Abstract：
Human amniotic epithelial cells (hAECs) are novel and promising therapeutic agents for patients suffering from degenerative diseases. Studies have demonstrated that the therapeutic effects of hAECs mainly depend on their paracrine components. Currently, appropriate pretreatment is a widely confirmed strategy for enhancing the repair potential of stem cells; however, the effect of proinflammatory factor pretreatment on hAECs and their secretome is still unclear. In this study, we used the well-characterized proinflammatory factors tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ) to stimulate hAECs and analyzed the effect of TNF-α and IFN-γ on hAECs, including gene expression profile, paracrine proteins, and microRNAs (miRNAs) in exosomes. Results showed that TNF-α and IFN-γ pretreatment improved the viability of hAECs but inhibited the proliferation of hAECs. TNF-α and IFN-γ pretreatment altered the gene expression profile of hAECs, and upregulated differentially expressed genes were predominantly enriched in biological adhesion, antioxidant activity, and response to IFN-beta. In addition, TNF-α and IFN-γ pretreatment enhanced the paracrine secretion of cytokines by hAECs. The upregulated differentially expressed proteins were mainly enriched in tissue remodeling proteins and cytokine-cytokine receptor. Notably, the expression of miRNAs in exosomes from hAECs was also changed by TNF-α and IFN-γ pretreatment. The target genes of upregulated exosomal miRNAs substantially contributed to the response to stimulus, metabolic pathways, and PI3K-Akt signaling pathway. Our findings improve our understanding of the biological characteristics of hAECs after proinflammatory factor pretreatment and provide novel insights to strengthen and optimize the therapeutic potential of hAECs and their secretome in regenerative medicine.
摘要：
人羊膜上皮细胞（hAECs）是一种新型且有前途的治疗退行性疾病的药物。研究表明，hAECs的治疗效果主要取决于其旁分泌成分。目前,适当的预处理是增强干细胞修复潜力的广泛确认的策略;然而,促炎因子预处理对hAECs及其分泌组的影响尚不清楚。在这项研究中,我们使用特征明确的促炎因子肿瘤坏死因子α（TNF-α）和干扰素γ（IFN-γ）刺激hAECs，并分析TNF-α和IFN-γ对hAECs的影响，包括基因表达谱，外泌体中的旁分泌蛋白和miRNA。结果表明,TNF-α和IFN-γ预处理提高了hAECs的活力,但抑制了hAECs的增殖。TNF-α和IFN-γ预处理改变了hAECs的基因表达谱，上调的差异表达基因(DEGs)主要富集在生物粘附中，抗氧化活性和对IFN-β的反应。此外,TNF-α和IFN-γ预处理增强了hAECs旁分泌细胞因子的分泌。上调的差异表达蛋白（DEP）主要富集在组织重塑蛋白和细胞因子-细胞因子受体中。值得注意的是,TNF-α和IFN-γ预处理也改变了hAECs外泌体中miRNA的表达。上调的外泌体miRNA的靶基因对刺激的反应有很大贡献，代谢途径和PI3K-Akt信号通路。我们的发现提高了我们对促炎因子预处理后hAECs生物学特性的理解，并为加强和优化hAECs及其分泌组在再生医学中的治疗潜力提供了新的见解。
