PKC-dependent, burn-induced adherens junction reorganization and barrier dysfunction in pulmonary microvascular endothelial cells.
Rat lung microvascular endothelial cell monolayers were exposed to donor plasma from burned rats (25% total body surface area) at 1:3 dilution for 30 min. Immunofluorescence analysis revealed that concomitant with gap formation alterations were seen in the adherens junction (AJ) proteins beta-catenin and vascular endothelial-cadherin. Both of these components were shown to exist in a smooth, uniform arrangement at the cell periphery in untreated cells. However, upon exposure to burn plasma, this uniformity was lost, and the AJ proteins showed a disrupted, zipper-like pattern at the cells' edge. In addition, these proteins were absent from areas of gap formation between the cells, and an increase in punctate staining throughout the cells suggests they were internalized in response to burn plasma. Measurements of both transendothelial electrical resistance and FITC-albumin flux across the cell monolayer were used to assess barrier integrity. Our study found that exposure to burn plasma rapidly caused the electrical resistance across confluent monolayers to decrease and albumin flux to increase, phenomena associated with barrier dysfunction. Furthermore, all the above responses to burn plasma were attenuated when cells were pretreated with the PKC inhibitor bisindolylmaleimide, suggesting that PKC is required for burn-induced pulmonary endothelial dysfunction.
Tinsley
John H
JH
Dept. of Surgery, Scott and White Memorial Hospital, 702 SW HK Dodgen Loop, Temple, TX 76504, USA. jtinsley@swmail.sw.org
Breslin
Jerome W
JW
Teasdale
Nicole R
NR
Yuan
Sarah Y
SY
D022005:Adherens Junctions / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002056:Burns / Q000097:blood*; D015820:Cadherins / Q000378:metabolism; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000378:metabolism; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000378:metabolism / Q000473:pathology*; D007211:Indoles / Q000494:pharmacology; D008168:Lung / Q000098:blood supply*; D008301:Maleimides / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015534:Trans-Activators / Q000378:metabolism; D051176:beta Catenin
D015820:Cadherins; C495268:Ctnnb1 protein, rat; D003598:Cytoskeletal Proteins; D007211:Indoles; D008301:Maleimides; D015534:Trans-Activators; D051176:beta Catenin; D011493:Protein Kinase C; C088060:bisindolylmaleimide