重组PinX1真核表达载体的构建及其对胃癌MKN28细胞增殖的抑制作用

Construction of recombinant eukaryotic expression vector of PinX1 and its roles in human gastric cancer MKN28 cell line

目的探讨PinX1基因对胃癌MKN28细胞生长及周期的作用,初步探讨该基因用于胃癌治疗方面的可能性。方法构建重组PinX1真核表达载体,酶切及测序鉴定无误后,应用脂质体转染法转染入胃癌MKN28细胞,建立稳定转染PinX1基因的MKN28细胞系;应用Western blot技术从蛋白水平检测转染前后目标蛋白PinX1的改变;MTT法检测转染前后细胞生长曲线的变化;流式细胞仪检测转染目的基因后细胞生长周期的改变。结果成功构建重组PinX1真核表达载体;建立了稳定转染入PinX1基因的胃癌MKN28细胞系;转染PinX1基因后,胃癌细胞细胞生长明显减缓(P<0.05),增殖变慢(P<0.05),细胞生长阻滞于G0/G1期。结论PinX1基因可抑制胃癌MKN28细胞的生长和增殖。

Objective To explore the effects of PinX1 gene on the growth and the cell cycles of gastric carcinoma cell line MKN28 and the possible functions of PinX1 gene in treatment of gastric cancer. Methods After construction of the recombinant eukaryotic expression vector of PinX1 confirmed by enzyme digestion, PinX1 gene was transfected into the gastric carcinoma cell line MKN28 by liposome method. The changes in PinX1, the cell growth, and the cell cycles were detected by Western blot, MTF, and flow eytometry, respectively. Results The recombinant eukaryotic expression vector of PinX1 was constructed successfully. Establishment of the stable transfection PinX1 into the gastric cancer MKN28 cell line was also successful. Transfection of PinX1 gene significantly inhibited the growth and proliferation of gastric cancer cells （ P 〈 0. 05 ）. The cell growth was arrested at G0/G1 stage. Conclusion As a potent telomerase inhibitor and a putative tumor suppressor, PinX1 gene may be used for the treatment of gastric cancer in clinical practice.