Perillyl alcohol modulates activation, permeability and integrity of human brain endothelial cells induced by Plasmodium falciparum

BACKGROUND Cerebral malaria (CM) is a severe immunovasculopathy caused for Plasmodium falciparum infection, which is characterised by the sequestration of parasitised red blood cells (pRBCs) in brain microvessels. Previous studies have shown that some terpenes, such as perillyl alcohol (POH), exhibit a marked efficacy in preventing cerebrovascular inflammation, breakdown of the brain-blood barrier (BBB) and brain leucocyte accumulation in experimental CM models. OBJECTIVE To analyse the effects of POH on the endothelium using human brain endothelial cell (HBEC) monolayers co-cultured with pRBCs. METHODOLOGY The loss of tight junction proteins (TJPs) and features of endothelial activation, such as ICAM-1 and VCAM-1 expression were evaluated by quantitative immunofluorescence. Microvesicle (MV) release by HBEC upon stimulation by P. falciparum was evaluated by flow cytometry. Finally, the capacity of POH to revert P. falciparum-induced HBEC monolayer permeability was examined by monitoring trans-endothelial electrical resistance (TEER). FINDINGS POH significantly prevented pRBCs-induced endothelial adhesion molecule (ICAM-1, VCAM-1) upregulation and MV release by HBEC, improved their trans-endothelial resistance, and restored their distribution of TJPs such as VE-cadherin, Occludin, and JAM-A. CONCLUSIONS POH is a potent monoterpene that is efficient in preventing P. falciparum-pRBCs-induced changes in HBEC, namely their activation, increased permeability and alterations of integrity, all parameters of relevance to CM pathogenesis.

【背景】脑型疟疾（Cerebral malaria, CM）是由恶性疟原虫（Plasmodium falciparum）感染引发的重症免疫性血管病变，其特征为被寄生的红细胞（parasitised red blood cells, pRBCs）在脑微血管内发生黏附滞留。既往研究表明，部分萜类化合物如紫苏醇（perillyl alcohol, POH）在实验性脑型疟疾模型中，可显著抑制脑血管炎症、血脑屏障（blood-brain barrier, BBB）破坏以及脑内白细胞聚集。
【研究目标】通过构建与pRBCs共培养的人脑内皮细胞（human brain endothelial cell, HBEC）单层模型，分析POH对内皮细胞的作用效果。
【研究方法】采用定量免疫荧光技术，检测紧密连接蛋白（tight junction proteins, TJPs）的表达缺失以及内皮细胞激活特征（如细胞间黏附分子1（ICAM-1）、血管细胞黏附分子1（VCAM-1）的表达水平）。通过流式细胞术（flow cytometry），评估恶性疟原虫刺激下HBEC释放的微泡（microvesicle, MV）情况。最后，通过监测跨内皮电阻（trans-endothelial electrical resistance, TEER），探究POH逆转恶性疟原虫诱导的HBEC单层通透性升高的能力。
【研究结果】POH可显著抑制pRBCs诱导的内皮黏附分子（ICAM-1、VCAM-1）上调以及HBEC释放微泡，改善跨内皮电阻，并恢复VE-钙粘蛋白（VE-cadherin）、闭合蛋白（Occludin）和连接黏附分子A（JAM-A）等紧密连接蛋白的正常分布。
【研究结论】POH是一种高效单萜（monoterpene）类化合物，可有效抑制恶性疟原虫感染红细胞诱导的HBEC异常改变，包括内皮细胞激活、通透性升高以及完整性受损，上述均为与脑型疟疾发病机制相关的关键参数。