title: Changes in endothelial connexin 43 expression inversely correlate with microvessel permeability and VE-cadherin expression in endotoxin-challenged lungs.
abstract: Endothelial barrier restoration reverses microvessel hyperpermeability and facilitates recovery from lung injury. Because inhibiting connexin 43 (Cx43)-dependent interendothelial communication blunts hyperpermeability in single microvessels, we determined whether endothelial Cx43 levels correlate with changes in microvessel permeability during recovery from lung injury. Toward this, bacterial endotoxin was instilled intratracheally into rat lungs, and at different durations postinstillation the lungs were isolated and blood perfused. Microvessel Cx43 expression was quantified by in situ immunofluorescence and microvessel permeability via a fluorescence method. To supplement the immunofluorescence data, protein levels were determined by immunoblots of lung tissue from endotoxin-instilled rats. Immunofluorescence and immunoblot together revealed that both Cx43 expression and microvessel permeability increased above baseline within a few hours after endotoxin instillation but declined progressively over the next few days. On day 5 postendotoxin, microvessel Cx43 declined to negligible levels, resulting in complete absence of intermicrovessel communication determined by photolytic uncaging of Ca(2+). However, by day 14, both Cx43 expression and microvessel permeability returned to baseline levels. In contrast to Cx43, expression of microvessel vascular endothelial (VE)-cadherin, a critical determinant of vascular barrier integrity, exhibited an inverse trend by initially declining below baseline and then returning to baseline at a longer duration. Knockdown of vascular Cx43 by tail vein injection of Cx43 shRNA increased VE-cadherin expression, suggesting that reduction in Cx43 levels may modulate VE-cadherin levels in lung microvessels. Together, the data suggest that endotoxin challenge initiates interrelated changes in microvessel Cx43, VE-cadherin, and microvessel permeability, with changes in Cx43 temporally leading the other responses.
authors.affiliation: Department of Physiology and.
authors.forename: Kathirvel
authors.initials: K
authors.lastname: Kandasamy
authors.affiliation: Department of Physiology and.
authors.forename: Rachel
authors.initials: R
authors.lastname: Escue
authors.affiliation: Department of Physiology and.
authors.forename: Jayeeta
authors.initials: J
authors.lastname: Manna
authors.affiliation: Department of Physiology and.
authors.forename: Adebowale
authors.initials: A
authors.lastname: Adebiyi
authors.affiliation: Department of Physiology and Department of Orthopedic Surgery and Biomedical Engineering, University of Tennessee Health Science Center, Memphis, Tennessee kparthas@uthsc.edu.
authors.forename: Kaushik
authors.initials: K
authors.lastname: Parthasarathi
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D015820:Cadherins / Q000378:metabolism*; D002199:Capillary Permeability; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D054426:Chemokine CXCL2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008168:Lung / Q000098:blood supply / Q000254:growth & development / Q000276:immunology* / Q000378:metabolism; D008810:Mice, Inbred C57BL; D055806:Microvessels / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D018919:Neovascularization, Physiologic / Q000276:immunology; D050794:STAT1 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction; D000099152:Cadherin 5
chemical_list: D015703:Antigens, CD; D015820:Cadherins; D054426:Chemokine CXCL2; D018031:Connexin 43; D008070:Lipopolysaccharides; D016328:NF-kappa B; D050794:STAT1 Transcription Factor; D000099152:Cadherin 5; C404915:Cxcl2 protein, mouse; C543076:GJA1 protein, mouse; C494080:Stat1 protein, mouse
keywords: Ca2+ communication; connexin 43; immunofluorescence; rat; short-hairpin RNA; uncaging; vascular endothelial cadherin