辛二酰苯胺异羟肟酸对人肝星状细胞增殖和凋亡的影响

Effects of suberoylanilide hydroxamic acid on proliferation and apoptosis of human hepatic stellate cells

目的研究组蛋白去乙酰化酶抑制剂辛二酰苯胺异羟肟酸(SAHA)对人肝星状细胞系LX-2增殖及凋亡的影响及其可能机制。方法体外应用SAHA作用于LX-2细胞,倒置显微镜观察LX-2细胞形态,MTT法检测细胞增殖;荧光显微镜及流式细胞仪Annexin V-FITC/PI法检测细胞凋亡率;Western blot检测α-SMA、Ⅰ型胶原、ac H3K9、ac H3K14和ac H3K18蛋白表达。结果 SAHA呈剂量依赖性显著抑制LX-2细胞增殖(P<0.05);SAHA对LX-2细胞凋亡具有呈时间依赖性的促进作用(P<0.05);SAHA处理LX-2细胞后,α-SMA和Ⅰ型胶原蛋白表达水平明显降低(P<0.05),而ac H3K9、ac H3K14和ac H3K18乙酰化修饰水平明显升高(P<0.01)。结论 SAHA抗肝纤维化的机制可能与下调α-SMA及Ⅰ型胶原蛋白表达,上调组蛋白ac H3K9、ac H3K14和ac H3K18乙酰化修饰水平有关。

Objective To determine the effects of histone deacetylase inhibitor suberoylanilide hydroxamic acid（ SAHA） on the cell proliferation and apoptosis of the human hepatic stellate cell line LX-2. The possible underlying mechanisms were also investigated. Methods The LX-2 cells were treated with SAHA in vitro. The morphology of LX-2 cells in different concentrations groups was observed by inverted microscope; the proliferation of LX-2 cells was measured by MTT assay; the Annexin V-FITC and PI staining was used to detect the apoptosis of LX-2 cells by flow cytometry and fluorescence microscope; the expression of α-SMA,collagen Ⅰ,ac H3K9,ac H3K14 and ac H3K18 were detected by Western blot. Results The morphology change of LX-2 cells showed that SAHA inhibited the proliferation rate of LX-2 cells and in a dose dependent manner（ P〈0. 05）. The LX-2 cells were sensitive to SAHA along with time increasing,and in a time-dependent manner（ P〈0. 05）. Western blot showed that the expression levels of α-SMA and collagen-Ⅰ were significantly lower（ P〈0. 05）,on the contrary,the acetylation levels of ac H3K9,ac H3K14 and ac H3K18 were significantly higher（ P〈0. 05）. Conclusions The increased acetylation of the histone ac H3K9,ac H3K14,ac H3K18 and the lower expressed α-SMA and collagen-Ⅰ in LX-2 cells may be one of the mechanisms of SAHA.