During this project we will examine human apo B-containing lipoproteins from surgical endarterectomy specimens and plasma. We will use selected-affinity immunosorption (SAI) to provide unperturbed lipoproteins for study, since there is evidence that ultracentrifugation may alter lipoproteins during their isolation (1). The work is divided into two parts. In part one we will fully characterize the lipid and protein content of SAI-isolated plasma VLDL, IDL, and LDL and compare them to centrifuged particules. Preliminary results for LDL have demonstrated several additional proteins not present after ultracentrifugation. As lipoprotein behavior is principally regulated by the protein components, isolation and identification of these new apoproteins may allow a better delineation of in vivo lipoprotein metabolism. We will also examine these unperturbed lipoproteins for subspeciation by comparing apoprotein molar ratios and will do subspecies separation when indicated. The receptor binding of SAI-isolated lipoproteins will be examined in human fibroblast and monocyte-macrophage tissue culture. In part two of this project we will also use SAI to isolate lipoproteins from the interstitial fluid of freshly isolated atherosclerotic plaque. Preliminary results demonstrate that intact lipoproteins of all three apo B-containing classes can be extracted from this tissue. The components of these lipoproteins will be analyzed and compared to plasma lipoproteins from the same patient. Previous studies of lipoproteins isolated from cadaveric aortas report substantial differences from plasma. Such differences may due to postmortem proteolytic degradation. Our extensive analysis of freshly isolated material should allow a determination of what alterations, if any, develop in vivo. The receptor binding of these plaque lipoproteins will also be assessed in human fibroblast and monocyte-macrophage tissue culture.
Showing the most recent 10 out of 15 publications
