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(Arteriosclerosis, Thrombosis, and Vascular Biology. 1997;17:394-402.)
© 1997 American Heart Association, Inc.

Articles

Reduced Responsiveness of Hypercholesterolemic Rabbit Aortic Smooth Muscle Cells to Nitric Oxide

Robert M. Weisbrod; Mark C. Griswold; Yue Du; Victoria M. Bolotina; Richard A. Cohen

the Vascular Biology Unit, Robert Dawson Evans Department of Clinical Research, Department of Medicine, Boston (Mass) University Medical Center.

Correspondence to Richard A. Cohen, MD, Vascular Biology Unit, R408, Boston University Medical Center, 80 E Concord St, Boston, MA 02118. E-mail racohen@med-med1.bu.edu.

The response to nitric oxide of intracellular free Ca²⁺ levels, measured by fura 2 fluorimetry, and cyclic GMP, measured by RIA, was evaluated on smooth muscle cells of the thoracic aorta in primary culture from normal and cholesterol-fed rabbits. Relaxation to acetylcholine and nitric oxide was also determined in isolated rings of aorta. After 10 weeks of high-cholesterol diet, the intact aorta relaxed less to both acetylcholine and nitric oxide. In cultured cells from hypercholesterolemic rabbits, intracellular Ca²⁺ oscillated, and the mean Ca²⁺ levels were approximately twofold greater than in normal aortic cells. Nitric oxide failed to affect basal Ca²⁺ in either cell type. The peak and sustained rise in intracellular Ca²⁺ induced by angiotensin II (10^-7 mol/L) were similar in the two cell types. However, nitric oxide (10^-10 to 10^-6 mol/L) decreased the sustained Ca²⁺ levels to a significantly smaller extent in cells from cholesterol-fed rabbits. In addition, in cells from hypercholesterolemic rabbits, nitric oxide added before angiotensin II inhibited to a smaller degree the transient increase in intracellular free Ca²⁺ caused by angiotensin II in the nominal absence of extracellular Ca²⁺, as well as the increase in Ca²⁺ associated with the addition of extracellular Ca²⁺. Measurements of fura 2 quenching caused by Mn²⁺ influx confirmed that nitric oxide inhibited the entry of extracellular divalent cations significantly less in cells from hypercholesterolemic rabbits. Basal levels of cyclic GMP were significantly less than normal, and nitric oxide increased levels of cyclic GMP to a significantly smaller degree in cells from cholesterol-fed rabbits. These data indicate a substantial resistance to nitric oxide action in aortic smooth muscle cells of cholesterol-fed rabbits. This observation is consistent with the notion that resistance of smooth muscle cells to nitric oxide contributes to abnormal endothelium-dependent vasodilatation during hypercholesterolemia and can play a role in the pathogenesis of atherosclerosis.

Key Words: nitric oxide • calcium • cyclic GMP • cholesterol • atherosclerosis • smooth muscle cell

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