Background Epidermal stem cells (ESCs) are primarily located in the basal layer of the epidermis and play a crucial role in wound healing. ESCs-derived exosomes (ESCs-Exo) are emerging as promising candidates for skin regeneration and wound healing. However, the underlying mechanisms remain unclear. This study aims to investigate the role and mechanisms of ESCs-Exo in promoting the proliferation, migration, and collagen synthesis of human skin fibroblasts (HSFBs). Methods This study generated, isolated, and characterized ESC-Exos. The effects of ESCs-Exo on the proliferation of human skin fibroblasts (HSFBs) were detected via Cell Counting Kit-8 (CCK8), 5-Ethynyl-2'-deoxyuridine (EdU), and Proliferating Cell Nuclear Antigen (PCNA) and Marker of Proliferation Ki-67 (MKI67) gene expression methods. The effect of ESCs-Exo on the migration of HSFBs was detected via a transwell assay and a scratch test. The concentrations of collagen secreted by the HSFBs and the mRNAs of the two kinds of collagen expressed by the HSFBs were analyzed. We also analyzed the phosphorylation of Protein Kinase N1 (PKN1) and the expression of cyclins via western blotting. Finally, the effect of ESCs-Exo on wound healing was verified by animal experiments, and the key genes and signaling pathways of ESCs-Exo were excavated by transcriptomic analysis. Results Western blotting revealed that the exosomes of ESCs highly expressed established markers such as Alix, CD63, and CD9. ESC-Exos significantly promoted HSFB proliferation and migration in a dose-dependent manner, as well as HSFB collagen synthesis, and effectively increased the ratio of collagen III/I. In addition, bioinformatics analysis showed that the expression of key gene C-X-C motif chemokine ligand 9 was lower in the ESCs-Exo group, which may promote wound healing by regulating PKN1-cyclin and tumor necrosis factor signaling pathways. Animal experiments demonstrated that ESCs-Exo could reduce inflammation and accelerate wound healing. Conclusions In this study, we found that ESCs-Exo may improve wound healing by promoting the proliferation and migration of HSFBs.

中文翻译：

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表皮干细胞来源的外泌体通过促进人体皮肤成纤维细胞的增殖和迁移来改善伤口愈合


背景 表皮干细胞 （ESC） 主要位于表皮的基底层，在伤口愈合中起着至关重要的作用。ESCs衍生的外泌体（ESCs-Exo）正在成为皮肤再生和伤口愈合的有前途的候选者。然而，潜在机制仍不清楚。本研究旨在探讨 ESCs-Exo 在促进人皮肤成纤维细胞 （HSFBs） 增殖、迁移和胶原蛋白合成中的作用和机制。方法 本研究生成、分离和表征 ESC-Exos。通过细胞计数试剂盒-8 （CCK8）、5-乙炔基-2'-脱氧尿嘧啶 （EdU） 和增殖核抗原标志物 Ki-67 （MKI67） 基因表达方法检测 ESCs-Exo 对人皮肤成纤维细胞 （HSFBs） 增殖的影响。通过 transwell 测定和划痕试验检测 ESCs-Exo 对 HSFBs 迁移的影响。分析 HSFBs 分泌的胶原蛋白浓度和 HSFBs 表达的两种胶原蛋白的 mRNA。我们还通过 western blotting 分析了蛋白激酶 N1 （PKN1） 的磷酸化和细胞周期蛋白的表达。最后，通过动物实验验证 ESCs-Exo 对伤口愈合的影响，并通过转录组学分析挖掘 ESCs-Exo 的关键基因和信号通路。结果 Western blotting 显示 ESCs 的外泌体高度表达已建立的标志物，如 Alix 、 CD63 和 CD9。ESC-Exos 以剂量依赖性方式显著促进 HSFB 增殖和迁移，以及 HSFB 胶原蛋白合成，并有效增加胶原蛋白 III/I 的比例。 此外，生物信息学分析显示，ESCs-Exo 组关键基因 C-X-C 基序趋化因子配体 9 的表达较低，可能通过调节 PKN1-cyclin 和肿瘤坏死因子信号通路促进伤口愈合。动物实验表明，ESCs-Exo 可以减少炎症并加速伤口愈合。结论 在本研究中，我们发现 ESCs-Exo 可能通过促进 HSFBs 的增殖和迁移来改善伤口愈合。