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目的:探讨pEGFP-N1-NPRL2真核表达载体对SGC-7901胃癌细胞体外增殖﹑细胞周期、凋亡、侵袭及迁移的影响。方法:构建pEGFP-N1-NPRL2真核表达载体,并进行酶切及测序鉴定。脂质体介导转染入SGC-7901胃癌细胞,应用荧光显微镜、RT-PCR、Western blot法检测NPRL2的基因及蛋白水平情况,CCK8法检测细胞增殖的变化,流式细胞仪分析细胞周期和凋亡率的变化,Transwell实验检测NPRL2对细胞迁移及侵袭能力的影响。结果:成功构建了pEGFP-N1-NPRL2真核表达载体。重组质粒转染SGC-7901细胞后,通过荧光显微镜观察到绿色荧光的表达,RT-PCR和Westernblot法检测到NPRL2 mRNA及蛋白的表达,CCK8法检测发现NPRL2能够明显抑制肿瘤细胞增殖(P<0.05),细胞周期分析显示NPRL2使细胞停在G0～G1期(P<0.05),细胞凋亡结果显示NPRL2能抑制细胞凋亡(P<0.05),Transwell侵袭和迁移实验显示NPRL2使细胞侵袭迁移能力均降低(P<0.05)。结论:NPRL2可抑制肿瘤细胞的增殖、周期、侵袭及迁移,并促进凋亡。
Objective: To investigate the effect of pEGFP-N1-NPRL2 eukaryotic expression vector on proliferation, cell cycle, apoptosis, invasion and migration in vitro of gastric cancer cell line SGC-7901. Methods: The eukaryotic expression vector pEGFP-N1-NPRL2 was constructed and identified by restriction enzyme digestion and sequencing. Liposome-mediated transfection into SGC-7901 gastric cancer cells, the fluorescence microscope, RT-PCR, Western blot detection of NPRL2 gene and protein levels, CCK8 assay of cell proliferation, flow cytometry analysis of cell cycle and Apoptosis rate changes, Transwell experiments to detect NPRL2 cell migration and invasion ability. Results: The eukaryotic expression vector pEGFP-N1-NPRL2 was successfully constructed. The expression of NPRL2 mRNA and protein was detected by RT-PCR and Westernblot after SGC-7901 cells were transfected with recombinant plasmids. The results of CCK8 showed that NPRL2 could significantly inhibit the proliferation of tumor cells (P <0.05) ). Cell cycle analysis showed that NPRL2 stopped cells at G0-G1 phase (P <0.05). The results of apoptosis showed that NPRL2 could inhibit cell apoptosis (P <0.05). Transwell invasion and migration assays showed that NPRL2 could promote cell invasion and migration (P <0.05). Conclusion: NPRL2 can inhibit tumor cell proliferation, cycle, invasion and migration, and promote apoptosis.