Continuous Blood Purification Ameliorates Endothelial Hyperpermeability in SAP Patients with MODS by Regulating Tight Junction Proteins via ROCK
Continuous Blood Purification Ameliorates Endothelial Hyperpermeability in SAP Patients with MODS by Regulating Tight Junction Proteins via ROCK
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DOI：
10.5301/ijao.5000216
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发表时间：
2013-08
期刊：
影响因子：
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通讯作者：
Mei Yang;Xue-mei Chen;Xiao-Gang Du;F. Cao;Sicharam Vijaya Luxmi;Qing Shen
中科院分区：
文献类型：
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作者：
Mei Yang;Xue-mei Chen;Xiao-Gang Du;F. Cao;Sicharam Vijaya Luxmi;Qing Shen
背景：严重急性胰腺炎（SAP）期间炎症介质级联过度激活是多器官功能障碍的主要原因，并与高死亡率相关。近年来，越来越多的研究表明，持续血液净化（continuous blood purification， CBP）可改善多器官功能障碍综合征（MODS）患者的预后，但其确切机制尚不清楚。许多研究者发现紧密连接屏障的破坏是内皮细胞高通透性的重要因素，在MODS的发病机制中起着关键作用。之前，我们发现CBP可以通过调节RhoA/ROCK介导的细胞骨架重组来减轻SAP肺损伤患者内皮细胞的高通透性。然而，CBP对SAP合并MODS患者紧密连接蛋白变化的影响尚不清楚。本研究旨在通过体外模型探讨紧密连接在SAP合并MODS患者内皮细胞高通透性中的作用，以及CBP对紧密连接屏障的影响。方法分别于CBP前和CBP后采血，观察肝肾功能、动脉血气，计算APACHE II评分，评价患者的严重程度。为了检测RhoA/ROCK信号通路是否参与其中，在CBP期间的特定时间点将人脐静脉内皮细胞（HUVECs）暴露于患者的血清样本中，或者用ROCK抑制剂Y-27632预孵育，然后用血清处理。然后观察内皮细胞通透性的变化以及紧密连接蛋白occludin和claudin-1的表达和分布。结果与CBP前比较，CBP后APACHEⅱ评分、血清肌酐、丙氨酸转氨酶均显著降低，PaO2/FiO2显著升高。同时，患者血清诱导的内皮通透性明显增加，紧密连接蛋白occludin和claudin-1的表达明显降低，occludin和claudin-1在这些细胞中出现严重的破坏。然而，用rho激酶抑制剂Y-27632预处理可以减轻所有这些异常，并且以剂量依赖的方式。用CBP治疗后患者血清处理HUVECs后，内皮细胞的高通透性、occludin和claudin-1的异常表达和分布减弱。结论ROCK介导的紧密连接异常是SAP合并MODS患者血清内皮细胞高通透性的重要机制。CBP可以改善紧密连接蛋白的紊乱和重分布，从而改善内皮细胞的通透性。
Background Excessive activation of inflammatory mediator cascade during severe acute pancreatitis (SAP) is a major cause of multiple organ dysfunction and is associated with a high mortality. Recently, more and more studies have shown that continuous blood purification (CBP) could improve the prognosis of patients with multiple organ dysfunction syndrome (MODS), but the exact mechanism is still unclear. Many researchers have found that the disruption of tight junction barrier was an important factor for endothelial hyperpermeability, which played a key role in the pathogenesis of MODS. Previously, we found CBP could attenuate endothelial hyperpermeability in SAP patients with lung injury through regulating cytoskeleton reorganization mediated by RhoA/ROCK. However, the effect of CBP on the change of tight junction proteins in SAP patients with MODS was still unknown. This study aimed to investigate the role of tight junctions in endothelial hyperpermeability in SAP patients with MODS using an in vitro model, and the effect of CBP on tight junction barrier. Methods Before CBP and after CBP, blood samples were collected to observe hepatic and renal function, and arterial blood gas, while the APACHE II score was calculated to evaluate the severity of patients. To test whether RhoA/ROCK signaling pathway was involved, human umbilical vein endothelial cells (HUVECs) were exposed to serum samples taken from patients at specific time points during CBP, or preincubated with ROCK inhibitor, Y-27632, followed by treatment with serum. Then, the changes in endothelial cell permeability and the expression and distribution of tight junction proteins occludin and claudin-1 were observed. Results Compared with before CBP, the APACHE II score, serum creatinine and alanine aminotransferase decreased significantly, while PaO2/FiO2 increased significantly after CBP. Meanwhile, endothelial permeability induced by serum from patients significantly increased, while the expression of tight junction proteins occludin and claudin-1 significantly decreased, and severe disruption of occludin and claudin-1 was found in these cells. However, pretreated with Rho-kinase inhibitor, Y-27632 could lessen all of these abnormalities, and in a dose-dependent way. Endothelial hyperpermeability, the abnormal expression and distribution of occludin and claudin-1 were attenuated in HUVECs treated with serum from patients after CBP treatment. Conclusions The abnormality of tight junctions mediated by ROCK was an important mechanism for endothelial hyperpermeability induced by serum from SAP patients with MODS. CBP could ameliorate the disorganization and redistribution of tight junction proteins, hence improve the endothelial permeability.