GRP78在胃癌中表达水平及沉默表达后对人胃癌SGC-7901/DDP细胞增殖和凋亡的影响

Expression of GRP78 in gastric cancer and its effect on proliferation and apoptosis of human gastric cancer cell line SGC-7901/DDP

目的研究内质网分子伴侣葡萄糖调节蛋白78(glucose regulated protein 78,GRP78)在人不同胃组织(正常胃组织和胃癌组织)和细胞(正常胃上皮GES1细胞、胃癌SGC-7901细胞、多耐药性胃癌SGC-7901/DDP细胞)中的表达水平;揭示GRP78沉默表达对SGC-7901/DDP细胞增殖和凋亡的影响。方法利用实时荧光定量PCR(Real-time PCR)和免疫印迹(Western blotting)方法分别检测人正常胃组织、胃癌组织、人正常胃上皮GES1细胞、胃癌SGC-7901细胞、多耐药性胃癌SGC-7901/DDP细胞中GRP78 mRNA和蛋白表达水平。利用LipfectamineTM2000将GRP78 siRNA转染至SGC-7901/DDP细胞中,同时设置未转染Control组和无义转染Control siRNA组。Real-time PCR和Western blotting方法检测各组SGC-7901/DDP细胞转染效率;MTT方法检测各组SGC-7901细胞的生长活力变化和不同浓度顺铂DDP作用下细胞增殖抑制率;流式细胞术检测各组SGC-7901细胞凋亡情况。结果胃癌组织中GRP78 mRNA和蛋白表达水平显著高于正常胃组织(P<0.05);多耐药性胃癌SGC-7901/DDP细胞中GRP78 mRNA和蛋白表达水平显著高于胃癌SGC-7901细胞和正常胃上皮GES1细胞(P<0.05);GRP78 siRNA转染SGC-7901/DDP细胞后GRP78 mRNA和蛋白表达水平显著下降(P<0.05);与Control组和Control siRNA组相比,GRP78 siRNA转染组SGC-7901/DDP细胞生长活力显著下降,DDP作用下细胞增殖抑制率和细胞凋亡率显著增加(P<0.05)。结论 GRP78 mRNA和蛋白在胃癌组织中高表达,在多耐药性胃癌SGC-7901/DDP细胞中也高表达,证实GRP78在胃癌中发挥促癌基因的作用。而GRP78 siRNA转染能够沉默SGC-7901/DDP细胞中GRP78基因表达,抑制SGC-7901/DDP细胞增殖并促进细胞凋亡。

Objective To study the expression of molecular chaperone glucose regulated protein 78 （GRP78） in different gastric tissues （normal and gastric cancer tissues） and cells （normal gastric epithelial cells GES1, gastric cancer ceils SGC-7901 and multi-drug resistance gastric cancer cells SGC-7901/DDP） ; to reveal the effect of GRP78 silence expression on proliferation and apoptosis of SGC-7901/DDP cells. Methods Real-time fluorescence quantitative PCR （ Real-time PCR） and Western blotting methods were used to detect the expressions of GRP78 mRNA and protein on human normal gastric tissues, gastric cancer tissues and normal human gastric epithelial cells GES1, gastric cancer ceils SGC-7901 and muhidrug resistance of gastric cancer cells SGC-7901/DDP. siRNA GRP78 were transfected into SGC- 7901/DDP cells by LipfectamineYM2000, and the siRNA group and control group without transfection were set up in the control cells. Real-time PCR and Western blotting SGC-7901/DDP methods were used to detect the transfection efficiency; changes of growth activity of SGC-7901 cells and cells proliferation inhibition rate of different concentrations of cisplatin DDP were detected by MTT method; apoptosis of SGC-7901 cells was measured by flow cytometry. Results The expressions of GRP78 gene mRNA and protein in gastric cancer tissues were significantly higher than those in normal gastric cancer （P 〈 0.05） ; the expressions of GRP78 mRNA and protein of muhi-drng resistance in gastric cancer SGC- 7901/DDP cells were significantly higher than those in SGC-7901 gastric cancer cells and normal gastric epithelial cells GES1 （P 〈 0.05） ; the expressions of GRP78 mRNA and protein were decreased significantly in SGC-7901/DDP cells transfected with GRP78 siRNA （P 〈 0.05 ） ; compared with control group and control siRNA group, cells growth activity was significantly decreased in GRP78 siRNA transfected SGC-7901/DDP group, cells proliferation inhibition rate and apoptosis rate were significantly increased （P 〈 0.05）. Conclusion GRP78 mRNA and protein show high expressions in gastric cancer and in multidrug resistance of gastric cancer cells SGC-7901/DDP, GRP78 plays a role in promoting cancer gene in gastric cancer, siRNA GRP78 transfection can silence the expression of GRP78 gene in SGC-7901/DDP cells, inhibit the proliferation of SGC-7901/DDP cells and promote cell apoptosis.