Increased lung vascular endothelial permeability leads to the leakage of fluid and plasma proteins into lung tissue, a primary factor in the pathogenesis of Acute Lung Injury (ALI). Our studies will investigate the fundamental question that Ca2+ entry mediated by transient receptor potential cation channel 6 (TRPC6), a receptor-operated channel (ROC), is a critical determinant of increased endothelial permeability. We will test the hypothesis that TRPC6 activation is required and sufficient for endothelial contraction that mediates increased lung vascular permeability. The proposed studies will use the agonist thrombin, which activates protease activating receptor (PAR-1) on the endothelial cell surface and produces diacylglycerol, the direct activator of the TRPC6 channel. Studies will determine the role of dual lipid/protein phosphatase PTEN in the mechanism of TRPC6 activation. We will address the role of TRPC6-mediated increase in intracellular Ca2+ as a requirement for signaling the activation of RhoA and MLCK-L, and thereby promoting endothelial cell contraction.
Our Specific Aims are to determine 1) role of endothelial ROC, TRPC6 in mediating Ca2+ signaling dependent increases in lung microvascular permeability, 2) PTEN-mediated mechanism of activation of TRPC6 and its consequences in the regulation of endothelial barrier function, and 3) downstream signaling mechanisms of TRPC6-induced increase in lung endothelial permeability involving the activation of PKCa and p60Src kinase and the effectors RhoA and MLCK-L isoform. These studies will be carried out at multiple levels including utilizing endothelial cells and lungs from mice carrying deletions in specific genes. Our hope is that the proposal will provide novel insights into the development of therapeutic strategies directed against TRPC6 that will prevent lung vascular "leakiness".

Public Health Relevance

The semi-selective endothelial cell monolayer partitions blood and tissue components. Increased permeability of this monolayer leads to leakage of fluid and macromolecules into lung tissue from the blood stream. If this leakage is not stopped it will lead to lung injury. Currently, there are no effective treatments for this problem. We believe that characterization of an important calcium channel in endothelial cells, TRPC6 will provide valuable insights to help us develop therapeutic strategies to reverse the vascular leakiness and thereby lung injury.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL071794-09
Application #
8386957
Study Section
Respiratory Integrative Biology and Translational Research Study Section (RIBT)
Program Officer
Peavy, Hannah H
Project Start
2002-12-01
Project End
2014-11-30
Budget Start
2012-12-01
Budget End
2014-11-30
Support Year
9
Fiscal Year
2013
Total Cost
$369,923
Indirect Cost
$134,303
Name
University of Illinois at Chicago
Department
Pharmacology
Type
Schools of Medicine
DUNS #
098987217
City
Chicago
State
IL
Country
United States
Zip Code
60612
Mehta, Dolly; Ravindran, Krishnan; Kuebler, Wolfgang M (2014) Novel regulators of endothelial barrier function. Am J Physiol Lung Cell Mol Physiol 307:L924-35
Rajput, Charu; Kini, Vidisha; Smith, Monica et al. (2013) Neural Wiskott-Aldrich syndrome protein (N-WASP)-mediated p120-catenin interaction with Arp2-Actin complex stabilizes endothelial adherens junctions. J Biol Chem 288:4241-50
Schmidt, Tracy Thennes; Tauseef, Mohammad; Yue, Lili et al. (2013) Conditional deletion of FAK in mice endothelium disrupts lung vascular barrier function due to destabilization of RhoA and Rac1 activities. Am J Physiol Lung Cell Mol Physiol 305:L291-300
Thennes, Tracy; Mehta, Dolly (2012) Heterotrimeric G proteins, focal adhesion kinase, and endothelial barrier function. Microvasc Res 83:31-44
Chava, Koteswara Rao; Tauseef, Mohammad; Sharma, Tiffany et al. (2012) Cyclic AMP response element-binding protein prevents endothelial permeability increase through transcriptional controlling p190RhoGAP expression. Blood 119:308-19
Chavez, Alejandra; Smith, Monica; Mehta, Dolly (2011) New insights into the regulation of vascular permeability. Int Rev Cell Mol Biol 290:205-48
Kini, Vidisha; Chavez, Alejandra; Mehta, Dolly (2010) A new role for PTEN in regulating transient receptor potential canonical channel 6-mediated Ca2+ entry, endothelial permeability, and angiogenesis. J Biol Chem 285:33082-91
Knezevic, Nebojsa; Tauseef, Mohammad; Thennes, Tracy et al. (2009) The G protein betagamma subunit mediates reannealing of adherens junctions to reverse endothelial permeability increase by thrombin. J Exp Med 206:2761-77
Tauseef, Mohammad; Kini, Vidisha; Knezevic, Nebojsa et al. (2008) Activation of sphingosine kinase-1 reverses the increase in lung vascular permeability through sphingosine-1-phosphate receptor signaling in endothelial cells. Circ Res 103:1164-72
Ramchandran, Ramaswamy; Mehta, Dolly; Vogel, Stephen M et al. (2008) Critical role of Cdc42 in mediating endothelial barrier protection in vivo. Am J Physiol Lung Cell Mol Physiol 295:L363-9

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