白藜芦醇对NIH3T3细胞Shh信号通路的影响

目的探讨白藜芦醇对NIH3T3细胞Shh信号通路的影响。方法实验分为对照组和白藜芦醇组。白藜芦醇处理NIH3T3细胞24 h。CCK-8法测定细胞活力,免疫荧光法检测初级纤毛蛋白(Ac-tu)、Smo和Gli-1的表达,Western blotting法检测Shh、Ptc、Smo、Gli-1蛋白的表达。结果 1.0.5和1μmol/L白藜芦醇组(0.679±0.047,0.774±0.054)细胞活力分别较对照组(0.585±0.039)明显增强(P<0.05),其中以1μmol/L白藜芦醇组最强。之后随白藜芦醇浓度(10、20、40、80μmol/L)的增高,细胞活力逐渐降低(0.428±0.043,0.395±0.031,0.373±0.017,0.361±0.016),且均较对照组明显减弱(P<0.05),而0.1μmol/L(0.602±0.065)和5μmol/L组(0.556±0.041)细胞活力与对照组比较差异无显著性(P>0.05)。2.NIH3T3细胞表达Ac-tu、Shh、Ptc-1、Smo、Gli-1从蛋白,有1根初级纤毛,Smo和Gli-1蛋白位于细胞质。白藜芦醇处理24 h时,Gli-1从细胞质进入细胞核,Smo从细胞质进入初级纤毛,且Shh(0.756±0.659比0.441±0.769,P<0.05)、Ptc-1(0.655±0.347比0.351±0.026,P<0.05)、Smo(0.779±0.064比0.451±0.035,P<0.05)和Gli-1(0.856±0.044比0.560±0.058,P<0.05)蛋白表达较对照组高。结论白藜芦醇可能通过激活Shh信号通路增强NIH3T3细胞的活力。

Effects of resveratrol on Shh signaling pathway of NIH3T3 cells

Objective To investigate the effect of resveratrol on Shh signaling pathway of NIH3T3 cells in vitro. Methods NIH3T3 cells were divided into the control and resveratrol groups. The cells were cultured with resveratrol for 24 hours. Cell viability was detected with CCK-8 assay．Immunofluorescence measured the expressions of Ac-tu, Smo, and Gli-1. Western blotting assayed the expressions of Shh, Ptc-1, Smo and Gli-1 proteins. Results Compared with the control group（0.585±0.039）, 0.5（0.679±0.047, P＜0.05 and 1.0 μmol/L（0.774±0.054, P＜0.05 resveratrol significantly enhanced the viability of NIH3T3 cells, and the peak was 1.0 μmol/L. On the contrary, 10, 20, 40, 80 μmol/L resveratrol significantly decreased the viability of NIH3T3 cells（0.428±0.043, 0.395±0.031, 0.373±0.017, 0.361±0.016, respectively). However, 0.1（0.602±0.065）and 5 μmol/L（0.556±0.041）resveratrol did not affect the viability（P>0.05）. NIH3T3 cells had one primary cilia and expressed the proteins of Ac-Tu, Shh, Ptc-1, Smo and Gli-1.Smo and Gli-1 proteins were located in the cytoplasm. At 24 hours for resveratrol treat, Gli-1 protein translocated into the nucleus from cytoplasm and Smo protein entered the primary cilia from the cytoplasm. Expressions of Shh（0.756±0.659 vs 0.441±0.769，P＜0.05,Ptc-1（0.655±0.347 vs 0.351±0.026，P＜0.05,Smo（0.779±0.064 vs 0.451±0.035，P＜0.05 and Gli-1（0.856±0.044 vs 0.560±0.058，P＜0.05 proteins significantly compared with the control group. Conclusion Resveratrol can enhance viability of NIH3T3 cells via activating Shh signaling pathway.