The hypothesis of the proposed studies is that cholesterol enrichment alters the structure and function of the plasma membrane which, in turn, alters cation fluxes and cytoplasmic ion concentrations, thereby triggering altered vasomotion and cell proliferation. The studies will combine studies in isolated arteries from normal and cholesterol-fed rabbit aorta, isolated cells from the arteries, and cultured smooth muscle and endothelium.
Aim 1 will determine the effects of dietary atherosclerosis on basal and agonist activated transmembrane Ca++, K+, and Na++ movements and free cytosolic levels. Ion fluxes will be determined in isolated arteries, cytosolic calcium levels will be determined in dispersed cells with fura-2, cytosolic K+ and Na+ levels will be estimated from back extrapolation to zero time from the ion efflux data. Changes in the ions will be correlated to vasoactive responses to agonists and potassium channel activators. Comparisons will be made in cultured cells where purer membrane preps will be available. In addition, changes in microvascular responses to ischemia as well as to K+ activators in the Langendorff rabbit heart as well as blood pressure responses in the anesthetized rabbit will be studied.
Aim 2 will measure membrane lipids, fluidity will be assessed, and a collaborator will assess membrane structure by x-ray diffraction. These parameters will be correlated with the ion flux data in aim 1.
Aim 3 will examine the potential role of LDL obtained form normal and hypercholesterolemic subjects (as well as modified LDL) for its ability to cause similar changes in ion fluxes and membrane structure.
Aim 4 will determine the mechanisms responsible for the increased proliferation of vascular cells by determining the potency of calcium channel antagonists and performing parallel ion flux measurements, as well as determining the influence of conditioned media from cells enriched with cholesterol.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL030496-08
Application #
3341522
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Project Start
1985-07-01
Project End
1995-06-30
Budget Start
1992-07-10
Budget End
1993-06-30
Support Year
8
Fiscal Year
1992
Total Cost
Indirect Cost
Name
Allegheny University of Health Sciences
Department
Type
Schools of Medicine
DUNS #
City
Philadelphia
State
PA
Country
United States
Zip Code
19129
Levitan, I; Christian, A E; Tulenko, T N et al. (2000) Membrane cholesterol content modulates activation of volume-regulated anion current in bovine endothelial cells. J Gen Physiol 115:405-16
Laury-Kleintop, L D; Gleason, M; Tulenko, T N (1999) Expression of the heterogenous nuclear ribonucleoprotein complex K protein and the prolyl-4-hydroxylase alpha-subunit in atherosclerotic arterial smooth muscle cells. Biochem Biophys Res Commun 260:382-9
Golden, G A; Mason, R P; Tulenko, T N et al. (1999) Rapid and opposite effects of cortisol and estradiol on human erythrocyte Na+,K+-ATPase activity: relationship to steroid intercalation into the cell membrane. Life Sci 65:1247-55
Tulenko, T N; Chen, M; Mason, P E et al. (1998) Physical effects of cholesterol on arterial smooth muscle membranes: evidence of immiscible cholesterol domains and alterations in bilayer width during atherogenesis. J Lipid Res 39:947-56
Cox, R H; Zhou, Z; Tulenko, T N (1998) Voltage-gated sodium channels in human aortic smooth muscle cells. J Vasc Res 35:310-7
Tulenko, T N; Laury-Kleintop, L; Walter, M F et al. (1997) Cholesterol, calcium and atherosclerosis: is there a role for calcium channel blockers in atheroprotection? Int J Cardiol 62 Suppl 2:S55-66
Cox, R H; Tulenko, T N (1995) Altered contractile and ion channel function in rabbit portal vein with dietary atherosclerosis. Am J Physiol 268:H2522-30
Chen, M; Mason, R P; Tulenko, T N (1995) Atherosclerosis alters the composition, structure and function of arterial smooth muscle cell plasma membranes. Biochim Biophys Acta 1272:101-12
Izzo Jr, N J; Tulenko, T N; Colucci, W S (1994) Phorbol esters and norepinephrine destabilize alpha 1B-adrenergic receptor mRNA in vascular smooth muscle cells. J Biol Chem 269:1705-10
Stepp, D W; Tulenko, T N (1994) Alterations in basal and serotonin-stimulated calcium permeability and vasoconstriction in atherosclerotic aorta. Arterioscler Thromb 14:1854-9

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