LPS Induces Occludin Dysregulation in Cerebral Microvascular Endothelial Cells via MAPK Signaling and Augmenting MMP-2 Levels.
LPS Induces Occludin Dysregulation in Cerebral Microvascular Endothelial Cells via MAPK Signaling and Augmenting MMP-2 Levels.
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LPS 通过 MAPK 信号传导和增强 MMP-2 水平诱导脑微血管内皮细胞中的 Occludin 失调
DOI：
10.1155/2015/120641
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发表时间：
2015
影响因子：
--
通讯作者：
Wu XH
中科院分区：
文献类型：
--
作者：
Qin LH;Huang W;Mo XA;Chen YL;Wu XH
在中枢神经系统感染期间，血脑屏障（BBB）完整性的破坏导致脑水肿。本研究探讨了脂多糖（LPS）诱导的紧密连接（TJ）蛋白表达失调的机制。将人脑微血管内皮细胞（hCMEC/D3）暴露于LPS、p38 MAPK抑制剂SB 203580和JNK抑制剂SP 600125，MTT法检测细胞活力。Western blot检测p38 MAPK、JNK和TJ（occludin和zonula occludens-（ZO-）1）蛋白的表达。采用实时定量聚合酶链反应（qRT-PCR）检测TJ组分和MMP-2的mRNA水平，采用酶联免疫吸附试验（ELISA）检测MMP-2的蛋白水平。LPS、SB 203580和SP 600125分别在10、7.69或0.22 µg/mL浓度下对细胞活力无影响。LPS处理降低了occludin和ZO-1的mRNA和蛋白水平，增强了p38 MAPK和JNK磷酸化以及MMP-2的表达。这些作用被SB 203580或SP 600125预处理减弱，但在ZO-1表达中没有减弱。盐酸多西环素（一种MMP-2抑制剂）和SB-3CT（一种特异性MMP-2抑制剂）均部分减弱LPS诱导的occludin下调。这些数据表明MMP-2过表达和p38 MAPK/JNK通路参与了LPS介导的hCMEC/D3中occludin的改变;然而，ZO-1水平不受p38 MAPK/JNK的影响。
Disrupted blood-brain barrier (BBB) integrity contributes to cerebral edema during central nervous system infection. The current study explored the mechanism of lipopolysaccharide- (LPS-) induced dysregulation of tight junction (TJ) proteins. Human cerebral microvascular endothelial cells (hCMEC/D3) were exposed to LPS, SB203580 (p38MAPK inhibitor), or SP600125 (JNK inhibitor), and cell vitality was determined by MTT assay. The proteins expressions of p38MAPK, JNK, and TJs (occludin and zonula occludens- (ZO-) 1) were determined by western blot. The mRNA levels of TJ components and MMP-2 were measured with quantitative real-time polymerase chain reaction (qRT-PCR), and MMP-2 protein levels were determined by enzyme-linked immunosorbent assay (ELISA). LPS, SB203580, and SP600125 under respective concentrations of 10, 7.69, or 0.22 µg/mL had no effects on cell vitality. Treatment with LPS decreased mRNA and protein levels of occludin and ZO-1 and enhanced p38MAPK and JNK phosphorylation and MMP-2 expression. These effects were attenuated by pretreatment with SB203580 or SP600125, but not in ZO-1 expression. Both doxycycline hyclate (a total MMP inhibitor) and SB-3CT (a specific MMP-2 inhibitor) partially attenuated the LPS-induced downregulation of occludin. These data suggest that MMP-2 overexpression and p38MAPK/JNK pathways are involved in the LPS-mediated alterations of occludin in hCMEC/D3; however, ZO-1 levels are not influenced by p38MAPK/JNK.