G蛋白偶联受体48基因参与人卵巢颗粒细胞增殖和凋亡的机制研究

Study on the mechanism of G protein-coupled receptor 48 involved in proliferation and apoptosis of human ovarian granulosa cells

目的探讨G蛋白偶联受体48(GPR48)基因通过β-catenin信号通路参与调控颗粒细胞增殖与凋亡的作用机制。方法应用RNA干扰(RNAi)技术建立GPR48基因低表达人卵巢颗粒细胞(KGN)细胞模型,作为GPR48降表达组,取处于增殖期的KGN细胞作为对照组。MTT法检测干扰GPR48表达对KGN细胞增殖的影响。采用流式细胞仪测定干扰GPR48表达对KGN细胞凋亡的影响。采用蛋白质印迹法检测KGN细胞中GPR48蛋白和β-catenin蛋白含量。结果MTT结果显示,GPR48降表达后,KGN细胞增殖活性显著被抑制,GPR48降表达组相对细胞活性为0.41±0.02,显著低于对照组(1.00±0.03),GPR48降表达组细胞凋亡百分比为(13.63±2.57)%,显著高于对照组[(0.23±0.04)%],差异有统计学意义(P<0.05)。GPR48降表达组的β-catenin和GPR48蛋白的表达为2.32±0.57、2.16±0.38、2.58±0.53,均显著低于对照组(0.68±0.17、1.06±0.21、2.51±0.26),差异均有统计学意义(P<0.05)。结论GPR48基因可能通过调控β-catenin信号通路抑制KGN细胞增殖,促进其凋亡。

Objective To investigate the mechanism of G protein-coupled receptor 48(GPR48)gene involved in regulating granulosa cell proliferation and apoptosis throughβ-catenin signaling pathway.Methods Human ovarian granulogranulose cell(KGN)cell model with low GPR48 gene expression was established by RNA interference(RNAi)technology.These cells were used as GPR48 down-expression group,and KGN cells in proliferation stage were used as control group.MTT method was used to detect the effect of downregulation of GPR48 on the proliferation of human ovarian granulocyte(KGN)cells.The effect of downregulation of GPR48 on apoptosis of KGN cells was determined by flow cytometry.The contents of GPR48 protein andβ-catenin protein in KGN cells were detected by Western blotting.Results MTT results showed that the proliferation activity of KGN cells was significantly inhibited after downregulation of GPR48.The relative cell activity of GPR48 down-expression group was 0.41±0.02,which was significantly lower than that of control group(1.00±0.03),and the percentage of apoptosis in GPR48 down-expression group was(13.63±2.57)%,which was significantly higher than that in control group[(0.23±0.04)%],the differences were statistically significant(P<0.05).The expressions ofβ-catenin and GPR48 proteins in the GPR48 down-expression group were 2.32±0.57,2.16±0.38,2.58±0.53,which were significantly lower than those in the control group(0.68±0.17,1.06±0.21,2.51±0.26),and the differences were statistically significant Academic significance(P<0.05).Conclusion GPR48 may inhibit KGN cell proliferation and promote apoptosis by regulating theβ-catenin signaling pathway.