【摘要】：目的探討抑制Sema 4D介導(dǎo)的成骨前體細(xì)胞AKT磷酸化水平上調(diào)在乳腺癌骨轉(zhuǎn)移中的作用。方法通過siRNA技術(shù)構(gòu)建穩(wěn)定的Sema 4D低表達(dá)人乳腺癌MCF-7細(xì)胞株,采用實(shí)時(shí)熒光定量PCR檢測細(xì)胞Sema 4D mRNA表達(dá),MTT法檢測各組細(xì)胞增殖能力,Transwell法檢測各組細(xì)胞侵襲能力;采用Transwell小室建立成骨細(xì)胞和乳腺癌細(xì)胞的共培養(yǎng)體系,茜素紅染色實(shí)驗(yàn)檢測骨礦化能力,Western blot檢測AKT、p-AKT蛋白表達(dá)。結(jié)果 MCF-7細(xì)胞Sema 4D mRNA相對表達(dá)量顯著高于Hs578Bst細(xì)胞(P0.05)。Sema 4D siRNA轉(zhuǎn)染后,Sema 4D siRNA組Sema 4D mRNA相對表達(dá)量顯著低于Control組和NC組(均P0.05),Control組和NC組Sema 4D mRNA相對表達(dá)量比較,差異無統(tǒng)計(jì)學(xué)意義(P0.05)。Sema 4D siRNA組細(xì)胞增殖率顯著低于NC組和Control組(均P0.05),Sema 4D siRNA組細(xì)胞侵襲率亦顯著低于NC組和Control組(均P0.05)。MCF-7+OM組和Sema 4D siRNA+OM組骨結(jié)節(jié)面積顯著低于Control+OM組(均P0.05),Sema 4D siRNA+OM組骨結(jié)節(jié)面積顯著高于MCF-7+OM組(均P0.05)。Control+OM組、MCF-7+OM組、Sema 4D siRNA+OM組AKT蛋白表達(dá)水平比較差異無統(tǒng)計(jì)學(xué)意義(P0.05);MCF-7+OM組和Sema 4D siRNA+OM組p-AKT蛋白表達(dá)水平顯著低于Control+OM組(均P0.05),Sema4D siRNA+OM組p-AKT蛋白表達(dá)水平顯著高于MCF-7+OM組(P0.05)。結(jié)論人乳腺癌細(xì)胞系MCF-7 Sema4D表達(dá)顯著上調(diào),并與細(xì)胞增殖、侵襲等生物學(xué)行為有關(guān);下調(diào)Sema 4D表達(dá),能夠提高AKT磷酸化水平,減弱對成骨細(xì)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.
【作者單位】： 河南省人民醫(yī)院骨科;
【基金】：河南省重大科技攻關(guān)項(xiàng)目(No.142102310081)
【分類號】：R737.9

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