基于PI3K/AKT/mTOR信号通路探讨莪术醇抗肝纤维化的分子机制

Molecular Mechanism of Curcumol Against Liver Fibrosis Based on PI3K/AKT/mT OR Signaling Pathway

目的:研究莪术醇对肝星状细胞PI3K/AKT/mTOR信号通路的作用,探讨莪术醇抗肝纤维化的作用机制。方法:将HSC-T6分为正常对照组、模型对照组、丹参酚酸B 70μg/mL组、莪术醇1.65、3.12、6.25μg/mL组,用10 ng/mL TGF-β1活化HSC-T6细胞模型24 h,以丹参酚酸B和莪术醇各浓度组干预造模细胞48 h,正常对照组和模型对照组用DMEM培养液进行处理;MTT比色法检测莪术醇对细胞的增殖;细胞流式检测各组细胞凋亡情况;蛋白免疫印迹法检测PI3K、AKT、mTOR的表达;RT-PCR法检测Pi3k、Akt、Mtor mRNA的表达。结果:与正常对照组比较,模型对照组肝星状细胞凋亡率显著下降(P<0.05);PI3K、AKT、mTOR蛋白及mRNA显著上调(P<0.01);与模型对照组比较,丹酚酸B 70μg/mL组、莪术醇1.65、3.12、6.25μg/mL组肝星状细胞的凋亡率明显提高(P<0.05或P<0.01);PI3K、AKT、mTOR蛋白表达明显上调,莪术醇3.12、6.25μg/mL及丹参酚酸B 70μg/mL组Pi3k、Akt、Mtor mRNA表达明显下调(P<0.05或P<0.01)。结论:莪术醇通过抑制PI3K/AKT/mTOR信号通路诱导肝星状细胞的凋亡,可能是其抗肝纤维化的分子机制之一。

Objective:To investigate the effect of curcumol on PI3 K/AKT/mTOR signaling pathway in hepatic stellate cells, and to explore the mechanism of curcumol against liver fibrosis. Methods:HSC-T6 cells were divided into a normal control group, a model group, a salvianolic acid B group(70 μg/mL), and low(1.65 μg/mL), medium(3.12 μg/mL), and high(6.25 μg/mL) dose curcumol groups. HSC-T6 cells were activated with 10 ng/mL TGF-β1 for 24 hours. Cells in the salvianolic acid B group and curcumol groups were treated with corresponding drugs for 48 hours, while those in the normal control group and the model group were treated with DMEM. MTT assay was used to detect cell proliferation in each group. Cell flow cytometry was used to detect cell apoptosis. Western blot was used to detect the expression of PI3 K, AKT, and mTOR. RT-PCR was used to detect the mRNA expression of PI3 K, AKT, and mTOR. Results:Compared with the normal control group, the model group showed decreased apoptosis rate of hepatic stellate cells(P<0.05) and up regulated protein and mRNA expression of PI3 K, AKT, and mTOR(P<0.01). Compared with the model group, the salvianolic acid B group and the curcumol groups displayed the increased apoptosis rate of hepatic stellate cells(P<0.05 or P<0.01) and up regulated protein expression of PI3 K, AKT, and mTOR. The salvianolic acid B group and the medium and high dose curcumol groups exhibited down regulated mRNA expression of PI3 K, AKT, and mTOR(P<0.05 or P<0.01).Conclusion: Curcumol induces hepatic stellate cell apoptosis by inhibiting PI3 K/AKT/mTOR signaling pathway, which may be one of the molecular mechanisms of its anti-liver fibrosis effect.