缺氧模拟剂CoCl2重塑猫间充质干细胞外泌体蛋白质组成

Hypoxia-mimetic agent CoCl2 remodels the proteomic profile of exosomes derived from feline mesenchymal stem cells

摘要:

本研究旨在揭示低氧预处理对猫脂肪间充质干细胞(adipose-derived mesenchymal stem cells, ADMSCs)外泌体蛋白质组成的重塑作用及潜在应用。探讨了低氧预处理猫ADMSCs来源外泌体的蛋白质组特征及其生物学功能，通过CoCl?模拟低氧环境处理ADMSCs，采用差速超速离心法分离外泌体，经透射电镜、纳米颗粒追踪分析和蛋白质印迹鉴定其物理特性。蛋白质组学分析显示，低氧预处理诱导外泌体蛋白质组发生显著改变，共鉴定出120个差异表达蛋白质(116个上调、4个下调)。生物信息学分析表明，差异蛋白质显著富集于低氧诱导因子-1 (hypoxia-inducible factor-1, HIF-1)信号通路、腺苷酸活化蛋白激酶(adenosine monophosphate-activated protein kinase, AMPK)信号通路、自噬和蛋白酶体功能等关键通路。基因本体(gene ontology, GO)功能注释显示，低氧预处理组代谢过程、细胞周期调控和信号转导等生物学过程显著富集。京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes, KEGG)分析进一步揭示其可能通过调控细胞周期、肾素-血管紧张素系统等多条通路发挥生物学功能。值得注意的是，差异蛋白质中HMOX1、TFRC等低氧应答蛋白质表达上调，而肾素-血管紧张素系统相关通路受到抑制。本研究首次系统阐明了低氧预处理对猫ADMSCs来源外泌体蛋白质组的重塑作用，为深入理解外泌体介导的细胞间通讯提供了新的理论基础。

Abstract:

This study investigated the proteomic characteristics and biological functions of hypoxic exosomes derived from hypoxia-preconditioned feline adipose-derived mesenchymal stem cells (ADMSCs), aiming to reveal the remodeling effect of hypoxic preconditioning on the protein composition of exosomes derived from feline ADMSCs and its potential applications. CoCl₂ was used to mimic a hypoxic environment for ADMSCs, and exosomes were isolated by differential ultracentrifugation. The physical properties of the exosomes were characterized by transmission electron microscopy, nanoparticle tracking analysis, and Western blotting. Proteomic analysis revealed that hypoxic preconditioning significantly altered the exosomal proteomic profile, identifying 120 differentially expressed proteins (116 upregulated and 4 downregulated). Bioinformatic analysis indicated significant enrichment of key pathways including the hypoxia-inducible factor-1 (HIF-1) signaling pathway, adenosine monophosphate-activated protein kinase (AMPK) signaling pathway, autophagy, and proteasome function. Gene ontology (GO) functional annotation demonstrated significant enrichment of biological processes such as metabolic processes, cell cycle regulation, and signal transduction in the hypoxia-preconditioned group. Kyoto encyclopedia of genes and genomes (KEGG) analysis further suggested potential biological functions through the regulation of pathways including the cell cycle and renin-angiotensin system. Notably, hypoxia-responsive proteins such as HMOX1 and TFRC were upregulated, while pathways related to the renin-angiotensin system were suppressed. This study systematically elucidates, for the first time, the remodeling effect of hypoxic preconditioning on the proteome of exosomes derived from feline ADMSCs, providing new molecular insights into exosome-mediated intercellular communication.