Fibrinogen induces alterations of endothelial cell tight junction proteins.
We previously showed that an elevated content of fibrinogen (Fg) increased formation of filamentous actin and enhanced endothelial layer permeability. In the present work we tested the hypothesis that Fg binding to endothelial cells (ECs) alters expression of actin-associated endothelial tight junction proteins (TJP). Rat cardiac microvascular ECs were grown in gold plated chambers of an electrical cell-substrate impedance system, 8-well chambered, or in 12-well plates. Confluent ECs were treated with Fg (2 or 4 mg/ml), Fg (4 mg/ml) with mitogen-activated protein kinase (MEK) kinase inhibitors (PD98059 or U0126), Fg (4 mg/ml) with anti-ICAM-1 antibody or BQ788 (endothelin type B receptor blocker), endothelin-1, endothelin-1 with BQ788, or medium alone for 24 h. Fg induced a dose-dependent decrease in EC junction integrity as determined by transendothelial electrical resistance (TEER). Western blot analysis and RT-PCR data showed that the higher dose of Fg decreased the contents of TJPs, occludin, zona occluden-1 (ZO-1), and zona occluden-2 (ZO-2) in ECs. Fg-induced decreases in contents of the TJPs were blocked by PD98059, U0126, or anti-ICAM-1 antibody. While BQ788 inhibited endothelin-1-induced decrease in TEER, it did not affect Fg-induced decrease in TEER. These data suggest that Fg increases EC layer permeability via the MEK kinase signaling pathway by affecting occludin, ZO-1, and ZO-2, TJPs, which are bound to actin filaments. Therefore, increased binding of Fg to its major EC receptor, ICAM-1, during cardiovascular diseases may increase microvascular permeability by altering the content and possibly subcellular localization of endothelial TJPs.
Patibandla
Phani K
PK
Department of Biochemistry and Molecular Biology, University of Louisville, School of Medicine, Louisville, Kentucky 40202, USA.
Tyagi
Neetu
N
Dean
William L
WL
Tyagi
Suresh C
SC
Roberts
Andrew M
AM
Lominadze
David
D
D000418:Albumins / Q000378:metabolism; D000818:Animals; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism*; D019332:Endothelin-1 / Q000494:pharmacology; D005340:Fibrinogen / Q000378:metabolism / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D062793:Occludin; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D019108:Tight Junctions / Q000187:drug effects* / Q000378:metabolism*; D014655:Vascular Resistance / Q000187:drug effects; D062826:Zonula Occludens-1 Protein; D062827:Zonula Occludens-2 Protein
D000418:Albumins; D019332:Endothelin-1; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; C568987:Ocln protein, rat; D010750:Phosphoproteins; D012333:RNA, Messenger; C569042:TJP1 protein, human; C569046:TJP2 protein, human; C569044:Tjp1 protein, rat; D062826:Zonula Occludens-1 Protein; D062827:Zonula Occludens-2 Protein; D005340:Fibrinogen