缺氧环境对PC12细胞损伤及HIF-1 mRNA表达水平的影响

Anoxic injury and HIF-1 mRNA expression changes after hypoxia in PC12 cell

目的探讨缺氧环境对PC12细胞的损伤作用及其初步机制。方法建立PC12细胞缺氧模型;倒置相差显微镜观察细胞形态;MTT法检测正常及缺氧环境对PC12细胞的增殖;PC12细胞缺氧培养0,2,4,8,12,24和48 h后测定乳酸脱氢酶(lactate dehydrogenase,LDH);用Real-time RT-PCR检测HIF-1 mRNA的表达水平。结果缺氧培养24 h,48 h后PC12细胞增殖受到抑制(P<0.05,P<0.01);PC12细胞缺氧培养2、4、8 h后培养液中的LDH活性从2～8 h逐渐增高,8 h达到最高值(P<0.01),8 h后逐渐下降;PC12缺氧培养0～4 h HIF-1 mRNA的表达水平逐渐增高,4 h达到最高,4～48 h逐渐降低。结论缺氧环境可引起PC12细胞损伤,缺氧8 h损伤最严重。缺氧导致HIF-1基因表达水平提高,4 h时达到峰值。

Aim To investigate the effect of the anoxic injury and HIF-1 mRNA expression changes after hypoxia in PC12 cell.Methods A hypoxia model was established;The inverted contrast phase microscope was used to observe cell morphology;MTT assay was used to determine the effect of normal and hypoxic environment on PC12 cell proliferation;Anaerobic incubation of PC12 cell lasted for 0,2,4,8,12,24 and 48 h,before lactate dehydrogenase（LDH） activity was measured;Real-time RT-PCR detected the expression of HIF-1 mRNA.Results Anaerobic cultivation after 24 h,the proliferation of PC12 cells was inhibited（P0.05）,hypoxia after 48 h the proliferation of PC12 cells was inhibited significantly（P0.01）;PC12 cells hypoxia 2,4,8,12,24 and 48 h,the cell culture medium LDH activity increased in a time-dependant manner within the 2～8 h range,and gradually decreases after the 8 h peak（P0.01）;the expression of HIF-1 mRNA level grodually increased within the 0～4 h range,and dropped after the 4 h peak.Conclusion Hypoxia environmental can cause PC12 cell damage,hypoxia 8 h damaged membrane most seriously,hypoxia 4 h the HIF-1 mRNA expression reached the highest level.