非肌肉肌球蛋白重链-9在骨肉瘤组织中的表达及其对细胞上皮间质转换和侵袭能力的影响

发布时间：2019-03-20 14:56

【摘要】：目的非肌肉肌球蛋白重链-9(MYH9)对肿瘤的发生发展具有重要的调控作用,文中旨在探讨MYH9在骨肉瘤组织中的表达及对骨肉瘤细胞上皮间质转换(EMT)和侵袭能力的影响。方法收集52例骨肉瘤癌组织及距肿瘤边缘5cm处的癌旁组织,RT-PCR和免疫组化检测骨肉瘤癌组织和癌旁组织中MYH9的mRNA和蛋白表达水平。脂质体法转染U2-OS细胞MYH9 shRNA表达质粒,沉默MYH9表达,转染后将细胞分成3组:以正常的U2-OS细胞为对照组、以转染空质粒的U2-OS细胞为空载组和以转染MYH9 shRNA的U2-OS细胞为干扰组。采用RT-PCR检测转染后U2-OS细胞MYH9的mRNA水平,Western blot检测U2-OS细胞转染后MYH9以及EMT相关蛋白E-cadherin和Vimentin的蛋白水平变化,Transwell侵袭实验检测细胞的侵袭能力变化。结果 RT-PCR检测结果表明,癌旁组织中MYH9 mRNA的相对表达量(1.526±0.148)较癌组织(3.547±0.195)明显减少(P0.05);免疫组化检测结果表明,MYH9蛋白在肿瘤组织中主要表达于细胞质中,且MYH9蛋白在癌组织中表达显著高于癌旁组织,其阳性表达率分别为59.6%(31/52)和26.9%(14/52),差异有统计学意义(P0.05)。RT-PCR和Western blot检测结果表明,沉默MYH9基因表达后,干扰组MYH9 mRMA相对表达量和蛋白水平较对照组和空载组明显降低(P0.05)。与对照组相比较,干扰组骨肉瘤细胞U2-OS中E-cadherin蛋白明显上调,Vimentin蛋白出现下调。Transwell实验表明,48 h后各组均有穿出微孔滤膜的细胞,干扰组穿出微孔滤膜的细胞[(41.2±15.1)个]较对照组[(117.3±12.4)个]和空载组[(193.5±14.7)个]明显减少(P0.05)。结论 MYH9蛋白在骨肉瘤组织中表达显著高于癌旁组织,沉默MYH9可以通过降低骨肉瘤细胞上皮间质转换来降低骨肉瘤的侵袭能力。
[Abstract]:Aim non-muscular myosin heavy chain-9 (MYH9) plays an important role in regulating the tumorigenesis and development of osteosarcoma. The aim of this study was to investigate the expression of MYH9 in osteosarcoma and its effect on epithelial mesenchymal transition (EMT) and invasion ability of osteosarcoma cells. Methods the mRNA and protein expression of MYH9 in 52 cases of osteosarcoma and 5cm adjacent to the margin of tumor were detected by RT-PCR and immunohistochemistry. The MYH9 shRNA expression plasmid of U2-OS cells was transfected with lipofectamine and the expression of MYH9 was silenced. After transfection, the cells were divided into three groups: normal U2-OS cells as control group; U2-OS cells transfected with empty plasmid were used as control group and U2-OS cells transfected with MYH9 shRNA as interference group. The mRNA level of MYH9 in U2-OS cells transfected with RT-PCR was detected by, Western blot. The protein levels of MYH9, EMT-associated proteins E-cadherin and Vimentin in U2-OS cells were detected by, Western blot, and the invasive ability of U2-OS cells was detected by Transwell invasion assay. Results the results of RT-PCR showed that the relative expression of MYH9 mRNA in paracancerous tissues (1.526 卤0.148) was significantly lower than that in cancer tissues (3.547 卤0.195) (P0.05). The results of immunohistochemistry showed that MYH9 protein was mainly expressed in cytoplasm of tumor tissue, and the expression of MYH9 protein in cancer tissue was significantly higher than that in paracancerous tissue. The positive rate of MYH9 gene expression was 59.6% (31 / 52) and 26.9% (14 / 52), respectively (P0.05). The results of RT-PCR and Western blot showed that after silencing the expression of MYH9 gene, the positive expression rate was 59.6% (31 / 52) and 26.9% (14 / 52). The relative expression and protein level of MYH9 mRMA in interference group were significantly lower than those in control group and control group (P0.05). Compared with the control group, the E-cadherin protein was up-regulated and the Vimentin protein was down-regulated in the osteosarcoma cells of the interference group. Transwell test showed that after 48 hours, the cells in each group penetrated the microporous membrane. (41.2 卤15.1) cells in the interference group [(41.2 卤15.1) cells] were significantly lower than those in the control group [(117.3 卤12.4) cells] and the control group [(193.5 卤14.7) cells] (P0.05). Conclusion the expression of MYH9 protein in osteosarcoma is significantly higher than that in paracancerous tissue. Silencing MYH9 can decrease the invasion ability of osteosarcoma by reducing the epithelial interstitial transition of osteosarcoma cells.
【作者单位】：华中科技大学同济医学院附属普爱医院骨科;
【分类号】：R738.1

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