肿瘤相关线粒体蛋白12对肝癌细胞凋亡的抑制作用  

Inhibitory effects of tumor associated mitochondrial protein 12 on HepG2 cell apoptosis

作　　者：金姝[1] 王俐[2] 王颖[1] 谢国化[1] 陈惠娟[1] 王树军[1] 张惠珍[1] 张勇[1] 葛瑜[1] 葛海良[1] 

机构地区：[1]上海交通大学医学院上海市免疫学研究所,上海200025 [2]上海交通大学医学院公共卫生学院营养与食品卫生系,上海200025

出　　处：《中国病理生理杂志》2009年第2期220-225,共6页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.30572117);上海市教委科研创新资助项目(No.08YZ39)

摘　　要：目的:探讨肿瘤相关线粒体蛋白12(TAMP12)对肿瘤细胞凋亡的抑制作用。方法:(1)构建重组逆转录病毒表达载体并转染包装细胞PA317,将获得的病毒颗粒感染TAMP12低表达的肝癌细胞株HepG2。应用RT-PCR检测TAMP12 mRNA表达变化;激光扫描共聚焦显微镜(CLSM)观察双色荧光标记蛋白的亚细胞定位和定量表达。(2)应用Hoechst 33258染色和FACS检测5-氟尿嘧啶(5-FU)处理后对诱导HepG2细胞凋亡的影响。结果:(1)CLSM检测显示TAMP12主要表达于HepG2细胞线粒体中。转染细胞中TAMP12基因和蛋白呈稳定高表达。(2)5-FU诱导细胞发生凋亡后,转染细胞的核形态较为完整,但对照细胞的染色质发生凝聚、边缘化、核固缩;且FACS检测显示转染细胞的凋亡率显著低于对照细胞(P<0.05或P<0.01)。结论:TAMP12具有抑制肝癌细胞凋亡的作用。AIM： To explore the inhibitory effects of tumor associated mitochondrial protein 12 （TAMPI2） on tumor cell apoptosis. METHODS ： （ 1 ） A retrovirus expression vector was recombinated and transfected into the packaging cell line PA317. The virus particles were obtained to infect the target cell line HepG2 low expressing of TAMP12. The expression of TAMP12 mRNA was detected by RT - PCR. The subcellular localization and quantification of TAMP12 protein labeled with double fluorescein were observed under colffocal laser scanning microscope （CLSM）. （2） Hoechst33258 staining and flow cytometry （FACS） were used to analysis the apoptosis of HepG2 ceils treated with 5 - fluorouracil （5 - FU）. RESULTS： （1） The CLSM observation showed that TAMP12 protein was mainly expressed in mitochondria of HepG2 cells. The expressions of TAMP12 gene and protein were stable and high in transfected HepG2 ceils. （2） Upon treatment with 5 - FU, the transfected HepG2 cells showed a fairly integrated nucelus while the control HepG2 cells exhibited chro- matin condensation, marginalization and karyorhexix. Moreover, the apoptosis rate of transfeced HepG2 cells was significandy lower than that in control HepG2 cells （P 〈0. 05 or P 〈0. 01） by FACS. CONCLUSION： TAMP 12 is an anti - apoptotic element in 5 - FU induced HepG2 cell apoptosis.

关 键 词：肿瘤相关线粒体蛋白12 线粒体 逆转录病毒载体 HEPG2细胞 氟尿嘧啶 细胞凋亡 

分 类 号：R363[医药卫生—病理学]