K7M2细胞转染Runx2基因后对MC3T3细胞RANKL基因表达的影响

Effect on the RANKL gene Expression of MC3T3 Cell of K7M2 Cell Transfected by Runx2

目的探讨转染Runx2基因后对RANKL及相关细胞因子的影响。方法将鼠骨肉瘤K7M2细胞分成3组,K7M2组,K7M2-neo组和K7M2-Runx2组,应用Westernblot方法检测转染是否成功,选择转染成功的细胞连同K7M2组,K7M2-neo组细胞,应用ELISA方法检测培养上清液中的IL-1α,IL-6,IL-11和PGE2的含量变化,并应用RT-PCR的方法检测应用K7M2组,K7M2-neo组和K7M2-Runx2组以及在K7M2-Runx2组细胞中加入IL-6抗体后的培养液来培养的MC3T3细胞RANKL基因表达的变化,并应用Westernblot方法检测K7M2细胞IL-6R蛋白的表达变化。结果转染Runx2基因后可以有效提高Runx2蛋白的表达,培养的上清液中的IL-6含量明显提高,应用K7M2-Runx2组细胞培液培养MC3T3细胞可以提高其RANKL基因表达,在K7M2-Runx2组细胞培液中加入IL-6抗体后则不会提高MC3T3细胞的RANKL基因表达。结论Runx2基因可以提高MC3T3细胞RANKL基因的表达,这种基因表达的提高是通过IL-6因子来介导的。

Objective To investigate the effect on the RANKL and related cytokines after transfecting Runx2 gene.Methods Dividing the mouse osteosarcoma KTM2 cell line into three groups, K7M2 group, K7M2-neo group and K7M2-Runx2 group,detected if transfected succeedly with Westernblot assay. Detected the level of IL-1α, IL-6, IL-11 and PGE2 in supernatant with ELISA assay in K7M2 group,K7M2-neo group, Runx2 transfected succeedly K7M2 group,the expression of RANKL gene of MC3T3 cells with RTPCR method which cultured with K7M2, K7M2-neo, K7M2-Runx2 and K7M2-Runx2 added IL-6 antibody cells supematant. IL-6R was detected with Westernblot method as well. Results Transfecting Rumx2 gene can increase the expression of Runx2 protein , level of IL-6 increased in supematant after tmngecting Runx2, the supematant from K7M2-Runx2 can elevated the RANKL expression , IL-6 antibody can antagonize the RANKL increasing of MC3T3. Conclusion Runx2 gene can increase the expression of RANKL, it can be mediated by IL-6.