RNA干扰对比观察HIF1α和HIF2α在人肾癌细胞中的作用及机制  

Effect of HIF1α and HIF2α on human renal cell carcinoma and their mechanisms: an RNAi approach comparison

作　　者：王丽 王素霞[2] 梅长林[2] 

机构地区：[1]上海武警总队医院肾内科,上海201103 [2]第二军医大学长征医院肾内科解放军肾脏病研究所,上海200003

出　　处：《第二军医大学学报》2006年第6期590-593,共4页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金面上项目(30271523)~~

摘　　要：目的:运用RNA干扰(RNAi)方法对比观察HIF1α和HIF2α在人肾癌细胞的作用并探讨相关机制。方法:分别将化学合成的HIF-1α和HIF-2α小片段寡核苷酸脂质体瞬时转染人肾癌细胞(A498),人正常肾小管上皮细胞(HK-2)和A498分别加入培养基和脂质体作为对照组。用RT-PCR或免疫印迹等观察RNAi 24 h后一些基因如VEGF、ET-1、bcl-2和Ki67的改变,并用流式细胞仪和荧光显微镜观察细胞的凋亡情况。结果:与HK-2相比,A498高表达HIF-1α和HIF-2α,RNAi后两基因基本被沉寂。HIF-1α干扰后ET-1、bcl-2和Ki67均下调,伴随凋亡或坏死细胞增多,而HIF2α干扰后,仅VEGF下调较为明显。结论:HIF-1α和HIF-2α均有一定程度调节血管生成基因的作用,但HIF-1α尚具备一定的调节肿瘤细胞增殖功能,可能通过调节凋亡抑制基因实现。提示HIF-1α和HIF-2α可能存在不同的转录调控功能和基因结合位点。Objective： To investigate and compare the effect and the possible mechanisms of 2 hypoxia-inducible factors （HIF） 1α and 2α on renal cell carcinoma （RCC） by RNA interference （RNAi） method. Methods： Chemically synthesized HIF- 1α and HIF-2α RNAi were transfected into RCC cell line A498 by liposome transient transfection. Normal proximal tubule cell line HK-2 and A498 were separately treated with medium and liposome as controls. The expressions of VEGF, ET-1, bcl-2 and Ki67 were observed by RT-PCR or Western blot 24 h after RNAi. Cell apoptosis was detected by flow cytometry and fluorescence microscopy. Results： Expressions of HIF-1α and HIF-2α RNAi were higher in A498 than those in HK-2, and the 2 genes were successfully blocked after RNAi. HIF-1α RNAi resulted in a significant decrease of ET-1, bcl-2 and Ki67 expression, with increased apoptosis and necrosis of cells. HIF-2α RNAi only resulted in a decrease of VEGF. Conclusion： Both HIF-1α and HIF-2α play important roles in regulation of angiogenesis; moreover, HIF-1α might also regulate proliferation of tumor cells, possibly through regulating anti-apoptotic genes. It is implied the existance of different transcription regulation functions and gene combining sites between HIF-1α and HIF-2α.

关 键 词：低氧诱导因子 肾肿瘤 细胞凋亡 RNA干扰 

分 类 号：R737.11[医药卫生—肿瘤]