title: The leading role of microtubules in endothelial barrier dysfunction: disassembly of peripheral microtubules leaves behind the cytoskeletal reorganization.
abstract: Disturbance of the endothelial barrier is characterized by dramatic cytoskeleton reorganization, activation of actomyosin contraction and, finally, leads to intercellular gap formation. Here we demonstrate that the edemagenic agent, thrombin, causes a rapid increase in the human pulmonary artery endothelial cell (EC) barrier permeability accompanied by fast decreasing in the peripheral microtubules quantity and reorganization of the microtubule system in the internal cytoplasm of the EC within 5 min of the treatment. The actin stress-fibers formation occurs gradually and the maximal effect is observed relatively later, 30 min of the thrombin treatment. Thus, microtubules reaction develops faster than the reorganization of the actin filaments system responsible for the subsequent changes of the cell shape during barrier dysfunction development. Direct microtubules depolymerization by nocodazole initiates the cascade of barrier dysfunction reactions. Nocodazole-induced barrier disruption is connected directly with the degree of peripheral microtubules depolymerization. Short-term loss of endothelial barrier function occurs at the minimal destruction of peripheral microtubules, when actin filament system is still intact. Specifically, we demonstrate that the EC microtubule dynamics examined by time-lapse imaging of EB3-GFP comets movement has changed under these conditions: microtubule plus ends growth rate significantly decreased near the cell periphery. The microtubules, apparently, are the first target in the circuit of reactions leading to the pulmonary EC barrier compromise. Our results show that dynamic microtubules play an essential role in the barrier function in vitro; peripheral microtubules depolymerization is necessary and sufficient condition for initiation of endothelial barrier dysfunction.
authors.affiliation: Electron Microscopy Department, AN Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia. irina_alieva@belozersky.msu.ru
authors.forename: Irina B
authors.initials: IB
authors.lastname: Alieva
authors.forename: Evgeny A
authors.initials: EA
authors.lastname: Zemskov
authors.forename: Ksenija M
authors.initials: KM
authors.lastname: Smurova
authors.forename: Irina N
authors.initials: IN
authors.lastname: Kaverina
authors.forename: Alexander D
authors.initials: AD
authors.lastname: Verin
mesh_terms: D003599:Cytoskeleton / Q000378:metabolism*; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D008870:Microtubules / Q000378:metabolism*; D011651:Pulmonary Artery / Q000166:cytology*; D013917:Thrombin / Q000378:metabolism
chemical_list: D013917:Thrombin
keywords: ENDOTHELIAL BARRIER DYSFUNCTION; ENDOTHELIAL BARRIER FUNCTION; HUMAN PULMONARY ENDOTHELIUM; MICROTUBULE DYNAMICS; MICROTUBULES