软骨终板干细胞外泌体活化巨噬细胞自噬抑制软骨终板炎的研究

Cartilage endplate stem cell-derived exosomes delay the progression of cartilage endplate inflammation by activating autophagy in macrophages

目的探讨软骨终板干细胞外泌体通过调控巨噬细胞进而抑制软骨终板炎的机制。方法采用免疫组化检测炎症因子和巨噬细胞在不同分级分期退变的软骨终板组织中的表达和浸润,NTA(nanoparticle tracking analysis)和透射电镜鉴定外泌体,外泌体质谱检测和KEGG富集分析预测相关信号通路,Western blotting和免疫荧光检测巨噬细胞中自噬因子LC3A/B和炎症因子IL-1β、IL-6及TNF-α的蛋白水平。结果在临床标本中,软骨终板的退变程度与巨噬细胞浸润数量IL-1β水平均正相关;40μg/ml软骨终板干细胞外泌体可有效活化巨噬细胞自噬,使P62表达降低,LC3B表达增加;自噬激动剂Rapamycin(20μmol/ml)则降低了TBHP诱导巨噬细胞炎症因子IL-6、IL-1β的水平。结论软骨终板干细胞外泌体通过活化巨噬细胞自噬,降低其炎症因子表达,延缓了软骨终板炎的进程。

This study was designed to study the mechanism of cartilage endplate stem cell-derived exosomes in regulating macrophages to inhibit the progression of cartilage endplate inflammation.Immunohistochemistry was used to detect the expression and infiltration of inflammatory factors and macrophages in different degenerated cartilage endplate;exosome were identified by nanoparticle tracking analysis(NTA)and transmission electron microscopy;mass spectrometry detection of exosomes and KEGG enrichment analysis were used to speculate related signaling pathways;Western blot and immunofluorescence were used to detect the difference in the levels of autophagy protein LC3A/B and inflammatory factors IL-1β,IL-6 and TNF-αin macrophages under different condition.Data showed that the degree of cartilage endplate degeneration was positively correlated with the number of macrophage infiltration and IL-1βlevels in clinical specimens.Exosomes of 40μg/ml could effectively activate macrophage autophagy,reduce the expression of P62,and increase the expression of LC3B.The autophagy agonist Rapamycin(20μmol/ml)decreased the levels of TBHP-induced macrophage inflammatory factors IL-6 and IL-1β.In conclusion,exosomes derived from cartilage endplatestem cells can reduce the expression of inflammatoryfactors by activating macrophage autophagy,thusdelaying the process of cartilage endplate inflammation.