title: Reduction-oxidation (Redox) and vascular tissue level of homocyst(e)ine in human coronary atherosclerotic lesions and role in extracellular matrix remodeling and vascular tone.
abstract: Hyperhomocyst(e)inemia in patients with coronary and peripheral arterial occlusion has been demonstrated by others. Redox-state of homocyst(e)ine causes dysfunction of endothelial cells and promote growth of vascular smooth muscle cells. The role of tissue, protein bound and unbound, oxidative mixed disulfides in the development of fibrous plaque in atherosclerotic lesion is not known. Redox-state around the fibroblasts and vascular smooth muscle cells modulates the expression of extracellular matrix (ECM) components (Tyagi et al. 1996, J Cell Biochem, 61: 139-151). To determine the role of tissue homocystine in fibrotic atherosclerotic plaque development, coronary arteries were isolated from ischemic explanted hearts (n = 10). Apparently normal vascular tissue was obtained from idiopathic cardiomyopathic explanted hearts (n = 10). Tissue extract were prepared from atherosclerotic lesions and from normal arteries devoid of adventitia. Interaction of homocystine with Ellman's reagent (5, 5'-dithio-bis-2-nitro benzoic acid) catalyzed by limiting amount of reducing agent (catalyst) generated change in optical density (OD) at 412 nm in dose dependent fashion. We have generated a standard curve between change at 412 nm and amount of homocystine. The change in OD at 412 nm with increasing amount (0-25 microg) of homocystine demonstrated linearity. The protein-bound oxidized disulfides were precipitated by trichloroacetic acid (TCA) and free-oxidative disulfides in the supernatant were collected. The pathophysiological amount of protein-bound disulfide in atherosclerotic tissue (1.0 +/- 0.2 microg/mg total protein) was 10 times that in normal tissue (0.1 +/- 0.01 microg/mg, p < 0.001). The amount of free oxidative disulfide in atherosclerotic tissue (1.5 +/- 0.3 microg/mg) was 15 times that in normal tissue (0.12 +/- 0.02 microg/mg, p < 0.001). To determine the role of homocystine in ECM expression, ECM collagenase activity in the presence and absence of homocystine was measured by zymography. The effect of homocysteine on collagenase activity was biphasic, increased at < [0.01 mM] and inhibited at > [0.1 mM]. To determine whether homocystine regulates vascular tone, isometric measurements were carried out using normal coronary rings. Results suggested that homocystine induced endothelial-modulated vasoconstriction in coronary vessels. Tissue oxidative disulfides and the homocystine may contribute to the development of fibrotic atherosclerotic lesions and vascular dysfunction.
authors.affiliation: Department of Physiology and Biophysics, University of Mississippi Medical Center, Jackson 39216-4505, USA.
authors.forename: S C
authors.initials: SC
authors.lastname: Tyagi
authors.forename: L M
authors.initials: LM
authors.lastname: Smiley
authors.forename: V S
authors.initials: VS
authors.lastname: Mujumdar
authors.forename: B
authors.initials: B
authors.lastname: Clonts
authors.forename: J L
authors.initials: JL
authors.lastname: Parker
mesh_terms: D001158:Arteries / Q000201:enzymology / Q000378:metabolism; D002457:Cell Extracts; D003324:Coronary Artery Disease / Q000378:metabolism*; D003331:Coronary Vessels / Q000201:enzymology / Q000378:metabolism*; D046508:Culture Techniques; D003545:Cysteine / Q000494:pharmacology; D003553:Cystine / Q000494:pharmacology; D004220:Disulfides / Q000378:metabolism; D004228:Dithionitrobenzoic Acid; D004730:Endothelium, Vascular / Q000502:physiology; D005109:Extracellular Matrix / Q000201:enzymology / Q000378:metabolism*; D006710:Homocysteine / Q000494:pharmacology; D006711:Homocystine / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007537:Isometric Contraction / Q000187:drug effects; D008666:Metalloendopeptidases / Q000378:metabolism; D010084:Oxidation-Reduction; D014661:Vasoconstriction / Q000187:drug effects
chemical_list: D002457:Cell Extracts; D004220:Disulfides; D006710:Homocysteine; D006711:Homocystine; D003553:Cystine; D004228:Dithionitrobenzoic Acid; D008666:Metalloendopeptidases; D003545:Cysteine