It is becoming increasingly clear that elevated blood cholesterol levels result in alterations in arterial wall function, quite apart from its role in the etiology of atherosclerosis. This is indicated by arterial hypersensitivity and reduced organ blood flows reported during dietary hypercholesterolemia. Excess cholesterol has been shown to alter membrane mediated functions in several cell types and in this way may contribute to altered cell function. Based on data from this and other laboratories, we have developed the hypothesis that elevated cholesterol levels result in the incorporation of excess cholesterol into membranes of the arterial wall cells, thereby altering membrane fluidity and transmembrane ion movements. We believe that these changes alter excitation-contraction coupling and contribute to the arterial hypersensitivity complicating hypercholesterolemia. The broad aim of this proposal is to determine the mechanism by which cholesterol alters cell membranes and contributes to abnormal cell function in arterial smooth muscle cells (SMC) and endothelial cells (EC). This proposal reflects the long-term goal of this laboratory which is in study the effects of altering membrane lipid composition on cell function in arterial SMC's and EC's.
The specific aims of this study are to determine: 1, the effects of excess cholesterol on the physical state (fluidity) of the plasmalemma membrane of SMC and EC, and 2, the effects of these membrane alteration on transmembrane calcium, potassium and sodium movements and function in smooth muscle cells, and 3, the effects of these membrane alterations on transmembrane calcium, potassium and sodium movements and function in endothelial cells. Two general and complimentary methodological approaches to this problem will be used. First, rabbit arteries will be perfused in vitro, and cholesterol enrichment of the cellular plasmalemma membranes of the vessel will be accomplished with cholesterol rich liposomes. Ring segments of these vessels will be used to study smooth muscle contraction in parallel with ion flux studies. Secondly, we will examine the effects of cholesterol enrichment on endothelial cell function (EDRF release) and transmembrane ion movement using primary cell cultures of rabbit endothelium. In addition, ion flux studies will also be performed in cultured rabbit SMC to define the details of transmembrane ion movements not otherwise discernable in intact vessel segments. The effects of cholesterol enrichment on the physical state (fluidity) of the membranes measured by fluorescence anisotropy will help define the molecular mechanism for cholesterol's effect. Integrating the results of these methodological approaches will permit a thorough and broad analysis of the molecular mechanisms by which cholesterol alters arterial reactivity. This study will provide new insights into the interplay between altered membrane cholesterol composition and cell function and will contribute to our understanding of the cellular mechanisms underlying inappropriate vasomotion associated with hypercholesterolemia.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL030496-05
Application #
3341520
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Project Start
1985-07-01
Project End
1991-06-30
Budget Start
1989-07-01
Budget End
1990-06-30
Support Year
5
Fiscal Year
1989
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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