聚焦超声诱导卵巢癌细胞株COC1凋亡及其机制的初步研究

Preliminary study of apoptosis in human ovarian cell line COC1 induced by ultrasound and its mechanism

目的 :研究低频脉冲聚焦超声波对人卵巢癌细胞株COC1的抗肿瘤作用 ,并探讨其诱导细胞凋亡的分子机制。方法 :以低频脉冲聚焦超声 (超声频率 0 .8MHz ,声强 83.6W /cm2 ,脉肿持续时间 :5 0 μs)辐照卵巢癌COC1细胞 10s ,然后采用MTT法观察超声对COC1细胞增殖的抑制作用 ;流式细胞仪 (FCM )、TUNEL法、荧光染色和光、电镜检测细胞凋亡 ;应用FCM免疫荧光技术观察Bcl- 2、bax基因蛋白表达的变化。结果 :超声可明显抑制存活卵巢癌COC1细胞的增殖 ,辐照后 12h、2 4h、36h、4 8h ,COC1细胞MTT光吸收值与对照组相比显著降低 ;FCM检测超声辐照后 6h的COC1细胞凋亡率达 ( 2 3.5 7± 4 .6 2 ) % ;Bcl- 2蛋白的荧光指数 (FI)降低 ,而bax蛋白FI升高。同时TUNEL法观察到棕褐色着染的凋亡细胞 ;荧光显微镜和电镜下观察到凋亡小体形成等典型的形态学改变。结论 :低频脉冲聚焦超声辐照对人卵巢癌细胞株COC1具有细胞毒作用 ,其机制为诱导辐照后存活的人卵巢癌细胞株COC1凋亡 ,bcl- 2和bax蛋白可能参与了COC1细胞凋亡过程。

Objective:To investigate the antitumor effect of Low-Frequency Pulse Forcusd Ultrasound(LFPFU) exposure in human cell line COC1,and to clarify its probable mechanism.Methods:The growth inhibitory effect of ultrasound exposure[0.8MHz,83.6W/cm 2,pulse duration(PD) 50μs;sonication time of 10s] on COC1 cell was observed by MTT colormetric assay.Apoptosis was analyzed using flow cytometry(FCM) and terminal deoxynucleotidyl transferase-mediated dUTP-digoxin nick end labeling(TUNEL),and fluorescent microscope and electron microscope were applied to detect the morphological changes of COC1 cell.The protein expression of bcl-2 and bax affected by LFPFU were determined by FCM and immunofluorescent techniques.Results:Sonication could obviously suppress proliferation of COC1 cells surviving ultrasound exposure.The absorbance value(A 570 )of MTT decreased significantly at 12h,24h,36h and 48h after sonication as compared with control group.The hightest apoptosis rate was found to reach 23.57±4.62% by FCM,and brown-colored positive apoptotic cells were observed with TUNEL method at 6h after sonication.Typical morphological changes of apoptosis such as apoptotic body was found under fluorescent microscope and transmission electron microscope.Fluorescence index(FI) of bcl-2 decreased,and Fl of bax increased after LFPFU exposure.Conclusion:LFPFU can induce apoptosis in vitro in human ovarian cell line COC1 surviving ultrasound exposure.The bcl-2 and bax protein may be involved in the apoptotic process.