Involvement of microtubules and Rho pathway in TGF-beta1-induced lung vascular barrier dysfunction.
Transforming growth factor-beta1 (TGF-beta1) is a cytokine critically involved in acute lung injury and endothelial cell (EC) barrier dysfunction. We have studied TGF-beta1-mediated signaling pathways and examined a role of microtubule (MT) dynamics in TGF-beta1-induced actin cytoskeletal remodeling and EC barrier dysfunction. TGF-beta1 (0.1-50 ng/ml) induced dose-dependent decrease in transendothelial electrical resistance (TER) in bovine pulmonary ECs, which was linked to increased actin stress fiber formation, myosin light chain (MLC) phosphorylation, EC retraction, and gap formation. Inhibitor of TGF-beta1 receptor kinase RI (5 microM) abolished TGF-beta1-induced TER decline, whereas inhibitor of caspase-3 zVAD (10 microM) was without effect. TGF-beta1-induced EC barrier dysfunction was linked to partial dissolution of peripheral MT meshwork and decreased levels of stable (acetylated) MT pool, whereas MT stabilization by taxol (5 microM) attenuated TGF-beta1-induced barrier dysfunction and actin remodeling. TGF-beta1 induced sustained activation of small GTPase Rho and its effector Rho-kinase; phosphorylation of myosin binding subunit of myosin specific phosphatase; MLC phosphorylation; EC contraction; and gap formation, which was abolished by inhibition of Rho and Rho-kinase, and by MT stabilization with taxol. Finally, elevation of intracellular cAMP induced by forskolin (50 microM) attenuated TGF-beta1-induced barrier dysfunction, MLC phosphorylation, and protected the MT peripheral network. These results suggest a novel role for MT dynamics in the TGF-beta1-mediated Rho regulation, EC barrier dysfunction, and actin remodeling.
Birukova
Anna A
AA
Division of Pulmonary and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21224, USA.
Birukov
Konstantin G
KG
Adyshev
Djanibek
D
Usatyuk
Peter
P
Natarajan
Viswanathan
V
Garcia
Joe G N
JG
Verin
Alexander D
AD
D000199:Actins / Q000378:metabolism; D000205:Actomyosin / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002199:Capillary Permeability / Q000187:drug effects; D002417:Cattle; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D042783:Endothelial Cells / Q000187:drug effects / Q000473:pathology; D047908:Intracellular Signaling Peptides and Proteins; D008168:Lung / Q000098:blood supply* / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008870:Microtubules / Q000187:drug effects* / Q000378:metabolism*; D018994:Myosin Light Chains / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D043263:Myosin-Light-Chain Phosphatase / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D053773:Transforming Growth Factor beta1; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D054460:rho-Associated Kinases
D000199:Actins; D047908:Intracellular Signaling Peptides and Proteins; D018994:Myosin Light Chains; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D000205:Actomyosin; D000242:Cyclic AMP; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D009219:Myosin-Light-Chain Kinase; D043263:Myosin-Light-Chain Phosphatase; D020741:rho GTP-Binding Proteins; D002118:Calcium