NLRP3炎性小体通过调控巨噬细胞分化促进子宫内膜癌细胞迁移、侵袭及上皮-间质转化

NLRP3 inflammasome promotes endometrial cancer cell migration,invasion and epithelial-mesenchymal transition by regulating macrophage differentiation

目的探究NLRP3炎性小体通过调控巨噬细胞分化对子宫内膜癌迁移、侵袭及上皮-间质转化的影响。方法 RTPCR检测CD68、TGF-β、IL-10和VEGF m RNA表达;Western blot检测NLRP3、ACS、caspase-1、E-cadherin、N-cadherin、Vimentin蛋白表达;流式细胞术检测F4/80+/CD68+M1巨噬细胞和F4/80+/CD163+M2巨噬细胞数量;Transwell实验检测人子宫内膜癌细胞迁移和侵袭能力。结果与人子宫内膜上皮细胞h EEC相比,人子宫内膜癌细胞RL95-2中NLRP3、ACS和caspase-1表达均显著升高(P<0.05)。干扰NLRP3、ACS和caspase-1表达可显著抑制RL95-2细胞迁移、侵袭及上皮-间质转化(P<0.01)。与siNC组相比,干扰NLRP3的hEEC和RL95-2细胞显著抑制巨噬细胞分化为M2表型(P<0.05),显著降低M2型细胞因子TGF-β、IL-10和VEGF的表达(P<0.01),干扰ACS或caspase-1的hEEC和RL95-2细胞对巨噬细胞分化为M1、M2表型无显著影响(P>0.05),干扰NLRP3的hEEC和RL95-2细胞对巨噬细胞分化为M1表型无显著影响(P>0.05)。与NC组相比,过表达NLRP3的hEEC和RL95-2细胞可显著促进巨噬细胞向M2表型分化(P<0.05),显著升高TGF-β、IL-10和VEGF的表达(P<0.01)。过表达NLRP3诱导产生的M2巨噬细胞可显著促进RL95-2细胞迁移、侵袭及上皮-间质转化(P<0.01)。结论 NLRP3炎性小体通过诱导巨噬细胞M2分化促进子宫内膜癌的迁移、侵袭及上皮-间质转化。

The purpose of this study was to investigate the effect of NLRP3 inflammasome on the migration,invasion and epithelial-mesenchymal transition of endometrial cancer cells by regulating macrophage differentiation.Human endometrial cancer cells RL95-2 were transfected with si-NLRP3, si-ACS, si-caspase-1 or si-NC, and then co-cultured with macrophages.RT-PCR was used to detect the mRNA expression levels of CD68,TGF-β, IL-10 and VEGF mRNA;Western blot was applied to detect the protein expressions of NLRP3, ACS,caspase-1, E-cadherin, N-cadherin and Vimentin protein;flow cytometry was used to detect F4/80+/CD68+M1 macrophages and F4/80+/CD163+number of M2 macrophages;Transwell test was utilized to detect the migration and invasion ability of human endometrial cancer cells.Data showed that the expression levels of NLRP3, ACS and caspase-1 in human endometrial cancer cells RL95-2 was significantly increased compared with human endometrial epithelial cells hEEC(P<0.05);hEEC and RL95-2 cells interfered with si-NLRP3 could significantly inhibit macrophage differentiation into M2 phenotype(P<0.05) and significantly reduce the expression of M2 cytokines TGF-β, IL-10, and VEGF(P<0.01);hEEC and RL95-2 cells interfered with si-ACS or si-caspase-1 had no significant effect on M1 and M2 differentiation of phagocytes(P>0.05);hEEC and RL95-2 cells interfered with NLRP3 had no significant effect on the differentiation of macrophages into M1 phenotype(P>0.05).On the other way, hEEC and RL95-2 cells overexpressing NLRP3 could significantly promote the differentiation of macrophages into M2 phenotype(P<0.05), and significantly increase the expression of TGF-β, IL-10 and VEGF(P<0.01);M2 macrophages induced by NLRP3 overexpression could significantly promote the migration, invasion and epithelial-mesenchymal transition of RL95-2 cells(P<0.01).In summary, NLRP3 inflammatory bodies promote the migration, invasion and epithelial-mesenchymal transition of endometrial cancer cells by inducing macrophage M2 differentiation.The results of this study provide new scientific data for elucidating the pathogenesis of endometrial cancer.