FasL、抗DR5单克隆抗体对结肠癌细胞株HT29的杀伤作用及其机制

目的 探讨FasL、抗DR5单克隆抗体(Anti-DR5 mAb)对结肠癌细胞株HT29的杀伤作用及机制.方法 采用逆转录-聚合酶链反应(RT-PCR)、噻唑蓝(MTT)比色法、DNA倍体分析,Western blot等.结果 HT29细胞表面Fas mRNA的表达低于DIL5 mRNA的表达.50 mg/L FasL和Anti-DR5 mAb对HT29细胞的杀伤率分别为(25.49±0.90)%和(48.90±3.15)%,这种作用呈剂量依赖性.流式细胞术分析细胞周期和凋亡实验表明FasL和Anti-DR5 mAb能够抑制HT29细胞的生长,并且诱导它的凋亡.25 mg/L FasL和Anti-DR5 mAb对HT29细胞的凋亡指数分别为(13.8±1.5)%和(22.6±1.1)%.FasL和Anti-DR5 mAb作用HT29细胞后,Caspase-3蛋白表达上升,bcl-2蛋白表达水平下降.结论 FasL、Anti-DR5 mAb能不同程度的诱导结肠癌细胞株HT29凋亡,其机制与其受体和Caspase-3、bcl-2的表达有关.

Fas ligand and anti-human DR5 monoclonal antibody induce colon cancer cell line HT29 apoptosis

Objective To study the cytotoxic effects of Fas ligand and anti-human DR5 mono-clonal antibodies (Anti-DR5 mAb) on colon cell line HT29 and the underlying mechanism. Methods Fas/DR5 mRNA was detected by RT-PCR. Cytotoxicity exerted by FasL/Anti-DR5 mAb on tumor cell lines was measured by MTr colorimetry and the induced apoptosis was determined by agarose gel electro-phoresis. The cell cycle and apoptosis was assessed by flow cytometry with PI staining. Caspase-3 and bcl-2 expression was assayed by Westem blot. Results The expression of Fas mRNA in HT29 cells was lower than DR5 mRNA. HT29 cells were sensitive to anti-DR5 mAb but partially sensitive to FasL in a dose de-pendent manner. The killing rate of 50 mg/L FasL and anti-DR5 mAb to HT29 cells was (25.49± 0, 90)% and (48.90 ± 3.15 )%, respectively. The cell cycle analysis suggested that FasL/ Anti-DR5 mAb could inhibit HT29 cells in G0/G1 phase, and then further inhibit cell growth. The apoptosis rate of 25 mg/L FasL and anti-DR5 mAb to HT29 cells was ( 13.8±1.5 ) % and (22.6±1.1 ) % ,respectively. The expression of Caspase-3 was increased, but the expression of bcl-2 was decreased obviously in HT29 cells after treated with FasL/Anti-DR5 mAb. Conclusion FasL/ Anti-DR5 mAb could induce apoptosis of HT29 cells. The underlying mechanism may be relevant to Fas/DR5 mRNA expression, and the release of Caspase-3 and bcl-2.