cyclin D1反义寡核苷酸对胃癌细胞生长和G_1期调控的影响

Effect of cyclin D1 Antisense Oligodeoxyneucleotides on the Growth and G_1 Phase Regulators in Gastric Carcinoma Cells

目的 研究cyclinD1反义寡核苷酸 (ASODN )对胃癌细胞SGC790 1和HS746T的生长、细胞周期和G1期调控的影响。方法 采用脂质体LipofectAMINE 2 0 0 0包裹硫代修饰的cyclinD 1ASODN转染细胞 ,观察量效曲线和生长曲线 ,应用RT PCR检测cyclinD1mRNA的表达 ,流式细胞仪检测细胞周期 ,并用免疫组织化学技术检测胃癌细胞中cyclinD 1、p2 1、p2 7、pRb蛋白的表达。结果 cyclinD 1ASODN对胃癌细胞产生剂量依赖性的抑制效应 ,0 .2 μmol/LcyclinD 1ASODN转染 2 4h能显著抑制胃癌细胞生长 ,cyclinD 1mRNA表达分别是对照组的 2 6.3 % (SGC790 1)和 17.3 % (HS746T ) ,G0 /G1期比例明显增加 ,并使 2种细胞中cyclinD 1、pRb表达降低 ,p2 1表达升高 ,在HS746T中p2 7表达下降 ,但在SGC790 1中 p2 7表达无改变。 结论 cyclinD1反义寡核苷酸能有效抑制胃癌细胞的生长 ,降低cyclinD1mRNA表达水平 。

Objective To investigate the effect of cyclin D1 antisense oligodeoxyneucleotides(ASODN) on the growth,cell cycle and expression of G 1 phase regulators in human gastric carcinoma cell lines SGC7901 and HS746T.Methods Phosphorothioate modified cyclin D1 ASODN were encapsulated by LipofectAMINE2000 and transfected into gastric carcinoma cells,the dose-effect curve and growth curve were observed,the expression of cyclin D1mRNA was detected by RT-PCR,the cell cycle was analyzed by flow cytometry,and the expression of cyclin D1,p21,p27,pRb in cancer cells were detected by immunohistochemical staining.Results Dose-dependent inhibitory effect was caused by cyclin D1 ASODN in two gastric carcinoma cells.0.2 μmol/L cyclin D1 ASODN used in gastric carcinoma cells for 24 hours could inhibit growth significantly,reduce expression of cyclin D1mRNA to 26.3%(SGC7901) and 17.3%(HS746T) respectively.The percentage of cells in G 0/G 1 phase increased,the expression of p21 increased and the expression of cyclin D1 and pRb decreased in two cell lines.The expression of p27 increased in HS746T,but kept unchanged in SGC7901.Conclusion cyclin D1 ASODN can inhibit the growth and the expression of cyclin D1 mRNA in gastric carcinoma cells.It can also influence the cell cycle and the expression of cell cycle regulator.