PATHOBIOLOGICAL DETERMINANTS OF ATHEROSCLEROSIS IN YOUTH

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This application is submitted as a participating unit in the USA investigator-initiated multicenter study of the pathobiological determinants of atherosclerosis in youth. The conceptual basis for the application rests on several considerations and observations. Chief among these are: 1) the functional integrity of the walls of major vessels is, for the most part, maintained through the presence of collagen fibers; and 2) the quantity of collagen as well as the relative proportions of various collagen types is generally altered during the processes leading to the appearance of atherosclerotic lesions in humans. In addition, it is believed that the processes leading to the formation of atherosclerotic lesions are operative over lengthy intervals. In this regard, participation in the contemplated consortium arrangement provides a unique opportunity to evaluate potential modifications of vascular collagen status in a defined preclinical population and correlate the findings with age, sex, race and habitat of the subjects included in the overall study. Accordingly, it is anticipated that the collecting centers involved in the study will provide approximately 50 samples of vascular tissue per year. The samples will be from 25 subjects and involve one anatomically-defined segment (20 mm) of thoracic aorta as well as one anatomically defined segment (40 mm) of the left circumflex coronary artery per subject. The complexity of the contemplated analyses precludes analyses of more than the indicated samples on a yearly basis. With respect to the analyses, each sample will be freed of adventitia, finely divided by physical means, and a small aliquot subjected to amino acid analysis in order to evaluate total collagen content. Since previous investigations have shown that limited proteolysis with pepsin is only marginally effective in quantitatively solubilizing collagens in vascular tissue, the collagens will be solubilized as peptide fragments utilizing specific chemical cleavage with CNBr. The resulting peptide fragments will then be chromatographed on CM-cellulose and portions of the column effluent known to contain marker peptides derived from the chains of type I, III, IV and V collagens will be chosen for further analyses by gel permeation HPLC in order to achieve resolution and quantitation of the peptides derived from various chains. Since type I, III, IV and V collagens constitute the major collagens present in vascular tissues, these analyses will provide basic data concerning any potential quantitative and/or qualitative alterations in vascular collagen status as a function of the parameters noted above.