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目的应用RNA干扰技术沉默细胞内LASP-1蛋白的表达,探讨LASP-1蛋白对人肝癌HepG2细胞增殖和迁移的作用。方法根据LASP-1 mRNA编码序列设计RNA干扰靶点并构建特异性小干扰RNA(siRNA),通过脂质体转染入HepG2细胞,采用Western blot法检测LASP-1蛋白的表达。采用CCK-8增殖实验、平板克隆形成实验、Transwell和划痕愈合实验方法,观察HepG2细胞增殖及迁移情况。结果 (1)LASP-1 siRNA对LASP-1蛋白表达水平有显著的下调作用(P<0.05)。(2)体外实验结果显示,与HepG2组及阴性对照组相比,siRNA组HepG2细胞增殖和克隆形成率明显被抑制(P<0.05),其迁移能力也下降(P<0.05)。结论下调LASP-1蛋白表达可显著抑制HepG2细胞的增殖和迁移能力,提示LASP-1在肝癌细胞恶性增殖和转移过程中具有重要作用,为临床上寻找抑制肝癌增殖转移的靶点提供新的思路和方法。
Objective To investigate the effect of LASP-1 protein on the proliferation and migration of HepG2 human hepatocellular carcinoma by RNAi technique. Methods According to the coding sequence of LASP-1 mRNA, RNAi targeting sites were designed and siRNAs were constructed. The siRNAs were transfected into HepG2 cells by lipofectamine. The expression of LASP-1 protein was detected by Western blot. The proliferation and migration of HepG2 cells were observed by CCK-8 proliferation assay, plate clone formation assay, Transwell and scratch healing assay. Results (1) LASP-1 siRNA had a significant down-regulation of LASP-1 protein expression (P <0.05). (2) The results of in vitro experiments showed that compared with HepG2 group and negative control group, the proliferation and colony formation rates of HepG2 cells in siRNA group were significantly inhibited (P <0.05), and their migration ability also decreased (P <0.05). Conclusion The down-regulation of LASP-1 protein expression can significantly inhibit the proliferation and migration of HepG2 cells, suggesting that LASP-1 plays an important role in the malignant proliferation and metastasis of hepatocellular carcinoma cells, providing a new idea for finding a target for inhibiting the proliferation and metastasis of hepatocellular carcinoma And method.