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Oxygen-Glucose Deprivation and Reoxygenation as an In Vitro Ischemia-Reperfusion Injury Model for Studying Blood-Brain Barrier Dysfunction
Published on: May 7, 2015
Association between adherens junctions and tight junctions via Rap1 promotes barrier protective effects of oxidized
Anna A Birukova1, Noureddine Zebda, Panfeng Fu
1Lung Injury Center, Section of Pulmonary and Critical Medicine, Department of Medicine, University of Chicago, Chicago, Illinois 60637, USA. kbirukov@medicine.bsd.uchicago.edu
Abstract:
Previous studies showed that cyclopenthenone-containing products resulting from oxidation of a natural phospholipid, 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (OxPAPC) exhibit potent barrier-protective effects in the in vitro and in vivo models of lung endothelial cell (EC) barrier dysfunction, and these effects are associated with enhancement of peripheral actin cytoskeleton, cell-cell and cell-substrate contacts driven by activation of Rac and Cdc42 GTPases. Rap1 GTPase is another member of small GTPase family involved in control of cell-cell interactions; however, its involvement in EC barrier-protective effects by OxPAPC remains unknown. This study examined a role of Rap1 in regulation of OxPAPC-induced interactions in adherens junctions (AJ) and tight junctions (TJ) as a novel mechanism of EC barrier preservation in vitro and in vivo. Immunofluorescence analysis, subcellular fractionation, and co-immunoprecipitation assays indicate that OxPAPC promoted accumulation of AJ proteins: VE-cadherin, p120-catenin, and β-catenin; and TJ proteins: ZO-1, occludin, and JAM-A in the cell membrane, and induced novel cross-interactions between AJ and TJ protein complexes, that were dependent on OxPAPC-induced Rap1 activation. Inhibition of Rap1 function suppressed OxPAPC-mediated pulmonary EC barrier enhancement and AJ and TJ interactions in vitro, as well as inhibited protective effects of OxPAPC against ventilator-induced lung injury in vivo. These results show for the first time a role of Rap1-mediated association between adherens junctions and tight junction complexes in the OxPAPC-induced pulmonary vascular EC barrier protection.
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