Wilson D W, Lamé M W, Dunston S K, Taylor D W, Segall H J
Departments of Veterinary Pathology, Microbiology, and Immunology, University of California at Davis, Davis, California 95616, USA.
Toxicol Appl Pharmacol. 1998 Sep;152(1):138-44. doi: 10.1006/taap.1998.8488.
One of the earliest morphologic changes evident in the monocrotaline (MCT) model of pulmonary hypertension in rats is microvascular leak. Whether this represents a direct effect of MCT metabolites or is secondary to inflammatory and thrombotic changes remains uncertain. To determine whether MCT directly affects endothelial cell permeability barrier function, we characterized the interaction of the reactive pyrrole intermediate of MCT (MCTP) with endothelial cell actin and characterized its effects on thrombin-mediated signal transduction and monolayer permeability. Bovine pulmonary endothelial cells (BPAEC) treated with MCTP had altered distribution of filamentous actin evident by fluorescence microscopy. Correlative Western blots and autoradiography of actin isolated from BPAEC treated with 14C-MCTP showed comigration of actin and MCTP-derived 14C. MCTP treatment did not alter cellular free Ca2+ concentrations nor did it interfere with thrombin-mediated intracellular Ca2+ signal. Pretreatment with MCTP significantly augmented the thrombin-mediated transudation of Evan's blue albumin in BPAEC monolayers apparently by increasing the size of intercellular gaps. We conclude that MCTP directly interacts with actin to alter its polymerization state but does not significantly affect endothelial cell response to contractile stimulus. Our results suggest that MCTP may affect endothelial cell barrier function through alterations in intracellular junctions.
大鼠肺动脉高压的野百合碱（MCT）模型中最早出现的明显形态学变化之一是微血管渗漏。这是MCT代谢产物的直接作用，还是继发于炎症和血栓形成变化，仍不确定。为了确定MCT是否直接影响内皮细胞通透性屏障功能，我们对MCT的活性吡咯中间体（MCTP）与内皮细胞肌动蛋白的相互作用进行了表征，并表征了其对凝血酶介导的信号转导和单层通透性的影响。用MCTP处理的牛肺动脉内皮细胞（BPAEC）通过荧光显微镜观察到丝状肌动蛋白的分布发生改变。从用14C-MCTP处理的BPAEC中分离出的肌动蛋白的相关蛋白质免疫印迹和放射自显影显示肌动蛋白和MCTP衍生的14C共迁移。MCTP处理未改变细胞内游离Ca2+浓度，也未干扰凝血酶介导的细胞内Ca2+信号。用MCTP预处理明显增加了BPAEC单层中凝血酶介导的伊文思蓝白蛋白渗出，显然是通过增加细胞间隙的大小。我们得出结论，MCTP直接与肌动蛋白相互作用以改变其聚合状态，但不会显著影响内皮细胞对收缩刺激的反应。我们的结果表明，MCTP可能通过改变细胞内连接来影响内皮细胞屏障功能。
