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该研究旨在探讨富含半胱氨酸的酸性分泌蛋白基因(secreted protein acidic and rich in cysteine gene,SPARC)过表达对卵巢癌淋巴结高转移细胞(SKOV3-PM4)生物学特性的影响。构建SPARC基因的慢病毒表达载体并转染SKOV3-PM4细胞,Real-time PCR和Western blot验证转染后的表达效率,激光共聚焦免疫荧光进行蛋白的细胞定位,细胞计数法和集落形成实验测定细胞增殖能力,流式细胞仪检测细胞周期,Transwell小室实验测定细胞体外侵袭、迁移能力。实验结果显示,SPARC蛋白存在于核周及胞质;过表达SPARC基因后,SKOV3-PM4细胞增殖受到明显抑制(P<0.05);细胞周期检测结果显示,各期改变无明显差异;体外侵袭、迁移实验结果显示,SKOV3-PM4细胞侵袭、迁移能力显著降低(P<0.05)。实验结果表明,SPARC基因在卵巢癌淋巴结转移中可能发挥抑癌基因的生物学作用。
The aim of this study was to investigate the effects of over-expression of cysteine-rich secreted protein acidic and rich in cysteine gene (SPARC) on the biological characteristics of lymph node metastatic cells (SKOV3-PM4) in ovarian cancer. The lentiviral expression vector of SPARC gene was constructed and transfected into SKOV3-PM4 cells. Real-time PCR and Western blot were used to verify the expression efficiency of the transfected SKOV3-PM4 cells. Laser confocal immunofluorescence was used to determine the cell localization, cell counting and colony formation assay Cell proliferation, flow cytometry cell cycle, Transwell chamber assay in vitro invasion and migration. The results showed that SPARC protein was present in the perinucleus and cytoplasm. The proliferation of SKOV3-PM4 cells was significantly inhibited (P <0.05) after overexpression of SPARC gene. The results of cell cycle assay showed that there was no significant difference between the two groups. Migration assay showed that invasion and migration of SKOV3-PM4 cells were significantly decreased (P <0.05). The experimental results show that SPARC gene may play a biological role of tumor suppressor gene in lymph node metastasis of ovarian cancer.