短发卡RNA抑制黑素瘤细胞BRAF^(V600E)突变基因的研究  

In vivo and in vitro suppression of BRAF^(V600E) in human melanoma cells by short hairpin RNA

作　　者：陈浩[1] 韩永智 周武庆[1] 曾学思[1] 孙建方[1] 

机构地区：[1]中国医学科学院,中国协和医科大学皮肤病研究所,南京210042 [2]广州省人民医院

出　　处：《中华皮肤科杂志》2007年第6期359-361,共3页Chinese Journal of Dermatology

基　　金：江苏省六大高峰人才基金资助

摘　　要：目的探讨BRAF^(V600E)突变基因在黑素瘤发展过程中的作用。方法用构建成功的质粒载体转染人黑素瘤细胞株A375,并建立动物模型来检测其在体内的干扰作用。分别采用RT-PCR和蛋白质印迹法检测体内外BRAF基因的mRNA和蛋白表达。结果特异性短发卡RNA在体内外均可抑制人黑素瘤细胞BRAF基因mRNA和蛋白的表达,使其蛋白表达量分别减少62%和90%。在动物模型中使黑素瘤细胞的成瘤性下降,可以抑制肿瘤增长缓慢,抑瘤率达到62%。结论BRAF^(V600E)突变基因在黑素瘤的发展中有一定的作用。Objective To investigate the role of BRAF^V600E in the development of melanoma, as well as in vivo and in vitro suppression of BRAF^V600E by short hairpin RNA （ shRNA ）. Methods Previously constructed shRNAs targeted against the BRAF gene were used. Three plasmids, including two BRAF- specific shRNA plasmids, braf 1 and braf 2, and a negative control plasmid, were constructed and transfected into human melanoma cell line A375. A xenograft nude mouse model was used to examine the inhibitory effect ofshRNA in vivo. The expressions ofBRAF mRNA and protein were detected by RT-PCR and Western blotting, respectively. Results The specific shRNA inhibited the in vivo and in vitro expression of BRAF mRNA and protein; protein level was decreased by 62% in vivo and 90% in vitro. In the mouse model, the tumorigenicity was decreased by the specific shRNA and the growth of tumor was inhibited by 62% Conclusion The specific shRNAs suppress the mRNA and protein expression of BRAF^V600E, and this gene may play an important role in the development of melanoma.

关 键 词：SHRNA 黑色素瘤 实验性 基因 BRAF RNA干扰 

分 类 号：R739.5[医药卫生—肿瘤]