Stem Cells International最新文献

Background: Atopic dermatitis (AD) is a chronic inflammatory skin disease that significantly impacts quality of life. Reducing inflammation and restoring the skin barrier are key to its management.

Objective: This study aimed to investigate the protective and therapeutic effects of secretory substances from induced pluripotent stem cell (iPSC)-derived mesenchymal stem cells (iMSCs) in AD.

Methods: The protective effects of iMSC secretome pretreatment were evaluated in HaCaT cells by assessing cell viability, AD biomarker expression, and cell migration. Therapeutic efficacy was examined in a 1-chloro-2,4-dinitrobenzene (DNCB)-induced AD mouse model through clinical, histological, and immunological assessments. Proteomic analyses were performed to relevant biological processes.

Results: iMSC secretome significantly reduced AD-induced cell death and AD biomarker expressions in vitro (p  < 0.05), with 200 μg/mL iMSC secretome promoting cell migration. In vivo, high dose (20 mg/mL) iMSC secretome alleviate clinical indicators compared to the vehicle group (p  < 0.05). Serum immunoglobulin (Ig) E, interleukin (IL)-4, IL-31, and IL-6 levels, along with the expression of AD biomarkers in skin, were significantly decreased (p  < 0.05). Proteomic analyses revealed upregulation of genes involved in the regulation of immune responses and the restoration of skin barrier.

Conclusion: iMSC secretome demonstrates significant anti-inflammatory and regenerative effects, making it a promising therapeutic option for AD.

Background: Protein kinase R-like endoplasmic reticulum kinase (PERK) mediates the endoplasmic reticulum stress (ERS) response. However, it remains unclear whether PERK regulates the odontogenic differentiation of human dental pulp stem cells (hDPSCs) and contributes to the repair and regeneration of the pulpo-dentinal complex (PDC) during inflammation. This study aimed to investigate the regulation of ERS-PERK in the differentiation and apoptosis of hDPSCs and its contribution to the repair and regeneration of PDC injury in the inflammatory microenvironment.

Methods: In vivo dentin defect (DD group) and pulp perforation (PP group) were established to evaluate the expression and healing-promoting properties of PERK in dental pulp at different injury stages. Using LPS with concentration gradients to simulate the inflammatory microenvironment, the activation of ERS-PERK pathway-related genes was investigated using quantitative real-time polymerase chain reaction (RT-qPCR). hDPSCs' apoptosis and odontogenic potential under inflammatory stimulation were also assessed using Calcein-AM/7AAD Live/Dead cell double staining, RT-qPCR, alkaline phosphatase (ALP) staining, and Alizarin Red S (ARS) staining. The potential role of PERK in odontogenesis and apoptosis of hDPSCs under inflammatory stimulation was explored.

Results: With the extension of dental defects in vivo, PERK expression was gradually upregulated in the DD group, whereas, in the PP group, it increased in the early stage of inflammation and then decreased. Under LPS stimulation, the expression of inflammatory factors increased with the activation of the PERK pathway, while the ALP activity of hDPSCs and mineralized nodules decreased. PERK knockdown attenuated mRNA levels of ERS-related genes and apoptosis-related genes, whereas the expression of odontogenic-related factors increased. The ALP activity and the number of mineralized nodules increased. PERK may regulate odontogenesis through the mitochondria-associated endoplasmic reticulum (ER) membrane.

Conclusion: Our results demonstrated PERK pathway activation, which enhances the expression of inflammatory factors and suppresses the odontogenic ability of hDPSCs in inflammatory microenvironments.

Background: Hyperthermia is a widely used adjunct treatment for different cancers. The GLI1 is upregulated in ESCC and its expression is associated with the stemness of ESCC.

Objective: We hypothesized that GLI1 constitutes an important hyperthermia treatment target, and investigated its contribution to hyperthermia responses in ESCC.

Methods: The growth of the human ESCC cell lines KYSE70 and KYSE140 was analyzed using CCK-8, clonogenicity and spheres formation assays after 43°C hyperthermia, under conditions of knockdown or overexpression of GLI1. Stemness-related proteins were determined using Western blotting and immunofluorescence staining. Last, the molecular mechanism of GLI1 degradation was studied using chemical inhibitors and immunoprecipitation assays.

Results: Hyperthermia increased the ubiquitination and proteasomal destruction of GLI1, causing a rapid decline in GLI1 protein levels of ESCC cells. Similar to GLI1 knockdown, ESCC cells treated with hyperthermia showed growth inhibition associated with the downregulation of cancer stemness proteins.

Conclusion: Our study reveals that hyperthermia can readily destabilize GLI1 levels in ESCC cells and inhibit ESCC cells growth. This proposes new strategies for implementing hyperthermia to target GLI1 driven cancers to improve therapeutic efficacy.

Objective: This study aimed to investigate the effect of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway inhibition on D-galactose (D-gal)-induced senescence in nucleus pulposus cells (NPCs) and its potential to delay intervertebral disc degeneration (IVDD), as well as to investigate the underlying mechanisms.

Methods: A cellular senescence model was established by treating rat NPCs with D-gal. The model was then intervened with a JAK2/STAT3 pathway inhibitor (ruxolitinib) or JAK2-specific small interfering RNA (siRNA). Cellular senescence was evaluated by senescence-associated β-galactosidase (SA-β-gal) staining. The expression of senescence markers (p16, p21, and p53), extracellular matrix (ECM) components (aggrecan and collagen II), catabolic enzymes (ADAMTS-4, ADAMTS-5, MMP-3, and MMP-13), and JAK2/STAT3 pathway proteins was analyzed by western blotting and immunofluorescence. The levels of inflammatory factors (interleukin [IL]-1β, IL-6, and tumor necrosis factor-α [TNF-α]) and advanced glycation end-products (AGEs) were measured by enzyme-linked immunosorbent assay (ELISA). Cell proliferation, apoptosis, and cell cycle distribution were assessed using cell counting kit-8 (CCK-8) and flow cytometry. In a parallel in vivo study, a rat model of IVDD was induced by D-gal and treated with the JAK inhibitor. Disc degeneration was evaluated by magnetic resonance imaging (MRI) and histopathological examination after 8 weeks.

Results: Both in vitro and in vivo, inhibition of the JAK2/STAT3 pathway, either pharmacologically or genetically, effectively attenuated D-gal-induced effects. It suppressed the phosphorylation of STAT3, reduced the expression of SA proteins (p16, p21, and p53), ECM catabolic enzymes (ADAMTS-4, ADAMTS-5, MMP-3, and MMP-13), and proinflammatory cytokines (IL-1β and IL-6). Consequently, this inhibition decreased SA-β-gal positivity, alleviated cell cycle arrest and apoptosis, and enhanced the synthesis of aggrecan and collagen II in NPCs. In the rat model, JAK inhibitor treatment improved MRI scores, restored disc signal intensity, and ameliorated histopathological degeneration.

Conclusion: : Inhibition of the JAK2/STAT3 pathway reduced the expression of inflammatory factors and oxidative stress markers in D-gal-treated NPCs. It also suppressed ECM degradation and apoptosis, delayed cellular senescence, and attenuated the progression of IVDD in rats.

Background: Adipose-derived stem cells (ADSCs) have important application prospects in the field of regenerative medicine, such as adjuvant autologous fat transplantation (AFT), due to their multidirectional differentiation and immunomodulatory functions. However, functional limitation of ADSCs in hypoxic environments may affect their effectiveness in clinical applications. Hypoxic preconditioning is a potential strategy to improve the function of ADSCs by enhancing the antioxidant capacity and metabolic adaptations of the cells, but the optimal hypoxic conditions and the mechanism of action have not yet been clarified.

Methods: ADSCs were extracted and pretreated with hypoxia in order to explore its effect on the function of ADSCs. The activity, apoptosis level, proliferation ability, and antioxidant capacity of ADSCs under normoxic and hypoxic conditions were evaluated using flow cytometry (FCM), live-dead cell fluorescence assay, and apoptosis flow assay. Further, the role of METTL3-mediated m6A modification in hypoxic preconditioning was explored by real-time fluorescence quantitative PCR, protein immunoblotting assay, and m6A modification level detection. Finally, the effect of hypoxic preconditioning of ADSCs on fat graft retention was verified by the mouse AFT model.

Results: We found that the survival of ADSCs was not affected by 5% O₂ pretreatment. Moreover, the cell viability and proliferation of ADSCs were enhanced after 24 h of anoxic preconditioning. Therefore, we determined that 5% O₂ treatment for 24 h was the best hypoxic pretreatment condition for ADSCs, which enhanced the antioxidant capacity of ADSCs, reduced apoptosis. METTL3-mediated m6A modification played a critical role in hypoxic preconditioning to reduce apoptosis in ADSCs. It was verified in a mouse model that hypoxia preconditioning of ADSCs significantly improved adipose graft retention and promoted neovascularization.

Conclusions: METTL3-mediated modification of m6A enhances the function of ADSCs under hypoxic conditions to improve adipose graft retention. These findings provide a new strategy and theoretical basis for improving the clinical outcome of fat grafting, as well as new molecular targets for future research.

Background: The therapeutic potential of human umbilical cord mesenchymal stem cells (HUCMSCs) for diabetic peripheral neuropathy (DPN) and the underlying mechanisms involving transient receptor potential vanilloid 1 (TRPV1) signaling remain incompletely defined.

Objective: This study aimed to elucidate the role of the TRPV1-[Ca²⁺]i-AMPK signaling axis in mediating the beneficial effects of HUCMSCs on neuropathic pain and Schwann cell (SC) dysfunction in DPN.

Methods: A murine model of DPN was established. Mechanical allodynia and thermal hyperalgesia were assessed using Von Frey filaments and the KW-LB hot plate test, respectively. Primary mouse SCs were isolated and cultured under high glucose (HG) conditions. Intracellular calcium ([Ca²⁺]i) levels were quantified by flow cytometry. Protein expression (TRPV1, p-TRPV1, AMPK, p-AMPK, cleaved-caspase-3, Bax, Bcl-2, Drp1, PGC-1α, TFAM, Mfn2) was analyzed via Western blotting. Apoptosis and cell proliferation were evaluated using TUNEL staining and the CCK-8 assay, respectively. Specific inhibitors (AMG9810 for TRPV1 and compound C for AMPK) were employed to probe pathway involvement.

Results: HUCMSC administration significantly alleviated mechanical allodynia and thermal hyperalgesia in diabetic mice. In vitro, HUCMSC coculture counteracted HG-induced effects in SCs by: (1) increasing the p-TRPV1/TRPV1 ratio and [Ca²⁺]i influx (effects blocked by AMG9810); (2) reducing apoptosis (decreased cleaved-caspase-3/Bax, increased Bcl-2); (3) enhancing the p-AMPK/AMPK ratio (attenuated by both AMG9810 and compound C); and (4) promoting mitochondrial homeostasis, increasing PGC-1α, TFAM, and Mfn2 expression, mitochondrial membrane potential and ATP levels, and decreasing Drp1 expression. These mitochondrial improvements were reversed by compound C.

Conclusion: HUCMSCs ameliorate diabetic neuropathic pain primarily through activation of the TRPV1-[Ca²⁺]i-AMPK signaling pathway in SCs, which may provide a new molecular target for enhancing the clinical therapeutic effect of HUCMSCs on DPN.

Periodontitis is a persistent inflammatory ailment that impacts periodontal tissues. Periodontal ligament stem cells (PDLSCs), also referred to as stem cells, possess advantageous attributes for tissue engineering and regenerative medicine due to their ability to self-renew with multi-directional differentiation potential. Nevertheless, the process of cellular senescence can compromise the restoration and regeneration of tissues, thereby impairing the normal regenerative and reparative functions of the periodontium. Exendin-4 (Ex-4) has protective effects against cellular senescence and apoptosis, but the impact of Ex-4 on inflammation-induced senescence of PDLSCs is unknown. This study used lipopolysaccharide (LPS) to simulate an inflammatory microenvironment, and then assessed the effect of Ex-4 on PDLSC senescence within that environment. Initially, PDLSCs were isolated and characterized and then cultured with LPS or LPS and Ex-4. Results demonstrated that the LPS-induced inflammatory microenvironment produced premature senescence of PDLSCs, which was reversible by treatment with Ex-4. Potential mechanisms underlying the effect were evaluated with regard to senescence-associated molecular pathways, and results demonstrated senescence of PDLSCs to be associated with Sirtuin 1 down-regulation and Notch1 upregulation. Our findings suggest that Ex-4 may mitigate the inflammation-induced senescence of PDLSCs through the SIRT1/Notch1 signaling pathway.

Epithelial cells are integral to tumor composition and engage with various immune cell types within the tumor microenvironment, influencing tumor progression and metastasis. A thorough exploration of the roles and mechanisms of these epithelial cells could enhance early detection strategies and treatment modalities for lung adenocarcinoma (LUAD). This research employed single-cell analysis techniques, complemented by machine learning algorithms, to identify genes associated with epithelial cells and evaluate their prognostic significance and implications for immunotherapy in LUAD patients. By leveraging multiple datasets and applying diverse clustering methods within machine learning, we successfully crafted and validated a diagnostic model specifically for LUAD. Among the genes linked to epithelial cells, the XGBoost and random forest techniques identified PMAIP1 as the most crucial gene in terms of prognosis. Additionally, this study investigated the relationship between PMAIP1 and the infiltration of immune cells. The expression levels of PMAIP1 and its relevance in LUAD were subsequently confirmed through immunohistochemical staining and in vitro cell experiments. This analysis revealed 17 key genes associated with epithelial cells by integrating single-cell analysis with clinical data from the TCGA-LUAD dataset, underscoring their significance in diagnosis, prognostic assessment, and possible treatment avenues for LUAD patients. Importantly, PMAIP1 is strongly linked to prognosis and responses to immunotherapy in LUAD, with experimental findings indicating its heightened expression in PRAD and its connection to adverse outcomes. Furthermore, reducing PMAIP1 expression has been shown to hinder the proliferation, metastasis, and stemness of LUAD cells. In summary, our findings indicate that PMAIP1 has potential as a prognostic biomarker and a target for immunotherapy in patients with LUAD.

Introduction: Umbilical cord-derived mesenchymal stem cells (UCMSCs) are promising candidates for the treatment of myocardial infarction (MI). However, their low mobility and survival limit their clinical applicability. This study aimed to enhance the therapeutic potential of UCMSCs by preincubating them with escin, a natural medicine derived from the dried mature seeds of Aesculus wilsonii.

Methods: We characterized the functional properties of UCMSCs before and after escin preconditioning in vitro. Additionally, we performed RNA sequencing (RNA-seq) to analyze the transcriptomic differences between untreated and escin-pretreated UCMSCs (E-UCMSCs), followed by Western blot (WB) validation of the differentially expressed genes. In vivo, an MI model was established in rats, which involved permanent ligation of the left anterior descending coronary artery, followed by intravenous administration of UCMSCs and E-UCMSCs through the tail vein. The therapeutic efficacy of UCMSCs and E-UCMSCs was assessed by cardiac function measurements and Masson's trichrome staining to quantify fibrosis.

Results: No significant differences were observed in the basic characteristics of the UCMSCs before and after escin pretreatment. RNA-seq results demonstrated higher expression of intercellular adhesion molecule 1 (ICAM1) and GATA-binding protein 4 (GATA4) in E-UCMSCs than in UCMSCs. Furthermore, WB results confirmed this phenomenon. Most importantly, E-UCMSCs significantly restored myocardial contractile function and reduced infarct size in MI rats.

Conclusions: The current study demonstrates that escin upregulated ICAM1 and GATA4 gene expression in UCMSCs, thereby enhancing the therapeutic efficacy of UCMSCs in rats with MI. Therefore, pretreatment of UCMSCs with escin is a promising approach for the treatment of MI.