Let-7b和miR-199a对B16F10细胞增殖生长的影响  

The effect of let-7b and miR-199a on B16F10 cell growth and proliferation

作　　者：周建大[1] 谭建湘[1] 谢慧清[1] 蒋碧梅[2] 周鸣[2] 刘海航[3] 李明[1] 张彦文[1] 徐丹[1] 陈继业[1] 李雄[1] 罗成群[1] 

机构地区：[1]中南大学湘雅三医院烧伤整形科,长沙410013 [2]中南大学基础医学院 [3]中南大学医学图书馆

出　　处：《中国医师杂志》2011年第6期721-726,共6页Journal of Chinese Physician

基　　金：国家自然科学基金（81071645）、湖南省自然科学基金（10JJ3030）、湖南省卫生厅科研基金（B2010031）、湘雅三医院“125”人才基金和中南大学杰青培育专项基金资助

摘　　要：目的研究let-7b和miR．199a对黑色素瘤增殖生长的影响。方法设计靶向let-7b和miR．199a基因的过表达质粒和inhibitor干扰片段，将同一株系B16FIO细胞分为七组：空白组、let-7b质粒组、miR-199a质粒组、空质粒组、let-7binhibitor组、miR-199ainhibitor组、inhibitor对照组，运用基因转染技术将对应组别的外源基因导人B16F10细胞中，从RNA水平、蛋白水平和细胞水平三方面检测let-7b和miR-199a基因对B16F10细胞的影响。结果let-7b质粒组和miR-199a质粒组B16FIO细胞中对应基因的表达为3．8776±0．1372和2．8660±O．2821，明显高于空白组（P〈0．05）；let-7binhibitor组和miR-199ainhibitor组B16F10细胞中对应基因的表达为0．2057±0．0263和0．2656±0．0253，明显低于空白组（P〈0．05）；B16F10细胞中cyclinDl的表达let-7b质粒组（2．023±0．315）和let-7binhibitor组（1．857±0．377）明显高于空白组（0．997±0．041）（P〈0．05），而B16F10细胞中Met的表达中miR．199a质粒组（5．19±0．309）和miR-199ainhibitor组（4．87±0．044）明显高于空白组（2．2±0．198）（P〈0．05）；let-7b质粒组和miR-199a质粒组B16FIO细胞的生长趋势较空白组缓慢，尤其到转染后第3天，此生长趋势下降至最低值（P〈0．05），此后又缓慢趋近正常；此外，let-7b质粒组和miR-199a质粒组B16FlO细胞凋亡率分别为（11．8-4-1．19）％和（11．3±1．59）％，明显高于空白组（5．77±1．74）％（P〈0．05）。结论let-7b和miR-199a可负调控黑色素瘤细胞的增殖生长。Objective To confirm whether or not let-7b and miR-199a were significantly associated with malignant melanoma growth and proliferation. Methods An over-expression plasmid and an inhibi-tor, which targeted on let-7b and miR-199a, was constructed. B16F10 ceils were divided into seven groups： control group, let-7b plasmid group, miR-199a plasmid group, empty plasmid group, let-7b inhibitor group, miR-199a inhibitor group, inhibitor control group. Foreign gene was transfected into B16F10 cells, let-7b and miR-199a expression were validated from RNA level, protein level and cell level. Results The relative let-7b or miR-199a gene expression of the let-7b plasmid group （3. 8776±0. 1372）and miR- 199a plasmid group （2. 8660 ±0. 2821 ）were significantly higher than control group （ P 〈0. 05）, the relative let-7b or miR-199a gene expression of the let-7 b inhibitor group （0. 2057 ±0. 0263 ） and miR-199 a in- hibitor group（0. 2656 ±0. 0253 ） were significantly lower than control group（ P 〈 0. 05 ）. The cyclinD1 expression of the let-7b plasmid group （2. 023 ± 0. 315 ） and let-7b inhibitor group （1. 857 ± 0. 377 ） were significantly higher than control group （0. 997 ±0. 041 ） （ P 〈0. 05） , whereas, the Met expression of the miR-199a plasmid group （5.19 ±0. 309） and miR-199a inhibitor group （4. 87±0. 044） were significantly higher than control group （2. 2±0. 198） （ P 〈 O. 05 ）. The let-7b plasmid group and miR-199a plasmid group B16F10 cell growth rate were slower than control group, especially on the third day after transfection, the growth rate gradually dropped to the lowest value （ P 〈 0. 05）. In addition, the apoptosis rates of the let-7b plasmid group and miR-199a plasmid group reach to （ 11.8±1.19）% and （ 11.3±1.59）% ,which were significantly higher than control group （ P 〈 0.05 ）. Conclusions let-7b and miR-199a may be a negative regulator on the B16FIO cell growth and proliferation.

关 键 词：微RNAs/药理学 黑色素瘤/病理学/药物疗法 细胞增殖 细胞生长过程 

分 类 号：R739.5[医药卫生—肿瘤]