Prosthetic grafts are used widely in vascular reconstructive surgery, but their long-term patency is limited by their thrombogenicity and the development of intimal hyperplasia. Oxidized LDL and lysophosphatidylcholine (lysoPC), a product of LDL oxidation, accumulate in grafts and alter cell function. The long-term goal of our research is to improve the patency of vascular grafts by promoting endothelial cell (EC) healing of graft surfaces. LysoPC inhibits EC migration in vitro, and hypercholesterolemia reduces EC migration into injured arteries and onto grafts. Old and lysoPC increase cellular production of reactive oxygen species, increase cell membrane fluidity, and open ion channels. These effects can inhibit EC migration. Specifically, lysoPC activates a canonical transient receptor potential (TRPC) ion channel, TRPC6, which opens TRPC5 through a unique TRPC activation cascade, leading to a prolonged rise in intracellular free calcium ion concentration ([Ca2+]i). Increased [Ca2+]i inhibits EC migration by activation of calpains that breakdown cytoskeletal proteins essential for migration. This proposal addresses the hypothesis that lipid oxidation products formed within synthetic vascular grafts inhibit their EC migration, in part through activation of TRPC6 and TRPC5 channels, and thereby limit endothelialization of grafts in vivo. The goals of this project are to identify mechanisms by which lipid oxidation products activate TRPC6 and TRPC5 channels and identify ways to counteract this. To accomplish these goals, the mechanism by which lipid oxidation products activate TRPC6, specifically the roles of Src kinases and phospholipase C-31, will be explored. In addition, and the mechanism by which TRPC6 activates TRPC5 will be studied, focusing on the role of intracellular calcium and myosin light chain kinase. The role of reactive oxygen species and changes in membrane fluidity in these actions will also be explored. Finally, the ability of an apoA-I mimetic or HDL, which we have shown to block the TRPC6-TRPC5 activation cascade in vitro, to improve EC migration in areas of arterial injury in mice and onto prosthetic grafts implanted in normal and hypercholesterolemic rabbits will be assessed. The proposed studies will investigate a mechanism by which lipid oxidation products limit EC healing of vascular injuries and synthetic vascular grafts. Studies will also address the ability of HDL to promote EC healing. These studies will lead to a better understanding of the role of lipids in the pathophysiology of graft failure, and provide impetus for development of TRPC6 channel inhibitors or agents that interrupt the TRPC6- TRPC5 activation cascade. These mechanism-based therapies will promote endothelial healing of angioplasty sites and prosthetic grafts to prolong their patency for the benefit of all people undergoing cardiovascular interventions.

Public Health Relevance

The long-term goal of our research is improve the healing of bypass grafts or arteries after balloon angioplasty and stenting. We will investigate how oxidized lipids block the movement of endothelial cells (cells that normally line blood vessels) into an area of injury or onto a bypass graft. Specifically, we will study the role of certain ion channels (TRPC channels) that when eliminated abolish the inhibitory effect of high cholesterol in a mouse model. The results of these studies will provide direction in the development of treatments to promote endothelial cell healing after vascular interventions. For example, a TRPC6 inhibitor could increase endothelial cell migration and promote healing of arterial injuries and vascular grafts for the benefit all patients who require cardiovascular interventions.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Bioengineering, Technology and Surgical Sciences Study Section (BTSS)
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Lundberg, Martha
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Cleveland Clinic Lerner
Other Basic Sciences
Schools of Medicine
United States
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Chaudhuri, Pinaki; Rosenbaum, Michael A; Birnbaumer, Lutz et al. (2017) Integration of TRPC6 and NADPH oxidase activation in lysophosphatidylcholine-induced TRPC5 externalization. Am J Physiol Cell Physiol 313:C541-C555
Chaudhuri, Pinaki; Rosenbaum, Michael A; Sinharoy, Pritam et al. (2016) Membrane translocation of TRPC6 channels and endothelial migration are regulated by calmodulin and PI3 kinase activation. Proc Natl Acad Sci U S A 113:2110-5
Rosenbaum, Michael A; Chaudhuri, Pinaki; Graham, Linda M (2015) Hypercholesterolemia inhibits re-endothelialization of arterial injuries by TRPC channel activation. J Vasc Surg 62:1040-1047.e2
Rosenbaum, Michael A; Chaudhuri, Pinaki; Abelson, Benjamin et al. (2015) Apolipoprotein A-I mimetic peptide reverses impaired arterial healing after injury by reducing oxidative stress. Atherosclerosis 241:709-15
Rosenbaum, Michael A; Miyazaki, Keiko; Graham, Linda M (2012) Hypercholesterolemia and oxidative stress inhibit endothelial cell healing after arterial injury. J Vasc Surg 55:489-96
Rosenbaum, Michael A; Miyazaki, Keiko; Colles, Scott M et al. (2010) Antioxidant therapy reverses impaired graft healing in hypercholesterolemic rabbits. J Vasc Surg 51:184-93
Miyazaki, Keiko; Colles, Scott M; Graham, Linda M (2008) Impaired graft healing due to hypercholesterolemia is prevented by dietary supplementation with alpha-tocopherol. J Vasc Surg 48:986-93
Chaudhuri, Pinaki; Colles, Scott M; Bhat, Manjunatha et al. (2008) Elucidation of a TRPC6-TRPC5 channel cascade that restricts endothelial cell movement. Mol Biol Cell 19:3203-11
Patel, Rajendra; Cardneau, Jeffry D; Colles, Scott M et al. (2006) Synthetic smooth muscle cell phenotype is associated with increased nicotinamide adenine dinucleotide phosphate oxidase activity: effect on collagen secretion. J Vasc Surg 43:364-71
Chaudhuri, Pinaki; Colles, Scott M; Fox, Paul L et al. (2005) Protein kinase Cdelta-dependent phosphorylation of syndecan-4 regulates cell migration. Circ Res 97:674-81

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