抑制Sema 4D表达介导的成骨前体细胞AKT磷酸化水平上调在乳腺癌骨转移中的作用

发布时间：2018-11-22 20:00

【摘要】：目的探讨抑制Sema 4D介导的成骨前体细胞AKT磷酸化水平上调在乳腺癌骨转移中的作用。方法通过siRNA技术构建稳定的Sema 4D低表达人乳腺癌MCF-7细胞株,采用实时荧光定量PCR检测细胞Sema 4D mRNA表达,MTT法检测各组细胞增殖能力,Transwell法检测各组细胞侵袭能力;采用Transwell小室建立成骨细胞和乳腺癌细胞的共培养体系,茜素红染色实验检测骨矿化能力,Western blot检测AKT、p-AKT蛋白表达。结果 MCF-7细胞Sema 4D mRNA相对表达量显著高于Hs578Bst细胞(P0.05)。Sema 4D siRNA转染后,Sema 4D siRNA组Sema 4D mRNA相对表达量显著低于Control组和NC组(均P0.05),Control组和NC组Sema 4D mRNA相对表达量比较,差异无统计学意义(P0.05)。Sema 4D siRNA组细胞增殖率显著低于NC组和Control组(均P0.05),Sema 4D siRNA组细胞侵袭率亦显著低于NC组和Control组(均P0.05)。MCF-7+OM组和Sema 4D siRNA+OM组骨结节面积显著低于Control+OM组(均P0.05),Sema 4D siRNA+OM组骨结节面积显著高于MCF-7+OM组(均P0.05)。Control+OM组、MCF-7+OM组、Sema 4D siRNA+OM组AKT蛋白表达水平比较差异无统计学意义(P0.05);MCF-7+OM组和Sema 4D siRNA+OM组p-AKT蛋白表达水平显著低于Control+OM组(均P0.05),Sema4D siRNA+OM组p-AKT蛋白表达水平显著高于MCF-7+OM组(P0.05)。结论人乳腺癌细胞系MCF-7 Sema4D表达显著上调,并与细胞增殖、侵袭等生物学行为有关;下调Sema 4D表达,能够提高AKT磷酸化水平,减弱对成骨细胞分化的抑制作用。
[Abstract]:Objective to investigate the role of inhibiting the up-regulation of AKT phosphorylation of osteoblasts mediated by Sema 4 D in bone metastasis of breast cancer. Methods stable human breast cancer MCF-7 cell line with low expression of Sema 4D was constructed by siRNA technique. The expression of Sema 4D mRNA was detected by real-time fluorescence quantitative PCR, the proliferation ability of each group was detected by MTT assay, and the invasion ability of each group was detected by Transwell assay. The co-culture system of osteoblasts and breast cancer cells was established by Transwell chamber. The bone mineralization ability was detected by alizarin red staining. The expression of AKT,p-AKT protein was detected by, Western blot. Results the relative expression of Sema 4D mRNA in MCF-7 cells was significantly higher than that in Hs578Bst cells (P0.05). After transfection with). Sema 4D siRNA, the relative expression of Sema 4D mRNA in Sema 4D siRNA group was significantly lower than that in Control group and NC group (P0.05). There was no significant difference in the relative expression of Sema 4D mRNA between Control group and NC group (P0.05). The cell proliferation rate of). Sema 4D siRNA group was significantly lower than that of NC group and Control group (P0.05). The cell invasion rate of Sema 4D siRNA group was significantly lower than that of NC group and Control group (P0.05). The bone nodule area in MCF-7 OM group and Sema 4D siRNA OM group was significantly lower than that in Control OM group (P0.05). The area of bone nodules in Sema 4D siRNA OM group was significantly higher than that in MCF-7 OM group (P0.05). There was no significant difference in AKT protein expression level between). Control OM group, MCF-7 OM group and Sema 4D siRNA OM group (P0.05). The expression of p-AKT protein in MCF-7 OM group and Sema 4D siRNA OM group was significantly lower than that in Control OM group (P0.05), and the p-AKT protein expression level in Sema4D siRNA OM group was significantly higher than that in MCF-7 OM group (P0.05). Conclusion the expression of MCF-7 Sema4D in human breast cancer cell line is significantly up-regulated, which is related to the biological behaviors such as cell proliferation and invasion, and down-regulating the expression of Sema _ 4D can increase the level of AKT phosphorylation and attenuate the inhibition of osteoblast differentiation.
【作者单位】：河南省人民医院骨科;
【基金】：河南省重大科技攻关项目(No.142102310081)
【分类号】：R737.9

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