摘要
目的研究丝裂原活化蛋白激酶(MAPKs)活化对热打击致小鼠肺微血管内皮细胞(PMVECs)凋亡的影响。方法建立重症中暑小鼠模型,采用TUNEL染色及免疫组化检测肺组织损伤情况。二次磁珠分选法分离乳鼠PMVECs,TUNEL染色检测PMVECs凋亡情况,Western blotting检测热打击恢复期(0、2、6h)MAPKs家族活化情况。通过检测单层内皮细胞跨膜电阻(TEER)及辣根过氧化物酶(HRP)值观察不同热打击温度对单层细胞通透性的影响,同时使用MAPKs家族抑制剂检测热打击对单层细胞通透性及凋亡的影响。结果在重症中暑小鼠恢复期肺组织中可观察到PMVECs发生凋亡。TUNEL染色发现随着恢复期时间的延长,PMVECs凋亡数目增多,热打击可使PMVECs MAPKs家族活化且微血管通透性增加,给予p38活化抑制剂SB203580及ERK活化抑制剂PD98059预处理后细胞通透性增加,凋亡数目增多,而给予JNK抑制剂SP600125预处理后细胞则出现相反的变化。结论重症中暑小鼠PMVECs可发生凋亡,p38及ERK起着抗凋亡的作用,JNK起着促凋亡的作用。
Objective To investigate the effect ofmitogen-activated protein kinases （MAPKs） activation on the heat stress- induced apoptosis of pulmonary microvascular endothelial cells （PMVECs）. Methods A mouse model of severe heat stroke was made and TUNEL and immunohistochemistry were employed to detect lung tissue damage. MACS separation was used for isolation of neonatal PMVECs, and TUNEL was utilized to detect the apoptosis of PMVECs. Western blotting was used for determining the MAPKs activation during heat stress recovery （0, 2, 6h）. The monolayer permeability of endothelial cells was detected in terms of transmembrane resistance （TEER） and horseradish peroxidase （HRP）. Cells were pretreated with MAPKs activation inhibitors to examine the effect of heat stress on the monolayer cell permeability and apoptosis. Results In mice with severe heat stroke~ extensive apoptosis of PMVECs was found in their pulmonary tissues. TUNEL revealed that the number of apoptotic cells increased over time during heat stress recovery period and heat stress could activate MAPKs in PMVECs. Compared with heat stress group, in the cells pretreated with p38 or ERK activation inhibitor PD98059 and SB203580, the monolayer permeability and apoptosis increased while in cells pretreated withJNK inhibitor SP600125, the cellular permeability and apoptosis decreased. Conclusion In mice with severe heat stoke, PMVECs might experience apoptosis and p38 and ERK could inhibit apoptosis while JNK could promote apoptosis.
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2017年第4期279-284,共6页
Medical Journal of Chinese People's Liberation Army
基金
国家自然科学基金(81671896,81471839)
南方医科大学南方医院院长基金(2016C016)~~
关键词
中暑
肺微血管内皮细胞
丝裂原活性蛋白激酶类
细胞凋亡
细胞通透性
heat stroke
pulmonary microvascular endothelial cells
mitogen-activated protein kinases
apoptosis
cellpermeability
