胃癌细胞外泌体调控M2型巨噬细胞极化及对胃癌细胞增殖和迁移的影响

目的:探究胃癌细胞外泌体调控M2型巨噬细胞极化及对胃癌细胞增殖和迁移能力的影响。方法:差速离心法提取胃黏膜上皮细胞GES-1和胃癌细胞株MGC-803、SGC-7901外泌体，透射电镜和Western blot鉴定外泌体，将U937细胞与PMA共孵育诱导U937细胞分化为巨噬细胞的同时分别与PBS、GES-1 exo、MGC-803 exo、SGC-7901 exo、IL-4孵育48 h，流式细胞术检测孵育后各组细胞CD206和HLA-DR表达，qRT-PCR检测CCL17、CXCL8、IL-10、TGF-β、TNF-α、IL-6、IL-1β表达，Western blot检测p-NF-κB、NF-κB、IκBα表达水平，将各组诱导后的U937细胞与MGC-803、SGC-7901共培养后，收集MGC-803、SGC-7901细胞，MTT检测各组细胞的增殖能力，Transwell小室法检测各组细胞的迁移能力。结果:差速离心法提取的外泌体符合外泌体的形态特征，并且在激光共聚焦显微镜下观察到外泌体可被U937细胞摄取，与MGC-803 exo、SGC-7901 exo共孵育后的U937细胞CD206显著高表达，HLA-DR低表达（P＜0.05），CCL17、CXCL8、IL-10、TGF-β表达显著上调（P＜0.05），TNF-α、IL-6、IL-1β表达显著下调（P＜0.05），NF-κB磷酸化水平显著增加（P＜0.05），IκBα表达显著增加（P＜0.05），并且与MGC-803 exo、SGC-7901 exo诱导极化后的巨噬细胞共培养组MGC-803、SGC-7901细胞增殖和迁移能力显著增强（P＜0.05）。结论:胃癌细胞能够通过激活NF-κB信号通路促进M2型巨噬细胞极化进而诱导MGC-803、SGC-7901细胞增殖和迁移能力的增强。

Gastric cancer cell exosomes regulate the polarization of M2 type macrophages and the effect on proliferation and migration of gastric cancer cells

Objective:To investigate the effect of gastric cancer cell exosomes on the proliferation and migration of gastric cancer cells by regulating the polarization of M2 type macrophages.Methods:Differential centrifugation was used to extract gastric mucosal epithelial cells GES-1 and gastric cancer cell lines MGC-803,SGC-7901 exosomes.Transmission electron microscopy and Western blot identified exosomes.Co-incubating U937 cells with PMA induces U937 cells to differentiate into macrophages,while incubating with PBS,GES-1 exo,MGC-803 exo,SGC-7901 exo,and IL-4 for 48 hours.After incubation,the expression of CD206 and HLA-DR in each group was detected by flow cytometry.The expressions of CCL17,CXCL8,IL-10,TGF-β,TNF-α,IL-6,IL-1β were detected by qRT-PCR,and the expression levels of p-NF-κB,NF-κB,and IκBα were detected by Western blot.After the induced U937 cells in each group were co-cultured with MGC-803 and SGC-7901,MGC-803 and SGC-7901 cells were collected.MTT was used to detect the proliferation ability of each group of cells,and the Transwell chamber method was used to detect the migration ability of each group of cells.Results:The exosomes extracted by differential centrifugation met the morphological characteristics of exosomes.Under a laser confocal microscope,the exosomes could be taken up by U937 cells.U937 cells incubated with MGC-803 exo and SGC-7901 exo showed high expression of CD206 and low expression of HLA-DR (P＜0.05).CCL17,CXCL8,IL-10,TGF-β expression was significantly up-regulated (P＜0.05).TNF-α,IL-6,IL-1β expression was significantly down-regulated (P＜0.05).The phosphorylation level of NF-κB was significantly increased (P＜0.05).The expression of IκBα was significantly increased (P＜0.05).And U937 cells were co-operated with MGC-803 exo and SGC-7901 exo induced polarization of macrophages and the proliferation and migration ability of MGC-803 and SGC-7901 in the culture group increased significantly (P＜0.05).Conclusion:Gastric cancer cells can promote the polarization of M2 type macrophages by activating the NF-κB signaling pathway,thereby inducing the increase of the proliferation and migration ability of MGC-803 and SGC-7901 cells.