RGC-32通过调节肝癌中Wnt/β-catenin信号诱导Treg/Th17失衡的研究

RGC-32 induces the imbalance of Treg/Th17 by regulating Wnt/β-catenin signaling in patients with liver cancer

目的:研究肝癌患者中RGC-32水平与Treg/Th17细胞比率相关性,初步探究RGC-32对Wnt/β-catenin信号通路的调节作用。方法:采用酶联免疫吸附试验(ELISA)检测健康对照和肝癌患者血浆中RGC-32、IL-17、IL-22、TGF-β1、IL-10表达水平;采用流式细胞术检测健康对照和肝癌患者外周血中Th17细胞和Treg细胞数;采用siRNA转染敲低单个核细胞(PMBC)中RGC-32水平,通过蛋白质免疫印迹(Western blot)检测RGC-32敲低后对Wnt/β-catenin信号通路的影响。结果:与健康对照比较,肝癌患者血浆中RGC-32水平显著升高(P<0.01),Th17细胞数升高而Treg细胞数下降,Treg/Th17细胞比率显著下降(P<0.01)。与健康对照比较,肝癌患者血浆中Th17细胞分泌的促炎细胞因子IL-17和IL-22显著升高(P<0.01),而Treg细胞分泌的抗炎细胞因子TGF-β1和IL-10显著下降(P<0.01)。线性回归分析显示,在健康对照中RGC-32水平与Treg/Th17无相关性(P>0.05),在肝癌患者中RGC-32水平与Treg/Th17呈负相关(P<0.01)。敲低RGC-32后,检测到促炎细胞因子IL-17表达下降(P<0.01),抗炎因子TGF-β1表达升高(P<0.01),Wnt/β-catenin信号的激活被抑制。结论:RGC-32的上调导致肝癌患者Treg/Th17细胞的失衡,RGC-32下调能抑制Wnt/β-catenin信号的激活。

Objective:To investigate the correlation between the level of RGC-32 and the ratio of Treg/Th17 cells in patients with liver cancer,and to explore the regulatory effect of RGC-32 on Wnt/β-catenin signaling pathway.Methods:The expression levels of RGC-32,IL-17,IL-22,TGF-β1 and IL-10 in plasma of healthy controls and liver cancer patients were detected by ELISA.Flow cytometry was used to detect Th17 cells and Treg cells in peripheral blood of healthy controls and patients with liver cancer.The level of RGC-32 in mononuclear cells(PMBC)was reduced by siRNA transfection,and the effect of RGC-32 knockdown on Wnt/β-catenin signaling pathway was detected by Western blot.Results:Compared with healthy controls,the level of RGC-32 in liver cancer patients was significantly increased(P<0.01),the number of Th17 cells was increased while the number of Treg cells was decreased,and the ratio of Treg/Th17 cells was significantly decreased(P<0.01).Compared with healthy controls,the proinflammatory cytokines IL-17 and IL-22 secreted by Th17 cells were significantly increased in liver cancer patients(P<0.01),while the anti-inflammatory cytokines TGF-β1 and IL-10 secreted by Treg cells were significantly decreased(P<0.01).By linear regression analysis,the results showed that there was no correlation between the level of RGC-32 and Treg/Th17 in healthy controls(P>0.05),but a negative correlation between the level of RGC-32 and Treg/Th17 in patients with liver cancer(P<0.01).After knockdown of RGC-32,the expression of pro-inflammatory cytokine IL-17 was decreased(P<0.01),the expression of anti-inflammatory cytokine TGF-β1 was increased(P<0.01),and the activation of Wnt/β-catenin signal was inhibited.Conclusion:The up-regulation of RGC-32 leads to the imbalance of Treg/Th17 cells in patients with liver cancer,and the down-regulation of RGC-32 can inhibit the activation of Wnt/β-catenin signal.