白藜芦醇对NIH3T3细胞Shh信号通路的影响

Effects of resveratrol on Shh signaling pathway of NIH3T3 cells

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。

Objective To investigate the effect of resveratrol on Shh signaling pathway of NIH3 T3 cells in vitro.Methods NIH3 T3 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 NIH3 T3 cells,and the peak was 1. 0 μmol/L. On the contrary,10,20,40,80μmol/L resveratrol significantly decreased the viability of NIH3 T3 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）. NIH3 T3 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 NIH3 T3 cells via activating Shh signaling pathway.