p38丝裂原活化蛋白激酶基因敲除对小鼠胚胎成纤维细胞增殖的影响

目的 研究p38丝裂原活化蛋白激酶(MAPK)基因敲除对小鼠胚胎成纤维细胞增殖的影响.方法 用蛋白质免疫印迹法(Western blotting)检测小鼠胚胎成纤维细胞中p38+/+和p38-/-的表达;利用噻唑蓝(MTT)比色法绘制p38+/+和p38-/-细胞的生长曲线,用流式细胞仪检测细胞周期各时相所占百分比.结果 绘制的生长曲线表明,p38-/-细胞的生长速率明显下降,增殖受到抑制,细胞倍增时间延长,培养96 h时,p38-/-细胞数量较p38+/+减少了15.5%,说明p38基因敲除明显抑制了G2/M的进程.流式细胞仪检测结果显示,p38+/+细胞中G0/G1期细胞占34.47%,G2/M期占10.81%,S期占54.72%;p38-/-细胞中G0/G1期占48.49%,G2/M期占4.06%,S期占47.44%.与p38+/+细胞相比,G1期的p38-/-细胞增加了40.7%,S期则减少了13.3%,说明p38基因敲除抑制了G1/S的进程.结论 p38基因敲除能够阻滞细胞周期进展,减慢细胞增殖速度.

Effect of p38 mitogen-activated protein kinase gene knockout on cell proliferation of embryonic fibroblasts in mice

OBJECTIVE: To investigate the effect of p38 mitogen-activated protein kinase (MAPK) gene knockout on the proliferation of embryonic fibroblasts in mice (MEFs). METHODS: The expression of p38 in MEFs p38(+/+)wwand p38(-/-)wwcells were detected by Western blotting. The growth curves of p38(+/+) and p38(-/-) cells were plotted with the results of methylthiazoletetrazolium (MTT) colorimetric assay, and the ratios of different cell phases of p38(+/+) and p38(-/-) cells were analyzed by flow cytometry. RESULTS: The growth curves showed that the growth rate was notably retarded and cell double time elongated in p38(-/-) cells, and there was 15.5% decrease of the number of p38(-/-) cells in comparison with that of p38(+/+) cells in 96-hour culture. G2/M transition was inhibited in p38(-/-) cells. Meanwhile, G1/S transition was also inhibited in p38(-/-) cells, as shown by the results of flow cytometry. The ratios of G0/G1, G2/M, and S phases of p38(+/+) cells were 34.47%, 10.81%, and 54.72%, respectively; while those of p38(-/-) cells were 48.49%, 4.06%, and 47.44%, respectively. There were 40.7% increase and 13.3% decrease in the cell numbers of G1 and S phases of p38(-/-) cells in comparison with those of p38(+/+) cells, respectively. CONCLUSION: p38 gene knockout in MEFs leads to cell cycle arrest and decreased cell proliferation.