摘要
目的建立犬肾细胞（Madin—Darby canine kidney，MDCK）缺氧模型，进一步探讨丝裂原活化蛋白激酶（MAPKs）在缺氧性细胞损伤中的活性改变。方法将MDCK细胞置于体积分数为5％CO：和95％N2的有机玻璃调节性密闭容器中分别培养24、36、48、72、84h，MTT实验检测细胞的存活力，蛋白免疫印迹法（Western blot）检测JNK（e—Jun NH2 terminal protein kinase）和p38MAPK的磷酸化活性。结果MDCK细胞缺氧24、36、48、72、84h后，与正常对照组相比，MTFD值均明显下降（P〈0．01）。MDCK细胞缺氧后，JNK和p38MAPK的磷酸化水平增高。结论此方法可以成功建立操作简便、有效的MDCK细胞缺氧模型，且是通过激活JNK、p38MAPK引起缺氧性细胞损伤。
Objective To establish a hypoxic model of Madin- Darby canine kidney （MDCK） cells and to investi- gate the changes of mitogen activated protein kinase （MAPKS） activity during hypoxic cell injury. Methods MDCK cells were incubated in a modular incubator chamber （5% CO2 and 95% N2 ） for 24 h, 36 h, 48 h, 72 h and 84 h~ Cell viability was measured by MTr assay. The phosphorylation of JNK and p38 MAPK was detected by Western blotting. Results Compared with the control, MDCK cells showed significantly decreases OD value in MTr assay after cultivated in hypoxic condition for 24 h, 36 h, 48 h, 72 h and 84 h （P 〈0.01 ）. Meanshile, the phosphorylation of JNK and p38MAPK was enhanced under hypoxic condition. Conclusion A simple and effective model of hypoxic MDCK cells was established by the present method, where hypoxia cell damages can be induced by activation of JNK and p38MAPK.
出处
《徐州医学院学报》
CAS
2016年第2期71-74,共4页
Acta Academiae Medicinae Xuzhou
基金
江苏省高校自然科学研究面上项目（12KJD320005）
徐州医学院“振兴计划”项目
