Stem Cell Research & Therapy最新文献

Aim: Circular RNAs (circRNAs) have been identified as key regulators in inflammatory diseases, yet their function in pulpitis is unclear. This study investigates their potential role in the progression of pulpitis.

Methodology: Microarray and single-cell RNA sequencing were applied to assess DNA damage responses (DDR) in inflammatory pulp and its derived stem cells, respectively. qRT-PCR and Western blot were employed to detect the DNA double-strand break (DSB) marker γ-H2AX and inflammatory cytokines in pulp tissue. Bioinformatics analysis was used to identify upregulated circRNAs in inflamed DPSCs. Functional assays were performed to assess the impact of circ_0042103 on LPS-driven cellular damage and inflammation in DPSCs. The interaction between circ_0042103 and TAF15 was investigated using RNA FISH, pulldown, and nuclear-cytoplasmic fractionation assays. Transfection with circ_0042103/TAF15-siRNA in DPSCs was carried out to evaluate activation of the nucleotide excision repair (NER) pathway and its regulatory effects on DNA damage and inflammation.

Results: DDR was activated in both pulpitis and inflamed DPSCs. DNA damage showed a positive correlation with inflammation in pulpitis. In vitro, circ_0042103 upregulation amplified LPS-stimulated DDR and inflammatory signaling, whereas its knockdown alleviated both effects. Mechanistically, circ_0042103 bound TAF15, leading to decreased levels of the NER-related proteins (ERCC1 and PCNA) and increased DNA damage and inflammation.

Conclusion: By interacting with TAF15, circ_0042103 reduces the levels of the NER-related proteins ERCC1 and PCNA, leading to increased DNA damage and inflammation in hDPSCs, thereby defining a circ_0042103/TAF15/NER axis in pulpitis progression.

Background: The human endometrium is a regenerative tissue essential for fertility, but pathological conditions like Asherman syndrome, endometrial atrophy, and thin endometrium can impair its function. Current therapies lack efficacy, driving demand for innovative regenerative therapies. In this context, endometrial-derived hydrogels and organoids have shown promise individually for tissue regeneration, but their combined therapeutic potential has not been previously evaluated in vivo. This study explores a dual regenerative strategy combining a hybrid hydrogel - composed of synthetic PuraMatrix^® and endometrial extracellular matrix hydrogel - with human endometrial organoids in an immunocompetent murine model with uterine damage.

Methods: Endometrial damage model was established in female C57BL/6 mice (n = 46) via uterine injury using 70° ethanol. After 4 days of endometrial damage, human endometrial organoids were co-injected with the hybrid hydrogel into the uterine horns. Two weeks post-injection, a subset of mice (n = 25) was sacrificed for biocompatibility, histological, and transcriptomic analyses. Functional recovery of the endometrium was assessed in the remaining animals (n = 21) through fertility outcome evaluation. For endometrial regeneration analyses, normally distributed data were analyzed by one-way ANOVA and Tukey's multiple comparisons, while non-normally distributed data were analyzed by the Kruskal-Wallis test with Dunn's multiple comparisons. For fertility outcomes, t-test or Mann-Whitney U tests for 2-by-2 comparisons were performed.

Results: Histological and molecular analyses revealed that the therapy improved endometrial thickness, gland density, and vascularization, and reduced fibrosis and ferroptosis, aligning tissue characteristics closer to healthy controls. However, fertility outcomes were not fully restored, potentially due to the persistence of the synthetic component of the hybrid hydrogel. Thus, further studies are needed to confirm complete hydrogel resorption and its impact on fertility restoration.

Conclusions: In conclusion, this study demonstrates the biocompatibility and regenerative potential of human endometrial organoids delivered within the hybrid hydrogel, highlighting a promising strategy for endometrial regeneration.

Mesenchymal stem cells (MSCs) have been extensively investigated and applied in autoimmune and inflammatory diseases owing to their tissue-repair capacity and immunomodulatory properties, and they hold significant promise in cellular immunotherapy. However, therapeutic outcomes of stem-cell-based immunotherapy can be inconsistent, because the immunomodulatory effects of MSCs may undergo dynamic shifts in response to changes in the microenvironment. Research has shown that the inflammatory state of the tissue microenvironment affects the tendency of MSCs to regulate immunity. Under highly inflammatory conditions, MSCs tend to exert immunosuppressive functions, while under low inflammatory conditions, MSCs tend to exhibit immune support. In addition, the physical culture method, pretreatment conditions, and tissue source of stem cells all affect the direction of their immune regulation. Therefore, gaining a deep understanding of the immune regulatory mechanisms of MSCs and their influencing factors is crucial for optimizing their application in stem cell immunotherapy and improving treatment outcomes. This review first explores the potential factors influencing MSCs' bidirectional immune regulation, then discusses the mechanisms underlying the immunosupportive and immunosuppressive effects of MSCs, and finally briefly describes the role of MSCs' bidirectional immune regulation function in disease treatment.

Spinal cord injury (SCI) remains a significant global health challenge with limited effective therapeutic options. Exosomes derived from mesenchymal stem cells (MSCs) have emerged as promising neuroprotective agents due to their biocompatibility and immunomodulatory properties. This study investigated the therapeutic potential of hypoxia-conditioned bone marrow MSC (BMSC)-derived exosomes in both in vitro and in vivo SCI models. Hypoxic preconditioning significantly enriched miR-615-3p in bone marrow mesenchymal stem cell (BMSC)-derived exosomes. In spinal neuron injury models, hypoxic exosomes enhanced cell viability, reduced apoptosis, and ameliorated dysfunction of the mitochondria-associated endoplasmic reticulum membranes (MAMs). Mechanistically, miR-615-3p directly targeted and suppressed phosphodiesterase 4 C (PDE4C), activating the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) pathway. This in turn modulated calcium signaling, attenuated mitochondrial calcium overload, and reduced endoplasmic reticulum stress (ERS). In a mouse model of SCI, short-term treatment with hypoxic exosomes promoted functional recovery within a 14-day post-injury period, as evidenced by improved locomotor performance, reduced lesion volume, attenuated tissue edema, and decreased inflammatory cell infiltration. Furthermore, in vivo administration of hypoxic exosomes upregulated miR-615-3p and downregulated PDE4C expression in injured spinal cord tissues. These results demonstrate that hypoxia-conditioned BMSC-derived exosomes exert neuroprotective effects via the miR-615-3p/PDE4C axis, highlighting their potential as a novel therapeutic strategy for SCI by targeting calcium homeostasis and mitochondrial-ER dysfunction. These findings demonstrate the short-term therapeutic potential of hypoxia-conditioned exosomes in SCI. However, further preclinical studies, including long-term follow-up to assess the durability of recovery and potential late-onset effects, alongside clinical validation, are warranted before clinical translation.

Background: Exosomes derived from mesenchymal stem cells (MSCs) are increasingly recognized as promising mediators of tissue regeneration. However, most studies have focused on exosomes from purified MSC populations, and the regenerative relevance of exosomes secreted by fibroblast-dominant oral cell populations remains poorly understood. This study aimed to characterize the cell type-specific miRNA-mRNA regulatory features of exosomes released by gingival fibroblasts, periodontal ligament fibroblasts, and dental pulp fibroblasts, and to evaluate their potential links to MSC-like molecular programs.

Methods: Fibroblast-rich cell populations were isolated from gingiva, periodontal ligament, and dental pulp tissue from the same extracted teeth, without MSC purification. Bulk RNA-seq was performed on the cells, and exosomes were collected from culture supernatants for miRNA-seq, small RNA-seq, and RNA-seq (n = 3 donors). Cell type-specific miRNA-mRNA regulatory axes were identified based on inverse expression patterns and confirmed using experimentally validated interactions from miRTarBase.

Results: Cellular transcriptomic profiling showed that dental pulp fibroblasts expressed higher levels of genes associated with stemness, osteogenic potential, and metabolic regulation, whereas gingival and periodontal ligament fibroblasts exhibited signatures related to inflammation, vesicle trafficking, and tissue homeostasis. Exosomal RNA profiling revealed distinct regulatory modules for each fibroblast type: gingival fibroblast-derived exosomes exhibited a miR-660-5p/XKR7 axis associated with apoptosis regulation; periodontal ligament fibroblast-derived exosomes displayed a miR-199a-5p/COL19A1 axis linked to extracellular matrix remodeling; and dental pulp fibroblast-derived exosomes contained multiple MSC-associated regulatory axes, including miR-1307-3p and miR-30b-3p targeting SNRPD1, miR-493-5p targeting HMGXB4, and miR-26b-5p targeting MB-HSPD1.

Conclusions: Exosomes derived from fibroblast-rich oral cell populations display distinct molecular signatures reflective of their tissue origins. Notably, exosomes from dental pulp fibroblasts exhibit MSC-like regulatory features. These findings suggest that exosomes from mixed fibroblast cultures, without requiring MSC purification, may hold promise as practical, cell-free regenerative tools, pending future functional validation.

Background: The development of vascular calcification (VC) in diabetes is closely related to the endothelial-to-mesenchymal transition (EndMT). We found that microRNA-32-5p (miR-32) was elevated in the plasma of calcification patients. However, it is unclear whether miR-32 mediates the function of bone marrow mesenchymal stem cell-derived extracellular vesicles (BMSC-EVs) in type 2 diabetes (T2D) VC.

Methods: BMSC-EVs were characterized by TEM, NTA, Western blotting, and confocal microscopy. Alizarin Red and ALP staining assessed the severity of VC. qRT-PCR and Western blotting evaluated the expression of BMP2, RUNX2, GPX4, SLC7A11, VE-cadherin, and N-cadherin, while immunofluorescence was used for detecting VE-cadherin and N-cadherin. In vivo validation was performed using miR-32^-/- and ApoE^-/- mice. RNA sequencing (RNA-seq) and bioinformatics analysis was conducted to explore underlying mechanisms.

Results: We demonstrated that BMSC-EVs attenuate VC in endothelial cells (ECs) and inhibit EndMT. In vivo, histological analysis showed that treatment with BMSC-EVs significantly reduced the severity of VC associated with T2D. Notably, knockout of miR-32 further enhanced the inhibitory effect of BMSC-EVs on VC. Mechanistically, transcriptomic and functional analyses suggest that the protective effect of BMSC-EVs on VC is associated with regulation of the MAPK/FoxO signaling pathway, potentially mediated by modulation of ferroptosis.

Conclusion: These findings demonstrate that BMSC-EVs attenuate T2D-associated VC, partially through miR-32-mediated suppression of EC ferroptosis.

Background: Bionic treatment is a strategy designed to facilitate functional recovery after clinical spinal cord injury (SCI) by emulating the natural morphological structure and regeneration process. We used zebrafish model, an animal with remarkable regenerative capabilities to investigate the regulatory pattern of spinal vascular regeneration following SCI, with the hope of providing inspirations for the development of bionic SCI treatment.

Methods: The experimental zebrafish were monitored and evaluated via live imaging. We first determined the formation time of the spinal perineural vessel plexus (PNVP) and used this as the timepoint to initiate SCI. Subsequently, a SCI model was established to observe the pattern of vascular repair without intervention; Furthermore, radial glial (RGs) of Tg(gfap: NTR-mCherry) report line fish were chemically ablated using metronidazole (Mtz) or nitrofuropyrinol (Nfp). We assessed the patterns of vascular repair, the vascular coverage of the injured area, and the number of vascular endothelial cells (ECs). Concomitantly, by analyzing the expression profile of vascular endothelial growth factor aa (Vegfaa) in the injured region following RGs ablation, and leveraging a public available single-cell sequencing dataset, we postulated the potential downstream pathways involved. The functional relevance of these pathways was finally evaluated by applying specific inhibitors.

Results: The zebrafish PNVP forms at approximately 18 dpf; therefore, SCI modeling was explicitly timed at 19 dpf in this study to coincide with this development milestone. In the Tg(gfap: NTR-mCherry) report line, RGs were successfully ablated using either Mtz or Nfp. Following ablation, both vascular coverage in the injured area and the number of ECs were significantly reduced in the Mtz/Nfp + SCI group compared to the DMSO + SCI group. Moreover, The vegfaa reporter line revealed a notable decline in vegfaa signal within the injured region post-ablation, suggesting its involvement in the repair process. This implication was further supported by inhibitor experiments, where intervention against the Notch and PI3K/Akt-mTOR pathways significantly altered the extend of vascular repair, indicating a potential correlation between these pathways and RGs-regulated vascular repair.

Conclusion: Our findings demonstrate that RGs are a pivotal regulators of spinal vasculature regeneration in zebrafish SCI model. The underlying mechanisms may involve the Vegfa-PI3K/Akt-mTOR and Notch signaling pathways. Therefore, it can be postulated that pro-vascular repair therapy in mammals following SCI could potentially be achieved by therapeutically mimicking pro-regenerative functions of RGs.

Friedreich's ataxia (FRDA) is an inherited, autosomal recessive, multisystem disorder that primarily manifests in children and affects the nervous system and the heart. FRDA is caused by an expansion of GAA repeats in the first intron of the frataxin (FXN) gene. The expansion disrupts transcription of FXN, resulting in significantly decreased FXN expression in FRDA patients' tissues. Frataxin is involved in biosynthesis of iron-sulfur (Fe-S) clusters, which are critical for the function of the electron transport chain and many metabolic enzymes. Frataxin deficiency leads to reduced energy production and accumulation of iron in mitochondria that exacerbates oxidative stress. Despite significant advancements in the field, FXN cellular functions and underlying pathological mechanisms of FXN deficiency in cell-type specific contexts remain to be elucidated. Inaccessibility to the most vulnerable cell types in FRDA patients, including neurons, cardiomyocytes, and β-cells, largely accounts for these limitations. Significant progress in recent years regarding the derivation and differentiation of human pluripotent stem cells (hPSCs), along with breakthroughs in gene editing technologies, enables the generation of patient-derived and isogenic control disease-relevant cell types and organoid-like structures as platforms for studying disease mechanisms and for drug discovery. Herein, we first provide an overview of hPSC derivation and intrinsic properties of these cells. We then discuss current advances and limitations of hiPSC-based cell models for FRDA. We also highlight the need to further refine and develop these in vitro cell models for pre-clinical advancement of therapeutic approaches for FRDA.

Human neural stem cells (hNSCs) are promising candidates for regenerative medicine due to their self-renewal capacity, differentiation potential, and ability to modulate inflammation. However, several reports showed that the regenerative properties of stem cells are tied to the extracellular vesicles (EVs) they secrete. This study aimed at characterizing hNSCs produced under Good Manufacturing Practice (GMP) conditions and at elucidating the molecular and functional properties of their secreted extracellular vesicles (hNSC-EVs). hNSCs were first assessed for proliferation, and differentiation potential, showing a stable growth profile and expression of neural stem cell markers. High-resolution proteomic analysis identified over 5000 proteins, with about 40% overlap with previous NSCs studies. hNSCs expressed mostly markers for different cell lineage precursors. The molecular characterization of hNSC-derived EVs (hNSC-EVs) showed a size distribution, as measured by nanoparticle tracking analysis, ranging from 140 to 200 nm and an enrichment in EV markers, detected by western blotting. Functional analyses showed that hNSC-EVs, reduce nitric oxide generation and inducible nitric oxide expression in LPS-treated microglial cells and inhibit caspase-1 activation in monocytic cell models through uptake-dependent and independent mechanism, respectively. Our findings show that hNSC possess a strong stemness signature and secrete EVs with immunomodulatory properties, suggesting the worth of hNSC-EVs as either alternative to cell-based therapies or primer to boost anti-inflammatory properties of hNSCs in the treatment of neurological disorders.

Background: Human umbilical cord mesenchymal stem cell-derived exosomes (HucMSC-Exo) have shown great therapeutic promise in the treatment of primary ovarian insufficiency (POI). Ferroptosis, a distinct form of cell death, has been associated with the pathogenesis of POI. However, whether HucMSC-Exo can mitigate POI by modulating ferroptosis remains unknown.

Methods: In a CTX-induced POI mouse model, HucMSC-Exo was administered. Ovarian function was assessed by monitoring the estrous cycle, hormone levels, ovarian index, fertility rate, and ovarian morphology. The molecular mechanisms underlying injury and repair were investigated through HucMSC-Exo tracing, immunohistochemical staining, western blot, and real-time polymerase chain reaction (PCR).

Results: HucMSC-Exo restored hormonal balance, preserved ovarian reserve, and reduced follicular atresia and developmental defects in a cyclophosphamide (CTX)-induced POI mouse model. Furthermore, HucMSC-Exo attenuated Fe²⁺ accumulation, oxidative stress, and ferroptosis in the granulosa cells (GCs) of atretic follicles in ovaries with POI. In vitro assays also demonstrated that HucMSC-Exo attenuated CTX-induced ferroptosis in GCs by alleviating Fe²-dependent oxidative damage. Interestingly, hucMSC-Exo specifically suppressed the CTX-induced upregulation of heme oxygenase-1 (HO-1), a key regulator of iron homeostasis, at the translational level. This suggests that post-translational modifications may play a regulatory role in HO-1 expression and iron homeostasis. Mechanistic studies revealed that HucMSC-Exo delivers SMURF1, an E3 ubiquitin ligase that promotes HO-1 degradation, thereby restoring iron homeostasis and inhibiting ferroptosis in GCs. Furthermore, HO-1 knockdown enhanced the protective effects of HucMSC-Exo against CTX-induced ferroptosis and cytotoxicity in GCs.

Conclusions: HucMSC-Exo delivers SMURF1 to promote HO-1 degradation, which in turn suppresses Fe²⁺ accumulation and lipid peroxidation, thereby preventing ferroptosis in GCs and ameliorating chemotherapy-induced POI.