缺氧诱导的结直肠癌外泌体通过激活PTEN/PI3Kγ信号通路介导巨噬细胞极化  

Hypoxia-induced colorectal cancer exosomes mediate macrophage polarization by activating the PTEN/PI3Kγsignaling pathway

作　　者：王宏力[1] 陈晓艳[2] 张鹏[1] 郝建军[1] 李健[1] 田建虎 王志远 WANG Hongli;CHEN Xiaoyan;ZHANG Peng;HAO Jianjun;LI Jian;TIAN Jianhu;WANG Zhiyuan(Department of Colorectal,Baoding First Hospital,Hebei Baoding 071000,China;Department of Prevention and Protection,Baoding First Hospital,Hebei Baoding 071000,China)

机构地区：[1]保定市第一医院结直肠外科,河北保定071000 [2]保定市第一医院防保科,河北保定071000

出　　处：《现代肿瘤医学》2021年第19期3363-3368,共6页Journal of Modern Oncology

基　　金：河北省医学科学研究重点课题计划项目(编号:20180023)。

摘　　要：目的:探究缺氧条件下,结直肠癌细胞外泌体对巨噬细胞极化的影响及机制。方法:差速离心法提取正常条件下和缺氧条件下结直肠癌细胞HT-29所分泌的外泌体(HT-29 N-exo和HT-29 H-exo),透射电镜和Western blot鉴定外泌体,荧光显微镜下观察外泌体的摄取。将U937细胞与100 ng/mL的PMA共孵育的同时分别与1μg/mL的HT-29 N-exo和HT-29 H-exo以及等体积的PBS共孵育,记为PBS组、HT-29 N-exo组、HT-29 H-exo组。qRT-PCR检测各组细胞CD163、IL-10、IL-1Rα、TGF-β1、CCL2、TNF-αmRNA的表达,Elisa检测各组细胞IL-10和TNF-α分泌,流式细胞术鉴定各组细胞CD163和CD11b表达,Western blot检测各组细胞PTEN/PI3Kγ信号通路激活情况。结果:差速离心法所提取的外泌体符合其结构和生物学特征,并且可被U937细胞摄取;相比于HT-29 N-exo组,HT-29 H-exo组细胞CD163、IL-10、IL-1Rα、TGF-β1、CCL2表达显著增加(P<0.05),IL-10分泌显著增加,TNF-α表达以及分泌显著减少(P<0.01),CD163+CD11b+巨噬细胞数显著增加(P<0.001),PTEN/PI3Kγ信号通路显著激活(P<0.001)。结论:缺氧条件下结直肠癌细胞外泌体能够显著促进巨噬细胞M2型极化,其机制为激活PTEN/PI3Kγ信号通路。Objective:To explore the effect and mechanism of colorectal cancer exosomes on the polarization of macrophages under hypoxic conditions.Methods:Differential centrifugation was used to extract exosomes(HT-29 N-exo and HT-29 H-exo)secreted by colorectal cancer cells HT-29 under normal conditions and under hypoxic conditions,and were identified by transmission electron microscopy and Western blot.The uptake of exosomes was observed under a fluorescence microscope.The U937 cells were incubated with 100 ng/mL PMA and simultaneously incubated with 1μg/mL HT-29 N-exo and HT-29 H-exo and an equal volume of PBS,which were recorded as PBS group,HT-29 N-exo group and HT-29 H-exo group.qRT-PCR detected the expression of CD163,IL-10,IL-1Rα,TGF-β1,CCL2,TNF-αmRNA in each group of cells.Elisa detected the secretion of IL-10 and TNF-αin each group of cells,and flow cytometry identified each group The expression of CD163 and CD11b in cells.Western blot was used to detect the activation of PTEN/PI3Kγsignaling pathway in each group.Results:The exosomes extracted by differential centrifugation conformed to their structure and biological characteristics,and could be taken up by U937 cells.Compared with HT-29 N-exo group,HT-29 H-exo group CD163,IL-10,IL-1Rα,TGF-β1,CCL2 expression increased significantly(P<0.05),IL-10 secretion increased significantly,TNF-αexpression and secretion decreased significantly(P<0.01),and the number of CD163+CD11b+macrophages increased significantly(P<0.001).The PTEN/PI3Kγsignaling pathway was significantly activated(P<0.001).Conclusion:Colorectal cancer exosomes can significantly promote the M2 polarization of macrophages under hypoxic conditions,and the mechanism is to activate the PTEN/PI3Kγsignaling pathway.

关 键 词：缺氧 结直肠癌 外泌体 巨噬细胞极化 

分 类 号：R735.37[医药卫生—肿瘤]