壁虎醇提物体外诱导人前列腺癌PC-3细胞凋亡及机制研究

Alcohol Extract from Gecko Induces Prostate Cancer Cell Line PC-3 Apoptosis in Vitro and Possible Mechanism

目的:研究壁虎醇提物(GAE)体外诱导激素非依赖性前列腺癌PC-3细胞凋亡作用及其可能机制,为激素非依赖性前列腺癌的治疗提供理论依据。方法:以前列腺癌PC-3细胞为研究对象,MTT法检测3.5,4.0,4.5,5.0,5.5 g·L^(-1)GAE作用细胞24,48,72 h后对激素非依赖性前列腺癌PC-3细胞增殖抑制作用;3.5,4.0,5.0 g·L^(-1)的GAE作用48 h后,另设空白组,Hoeehst33342荧光染色法,Annexin V-FITC/PI双染流式细胞仪检测细胞凋亡,SP免疫组化法检测细胞内半胱氨酸蛋白酶3(Caspase 3)和Fas的表达情况并做统计学处理。结果:3.5,4.0,4.5,5.0,5.5 g·L^(-1)GAE作用细胞24,48,72 h后能显著抑制PC-3细胞的生长且呈剂量和时间依赖性;与空白组比较,3.5,4.0,5.0 g·L^(-1)的GAE作用48 h后Hoeehst33342荧光染色发现部分PC-3细胞发生典型的凋亡形态学改变;Annexin V-FITC/PI双染流式细胞仪检测显示3.5,4.0,5.0 g·L^(-1)的GAE作用48 h后,早期凋亡细胞分别占6.51%,12.48%和22.81%,且免疫组化显示细胞内蛋白Caspase-3和Fas的表达均上调且呈浓度依赖性(P<0.05)。结论:GAE能够诱导激素非依赖性前列腺癌PC-3细胞凋亡,其作用机制可能与死亡受体介导的信号通路有关。

Objective： To study the possible mechanism of the alcohol extract from gecko （GAE） on inducing prostate cancer cell line PC-3 apoptosis and provide theoretical foundation for the therapy of hormone independent prostate cancer. Method： The drug was extracted from gecko by alcohol. MTT assay was used to detect the growth inhibition effects of GAE on PC-3 ceils of AIPC. With prostate cancer （PC） -3 ceils as the research object, MTT method was used to detect the inhibition effect of 3.5,4.0, 4.5, 5.0, 5.5 g·L-1 GAE after 24, 48 and 72 h on hormone dependence after prostate cancer （PC） -3 cell proliferation; after 48h treatment by 3.5, 4.0, 5.0 g ·L-1 GAE, a blank group was established. Apoptosis-inducing effect was detected by Hoechest 33342 staining and Annexin V-FITC/PI double stained flow cytometry. The protein expression of Caspase-3 and Fas in PC-3 cells was measured by SP immunohistochemistry assay. Result ： The proliferation of PC-3 cells treated with GAE （3.5, 4.0, 4.5, 5.0, 5.5 g·L-1） for 24, 48, 72 h, respectively, was significantly inhibited in dose-and time-dependent manner. Compared with the blank group, after 48 h treatment by 3.5, 4.0, 5.0 g·L-1 GAE,typical apoptotic morphological changes were found in part of PC-3 cells by Hoechest 33342 staining. Annexin V- FITC/PI double stained flow eytometry showed that the early apoptotic cells account for 6.51% , 12.48% and 22. 81% respectively after 48 h treatment by 3.5,4.0, 5.0 g ·L-1 GAE. Immunohistochemistry assay showed that the protein expression of Caspase-3 and Fas was increased in a concentration-dependent manner （ P 〈 0.05 ）. Conclusion： GAE can induce prostate cancer cell line PC-3 apoptosis and the mechanism of action may be associated with the signal pathway mediated by death receptor.