Microvascular leakage has been implicated in the pathogenesis of multiple organ dysfunctions in inflammation, sepsis, and ischemic reperfusion injury. The abnormality is largely attributable to neutrophil (PMN) activation and releasing edemagenic agents. The overall goal of this research project is to define the precise molecular mechanisms of PMN adhesion-dependent and adhesion-independent microvascular barrier injury. Our studies during the prior funding cycle have revealed a series of coordinated signaling and structural reactions at the microvascular endothelium during PMN stimulation, characterized by focal adhesion-supported cytoskeleton contraction and junction disorganization. Comparative analysis of the complex pathways points to a predominant role for myosin light chain (MLC)-mediated contractile force development as a triggering event to open the paracellular pathway for protein leakage across the endothelium. As a continuing effort, this study is designed to further characterize the endothelial contractile mechanism by testing the hypothesis that PMNs induce actin polymerization and MLC phosphorylation via p60Src-activation of endothelial-specific myosin light chain kinase (eMLCK) and RhoA-inhibition of the myosin targeting subunit of myosin light chain phosphatase (MLCP).
Three specific aims are proposed: 1) to study the eMLCK mechanism of PMN-elicited microvascular leakage, 2) to study the MLCP regulation of microvascular barrier function during PMN activation, and 3) to characterize the mechanical and molecular basis of endothelial contractile response in PMN-induced hyperpermeability. The research plan incorporates genetic, molecular, and pharmacological approaches into an integrative analysis of molecular reactions and barrier function at both the microvascular and subendothelial levels. Intravital microscopic studies will be complemented by in situ and in vitro experiments for a comprehensive evaluation of hydraulic conductivity and solute permeability. The study will provide novel mechanistic insights into the regulation of microvascular endothelial barrier function by PMNs, with broad implications in various types of vascular diseases or injuries. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL061507-10
Application #
7393792
Study Section
Hypertension and Microcirculation Study Section (HM)
Program Officer
Goldman, Stephen
Project Start
1999-01-01
Project End
2012-03-31
Budget Start
2008-04-01
Budget End
2009-03-31
Support Year
10
Fiscal Year
2008
Total Cost
$342,000
Indirect Cost
Name
University of California Davis
Department
Surgery
Type
Schools of Medicine
DUNS #
047120084
City
Davis
State
CA
Country
United States
Zip Code
95618
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Sun, Chongxiu; Beard Jr, Richard S; McLean, Danielle L et al. (2013) ADAM15 deficiency attenuates pulmonary hyperpermeability and acute lung injury in lipopolysaccharide-treated mice. Am J Physiol Lung Cell Mol Physiol 304:L135-42
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Lee, Eugene S; Shen, Qiang; Pitts, Robert L et al. (2011) Serum metalloproteinases MMP-2, MMP-9, and metalloproteinase tissue inhibitors in patients are associated with arteriovenous fistula maturation. J Vasc Surg 54:454-9; discussion 459-60
Sun, Chongxiu; Wu, Mack H; Yuan, Sarah Y (2011) Nonmuscle myosin light-chain kinase deficiency attenuates atherosclerosis in apolipoprotein E-deficient mice via reduced endothelial barrier dysfunction and monocyte migration. Circulation 124:48-57

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