cell division kinetics. All progenitor subgroups expressed stemness-related genes such as ANLN, AURKB, and HMGB2 and were detected at all stages of the cell cycle. Notably, Progenitor3 exhibited the highest levels of genes associated with stemness and the G2/M checkpoint, including ANLN, PLK1, AURKA, HMGB2, and TOP2A, and had the largest proportion of cells in G2/M. Progenitor2 was marked by histone H1 expression, while Progenitor1 displayed distinctive cell cycle kinetics. Despite stable proportions of the three progenitor populations throughout prolonged passaging, mitochondrial gene downregulation, and ribosomal gene upregulation were observed. Treatment with the small molecule RepSox partially preserved LSC maintenance in long-term culture by inhibiting the epithelial-mesenchymal transition program and modulating energy and metabolic pathways. These findings provide insights for optimizing in vitro LSC expansion for cell-based therapies.

The development of committed erythroid progenitors and their continued maturation into erythrocytes requires the cytokine erythropoietin (Epo). Here, we describe the immunophenotypic identification of a CD34- colony-forming unit-erythroid (CFU-E) progenitor subtype, termed late CFU-E (lateC), that arises in an Epo-dependent manner during human early erythropoiesis (EE). LateC cells lack CD235a (glycophorin A) but have high levels of CD71 and CD105, characterized as Lin-CD123-CD235a-CD49d+CD117+CD34-CD71hiCD105hi. Analysis of ex vivo cultures of bone marrow (BM) CD34+ cells showed that acquisition of the CD71hiCD105hi phenotype in lateC occurs through the formation of four other EE subtypes. Of these, two are CD34+ burst-forming unit-erythroid (BFU-E) cells, distinguishable as CD71loCD105lo early BFU-E (earlyB) and CD71hiCD105lo late BFU-E (lateB), and two are CD34- CFU-E, also distinguishable as CD71loCD105lo early CFU-E (earlyC) and CD71hiCD105lo mid CFU-E (midC). The EE transitions are accompanied by a rise in CD36 expression, such that all lateC cells are immunophenotypically CD36+. Patterns of CD34, CD36, and CD71 indicate two differentiation routes-in one earlyB lose CD34 to form earlyC, and in another, earlyB gain CD36 and CD71hi expression prior to losing CD34 to form midC, bypassing the earlyC stage. Regardless of the route, the transition from midC to lateC requires Epo. All five EE subtypes could be prospectively detected in human BM cells and, upon isolation and reculture, exhibited the potential to continue differentiating along the erythroid trajectory. Finally, we find that all five EE populations can also be detected in cultures of cord blood-derived CD34+ cells at levels similar to those observed in BM CD34+ cell cultures.

Introduction: Systemic lupus erythematosus (SLE) is driven by abnormal type-I and -II interferon activation, affecting a variety of immunocompetent cells. Mesenchymal stromal cells (MSCs) can modulate inflammation but often lack consistent potency. We developed HXB-319, an MSC-based therapy targeting inflammatory pathways in SLE. Previously, HXB-319 was shown to reduce alveolar hemorrhage in an SLE model. Here, we report its effects in a model of SLE that progresses to end stage kidney disease.

Materials and methods: SLE-like disease was induced via intraperitoneal (IP) pristane injection in female BALB/cJ mice, followed by treatment with naïve MSCs or HXB-319. Over 9 months, survival and proteinuria were monitored. Upon euthanasia, kidneys were analyzed for histopathology and gene expression, splenocytes for immune subsets by flow cytometry, and serum for autoantibodies, growth factors, and cytokines.

Results: HXB-319 significantly altered plasmacytoid dendritic cells, CD4+PD-L1+ cells, and both CD4+ and CD8+ RORγt+ (Th17 cells) subsets. HXB-310 lowered IFN-γ (P < 0.001), IL-17A (P = 0.01), BAFF (P < 0.05), and anti-dsDNA (P < 0.05), compared to untreated mice. HXB-319, but not naïve MSCs, significantly improved survival, halted progression of kidney disease, and stabilized proteinuria (all P < 0.05).

Conclusion: HXB-319 demonstrates potential for mitigating SLE-associated glomerulonephritis, improving survival, and reducing proteinuria and glomerulosclerosis.

The hypothalamus-pituitary-adrenal (HPA) axis is crucial for energy metabolism, cardiovascular function, and stress response. Importantly, neuronal signaling circuits in the hypothalamus, along with hormones released from the pituitary and adrenal gland, must adapt to physiological demands or pathological conditions. Stem and progenitor cells are pivotal in this regulation, either by giving rise to distinct cell types or by interacting with progenitor or hormone-producing cells. While lineage-tracing studies in rodent models have explored the role of stem cells in the HPA axis, our understanding of the mechanisms underlying this dynamic tissue plasticity remains limited, especially in humans. Moreover, single-cell RNA sequencing has revealed significant heterogeneity among stem cell populations in the HPA-axis, raising questions about the functional relevance of individual subclusters during development and adulthood. In this concise review, we summarize current knowledge on stem cells in the HPA axis, focusing on their origins, localization of different stem cell populations, and sex-specific activity in maintaining tissue integrity. We further address their role under pathophysiological conditions, including metabolic disease, cancer, and stress. Lastly, we discuss emerging strategies for replacing lost or damaged stem or progenitor cells during aging, highlighting recent achievements in the in vitro differentiation of hypothalamic, pituitary, and adrenal stem cells.

Residual beta cell function has been documented in "medalist" patients who have lived with Type 1 diabetes (T1D) for >50 years. In addition, endocrine cell neogenesis first occurs in the developing human embryo from progenitor cells derived from pancreatic ductal epithelial structure. Thus, beta cell conversion from a dormant epithelial precursor remains a promising approach to regenerate islets during T1D. We have previously shown that intra-pancreatic (iPan) injection of Wnt pathway-stimulated conditioned media (Wnt+ CdM) generated from human bone marrow-derived multipotent stromal cells (MSC) contained islet regenerative factors that reduced hyperglycemia and recovered beta cell mass in streptozotocin-treated mice. However, the endogenous source of regenerated beta cells remains unknown. Herein, we employed cytokeratin 19 (CK19)-CreERT Rosa26-mTomato lineage-tracing mice to assess the endocrine conversion of CK19+ cells during MSC CdM-induced islet regeneration. Mice iPan-injected with Wnt+ CdM demonstrated reduced blood glucose levels and improved glucose tolerance compared to mice injected with unconditioned basal media. CdM-injected mice also showed increased islet number and beta cell mass, as well as CK19+ cells within regenerating islets. The frequency of insulin + cells that co-expressed tdTomato within dissociated pancreas samples observed via flow cytometry was 5-fold higher in Wnt+ CdM-injected mice (~5%) compared to basal media-injected controls (~1%). Collectively, in vivo lineage tracing revealed conversion of CK19+ cells to functional beta cells partially contributed to islet regeneration induced by Wnt-activated MSC CdM. Future studies are required to delineate alternate cell types and mechanisms participating in islet regeneration induced by direct delivery of MSC-CdM.

A systems-level understanding of immunomodulatory, regenerative, and pro-/antifibrosis functions of mesenchymal stromal cells (MSCs) is critical to advance MSCs as a viable therapeutic option. Given the complexity of MSCs and their interactions with microenvironmental cells, a systems biology approach may enable such understanding to achieve practical objectives such as target identification, combination therapeutics, clinical strategies, and avoidance of adverse effects. In this study, a molecular systems architecture of MSCs microenvironment is developed to organize the complexity of biomolecular interactions between MSCs and other microenvironmental cells. This architecture provides a visual mapping of MSC interactions, identifies the complex crosstalk between MSCs and cells in the microenvironment, reveals potential targets, and offers a framework for creating future predictive, quantitative computational (in silico) models of the MSC microenvironment. The development of combination therapeutics, clinical strategies to improve therapeutic efficacy, and avoidance of adverse effects can be facilitated by such in silico models. However, it must all begin with a molecular systems architecture of MSCs-the objective and result of this study.

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are a valuable cell type for studying human cardiac health and disease in vitro. However, it is not known whether hiPSC-CMs display sex dimorphism and therefore whether sex should be incorporated as a biological variable in in vitro studies that include this cell type. To date, the vast majority of studies that utilize hiPSC-CMs do not include both male and female sex nor stratify results based on sex because it is challenging to amass such a cohort of cells. Here, we generated 3 female and 3 male hiPSC lines from adult left ventricular cardiac fibroblasts as a resource for studying sex differences in in vitro cardiac models. We used this resource to generate hiPSC-CMs and maintained them in basal media without exogenous hormones. Functional assessment of CMs showed enhanced calcium handling in female-derived hiPSC-CMs relative to male. Bulk RNA sequencing revealed over 300 differentially expressed genes (DEGs) between male and female hiPSC-CMs. Gene ontology analysis of DEGs showed distinct differences in pathways related to cardiac pathology including cell-cell adhesion, metabolic processes, and response to ischemic stress. Differential expression of the sodium channel auxiliary unit SCN3B was found and validated through patch-clamp measurements of sodium currents, showing increased peak amplitude and window current in female hiPSC-CMs. These findings highlight the importance of considering sex as a variable when conducting studies to evaluate aspects of human cardiac health and disease related to CM function.

Bone marrow mesenchymal stem cells (BMSCs) have chondrogenic differentiation potential to treat cartilage injury. N6 methyladenosine (m6A), one of the most prevalent mRNA modifications, has been reported to be crucial in cartilage disease. Herein, we further investigated the effects and underlying mechanisms in the modification of m6A on the chondrogenic differentiation of MSCs. This study showed that the m6A level was decreased in the chondrogenic differentiation of MSCs and m6A mRNA demethylation fat mass and obesity-associated protein (FTO) played an important role in these processes. The overexpression of FTO has been demonstrated to improve the levels of chondrogenic markers. We confirmed that FTO directly bound to SMAD3 mRNA and increased its demethylation, which promoted the chondrogenic differentiation of MSCs. We further indicated that the m6A "reader" YTHDF2 was probably related to the chondrogenic differentiation of MSCs. SiFTO attenuated the SiYTHDF2-increased mRNA stability of SMAD3, leading to the declining levels of chondrogenic markers. Collectively, these results reveal FTO could act as an important mediator of SMAD3 mRNA demethylation and improve the chondrogenic differentiation of MSCs.

Background: Engaging in appropriate exercise is advantageous for our well-being. We investigated whether exercise could affect the paracrine function of BMSCs and whether exosomes derived from treadmill exercise-trained mouse (Exo-tread) BMSCs could engender more pronounced effects on wound healing.

Methods: First, the effects of treadmill exercise on mouse BMSCs biological functions, exosomes secretion quantity, and identification were assessed. Furthermore, we assessed the effects of Exo-tread on M1 macrophage by qPCR and FCM in vitro. Additionally, the expressions and phosphorylation status of p65 and p38 proteins were analyzed via Western blotting. For the in vivo component, we induced wound models of mice. Subsequently, we assessed the effects of Exo-tread using various methodologies including imaging, H&E, Masson, immunohistochemical, and immunofluorescence staining. To demonstrate whether Exo-tread could act through macrophages, we further depleted mouse macrophages.

Results: Exercise accelerated the proliferation of BMSCs and the secretion of exosomes. In vitro, Exo-tread markedly decreased the expression of inflammatory factors while concurrently suppressing M1 polarization in mouse peritoneal macrophages compared with the Exo-ctrl group. These observed effects were potentially mediated by the reduction in the M1 polarization ratio, achieved through the inhibition of p65 and p38 phosphorylation. Similarly, in vivo experiments demonstrated that Exo-tread significantly enhanced wound healing by attenuating inflammatory cytokines and M1 macrophages.

Conclusions: Exo-tread facilitates wound healing by mitigating the inflammatory response, achieved through a reduction in the M1 polarization ratio.