TGF-β和IGF-1信号通路调控肾母细胞瘤细胞增殖机制探讨  

TGF-β and IGF-1 signaling pathways regulate the prolifetation of Wilms' tumor cells

作　　者：刘卓[1] 何峰[2] 李媛媛[1] 欧阳胜荣[1] 曹丁丁 常会波[1] 马斐斐[1] 吴建新[1] 

机构地区：[1]首都儿科研究所生物化学研究室,北京100020 [2]首都儿科研究所分子免疫研究室,北京100020

出　　处：《中华肿瘤防治杂志》2017年第12期813-816,共4页Chinese Journal of Cancer Prevention and Treatment

基　　金：北京市自然科学基金(7042017)

摘　　要：目的肾母细胞瘤是一种高发的儿童实体瘤,发病率占儿童恶性肿瘤8%~10%,多发生于<5岁儿童,有研究表明某些信号通路可调控其增殖过程。本研究探讨TGF-β和IGF-1信号通路是否共同参与调控肾母细胞瘤细胞的增殖过程。方法采用siRNA转染实验将TGFBR1-siRNA和IGF1R-siRNA转至肾母细胞瘤细胞株G401,以转染不同siRNA分为TGFBR1-siRNA组和IGF1R-siRNA组,siRNA无关序列(non-siRNA)转染组作为对照组,每组设6个复孔。MTS方法检测TGFBR1-siRNA和IGF1R-siRNA对细胞增殖率的影响,蛋白质印迹法检测TGF-β信号通路TGFBR1及下游蛋白p-ERK、SMAD2/3和p-AKT蛋白表达情况;蛋白质印迹法检测IGF-1信号通路IGF1R及下游蛋白p-AKT蛋白表达情况。ELISA法检测细胞上清中FGF9蛋白表达。结果抑制TGFBR1和IGF1R72和96h后,细胞的增殖率受到抑制,对照组增殖率设为100%。与其相比,转染TGFBR1-siRNA 72h后的实验组和对照组的增殖率分别为(80±14)%和(100±7)%,实验组低于对照组,P=0.034;转染IGF1R-siRNA 72h后实验组和对照组的增值率分别为(85±21)%和(100±7)%,实验组低于对照组,P=0.028。转染TGFBR1-siRNA 96h后的实验组和对照组的增殖率分别为(81±13)%和(100±11)%,实验组低于对照组,P=0.021;转染IGF1R-siRNA 96h后实验组和对照组的增殖率分别为(82±17)%和(100±11)%,实验组低于对照组,P=0.038。抑制TGFBR1后,与non-siRNA组相比,下游蛋白FGF-9、p-ERK、SMAD2/3和p-AKT明显受到抑制。TGFBR1-siRNA转染72h后,实验组细胞上清中FGF9蛋白表达量显著下降,实验组为(533.85±54.64)ρg/mL,对照组为(587.34±46.83)ρg/mL,P=0.041;96h后实验组FGF9蛋白表达量仍显著低于对照组,实验组为(495.03±59.26)ρg/mL,对照组为(605.34±49.28)ρg/mL,P=0.018。TGFBR1-siRNA转染48h后,p-ERK、SMAD2/3和p-AKT蛋白表达量出现降低;72h后,p-ERK和SMAD2/3蛋白表达量仍降低。与non-siRNA转染对照组相比,IGF1R-siRNA转染72和96h后,p-AKT蛋白表达量出现降低。�OBJECTIVE Nephroblastoma is one of the most common solid tumours in children,with an annual incidence of approximately 8%-10% of all neoplasms in that group.The peak incidence occurs between 1to 5years of age.The objective of this study was to explore whether TGF-βand IGF-1signaling pathways interact to regulate the proliferation of the Wilms’ tumor.METHODS siRNA transfection experiment was used for TGFBR1-siRNA,IGF1R-siRNA being transferred to the G401 cell line.Cells were grouped as TGFBR1-siRNA,IGF1R-siRNA and non-siRNA（control group）.Each test was repeated in six wells.MTS assay was used for cell proliferation assay.Western-blot was performed to detect the expression of p-ERK,SMAD2/3and p-AKT,the concentration of FGF9 in the supernatant was determined by ELISA.RESULTS The proliferation of G401 cells was significantly decreased at the time point of 72,96-hour of TGFBR1-siRNA,IGF1R-siRNA transferred to the G401.Proliferation of non-siRNA cells were considered as 100%,and the relative proliferation rate of TGFBR1-siRNA group and control group at 72-hour were（80±14）% and（100±7）%,P=0.034.IGF1R-siRNA group and control group at 72-hour were（85±21）% and（100±7）%,P=0.028.For96-hour of TGFBR1-siRNA group and control group were（81±13）% and（100±11）%,P=0.021;IGF1R-siRNA group and control group were（82±17）% vs（100±11）%,P=0.038.The expression of p-ERK,SMAD2/3and p-AKT were decreased significantly at the 48 hfor TGFBR1-siRNA group compared with non-siRNA group detected by Western-blot.p-ERK,SMAD2/3were still decreased significantly at the 72 hcompared with non-siRNA group.The concentrations of FGF9 at 72hwere（533.85±54.64）ρg/mL in TGFBR1-siRNA group compared with non-siRNA group（587.34±46.83）ρg/mL（P=0.041）,and TGFBR1-siRNA group was（495.03±59.26）ρg/mL compared with non-siRNA group（605.34±49.28）ρg/mL for 72 htime point（P=0.018）.p-AKT was decreased significantly at the 72,96 hfor IGF1RsiRNA group compared with non-siRNA group.CONCLUSIONS The TGF-βan

关 键 词：肾母细胞瘤 信号通路 TGFBR1 IGF1R 肿瘤增殖 

分 类 号：R737.11[医药卫生—肿瘤]