磷脂酰肌醇蛋白多糖-3对肝癌细胞增殖和凋亡的影响

Effects of silencing glypican- 3 gene transcription on proliferation and apoptosis of hepatoma cells

目的探讨shRNA干预磷脂酰肌醇蛋白多糖(GPC)-3基因转录对肝癌细胞增殖和凋亡的影响。方法将GPC-3-shRNA插入pGPU6/GFP/Neo质粒,转染人肝癌HepG2细胞,以Western Blot和荧光定量PCR分别分析GPC-3蛋白和mRNA表达;以MTT法分析HepG2细胞增殖;以流式细胞术、Annexin-V-PE/7-AAD及细胞凋亡-DNA ladder等分析细胞周期及细胞凋亡率。结果 shRNA1转染HepG2细胞,GPC-3 mRNA沉默效率为89.3%,与GPC-3蛋白下调一致;以shRNA转染HepG2细胞,增殖抑制率为71.1%;细胞周期阻滞在G1期,细胞凋亡率达65.6%。结论资料显示shRNA干预GPC-3基因转录,可显著抑制肝癌细胞增殖,促进肝癌细胞凋亡。

Objective To investigate the effects of silencing glypican -3 （GPC -3） gene transcription by shRNA on the proliferation and apoptosis of hepatoma cells. Methods GPC - 3 - shRNA was inserted into pGPU6/GFP/Neo vector, and HepG2 cells were transfected with the vector. The mRNA and protein expression of GPC -3 was measured by fluorescence quantitative PCR and Western blot; the proliferation of HepG2 cells was evaluated by MTT assay; the cell cycle and apoptosis of HepG2 cells were analyzed by flow cytometry, Annexin -V -PE/7 -AAD staining, and apoptotic DNA ladder kit. Results After the HepG2 cells were transfected with GPC -3 -shRNA, the GPC -3 mRNA silencing rate was up to 89.3%, in accordance with the down - regulation of GPC - 3 protein ; the proliferation inhibition rate of HepG2 cells was as high as 71.1% ; the HepG2 cells were arrested in GI phase, and the apoptosis rate of HepG2 cells reached 65.6%. Conclusion Silencing GPC - 3 gene transcription by shRNA can significantly inhibit the proliferation and promote the apoptosis of hepatoma cells.