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. 1990 Aug;137(2):457–465.

Lipoprotein degradation and cholesterol esterification in primary cell cultures of rabbit atherosclerotic lesions.

O Jaakkola 1, T Nikkari 1
PMCID: PMC1877621  PMID: 2201201

Abstract

Lipoprotein metabolism and cholesterol accumulation in atherosclerotic lesions was studied using enzymatically isolated primary cell cultures from aortas of rabbits made atherosclerotic by cholesterol feeding. The cultures consisted of macrophages and smooth muscle cells, thus resembling, in composition, fatty streak lesions. The mean (+/- SD) cholesteryl ester content of the dispersed cells was 1059 +/- 445 micrograms/mg cell protein, but it declined steeply during 1 week in primary culture. The uptake of low-density lipoprotein (LDL), beta-migrating very low-density lipoprotein (beta-VLDL), and acetylated LDL (acetyl-LDL), labeled with 125I or with the fluorescent probe 1,1'-dioctadecyl-3,3,3',3'- tetramethylindocarbocyanine (DiI), was studied in 2-day-old primary cultures. DiI-acetyl-LDL was avidly taken up by the macrophages and, to a lesser extent, by some smooth muscle cells. The uptake of DiI-beta-VLDL by the macrophages was weaker and less homogeneous than that of DiI-acetyl-LDL. The degradation rates of 125I-labeled beta-VLDL, LDL and acetyl-LDL were 135 +/- 54, 195 +/- 20, and 697 +/- 14 ng/mg cell protein/8 hours, respectively. Incubation with unlabeled acetyl-LDL enhanced the incorporation of [3H]oleate into cholesteryl esters and increased the cellular cholesteryl ester content. These results suggest that arterial macrophages and, to some extent, smooth muscle cells from cholesterol-fed rabbits actively metabolize acetyl-LDL and are thus capable of accumulating cholesteryl esters by uptake of modified forms of LDL.

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Selected References

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  1. Basu S. K., Goldstein J. L., Anderson G. W., Brown M. S. Degradation of cationized low density lipoprotein and regulation of cholesterol metabolism in homozygous familial hypercholesterolemia fibroblasts. Proc Natl Acad Sci U S A. 1976 Sep;73(9):3178–3182. doi: 10.1073/pnas.73.9.3178. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bierman E. L., Stein O., Stein Y. Lipoprotein uptake and metabolism by rat aortic smooth muscle cells in tissue culture. Circ Res. 1974 Jul;35(1):136–150. doi: 10.1161/01.res.35.1.136. [DOI] [PubMed] [Google Scholar]
  3. Bilheimer D. W., Eisenberg S., Levy R. I. The metabolism of very low density lipoprotein proteins. I. Preliminary in vitro and in vivo observations. Biochim Biophys Acta. 1972 Feb 21;260(2):212–221. doi: 10.1016/0005-2760(72)90034-3. [DOI] [PubMed] [Google Scholar]
  4. Brown M. S., Goldstein J. L., Krieger M., Ho Y. K., Anderson R. G. Reversible accumulation of cholesteryl esters in macrophages incubated with acetylated lipoproteins. J Cell Biol. 1979 Sep;82(3):597–613. doi: 10.1083/jcb.82.3.597. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brown M. S., Goldstein J. L. Lipoprotein metabolism in the macrophage: implications for cholesterol deposition in atherosclerosis. Annu Rev Biochem. 1983;52:223–261. doi: 10.1146/annurev.bi.52.070183.001255. [DOI] [PubMed] [Google Scholar]
  6. Brown M. S., Ho Y. K., Goldstein J. L. The cholesteryl ester cycle in macrophage foam cells. Continual hydrolysis and re-esterification of cytoplasmic cholesteryl esters. J Biol Chem. 1980 Oct 10;255(19):9344–9352. [PubMed] [Google Scholar]
  7. Daugherty A., Lange L. G., Sobel B. E., Schonfeld G. Aortic accumulation and plasma clearance of beta-VLDL and HDL: effects of diet-induced hypercholesterolemia in rabbits. J Lipid Res. 1985 Aug;26(8):955–963. [PubMed] [Google Scholar]
  8. Day A. J., Tume R. K. In vitro incorporation of 14C-labelled oleic acid into combined lipid by foam cells from rabbit atheromatous lesions. J Atheroscler Res. 1969 Mar-Apr;9(2):141–149. doi: 10.1016/s0368-1319(69)80049-9. [DOI] [PubMed] [Google Scholar]
  9. Day A. J., Wahlqvist M. L. Uptake and metabolism of 14C-labeled oleic acid by atherosclerotic lesions in rabbit aorta. Circ Res. 1968 Dec;23(6):779–783. doi: 10.1161/01.res.23.6.779. [DOI] [PubMed] [Google Scholar]
  10. Ellsworth J. L., Kraemer F. B., Cooper A. D. Transport of beta-very low density lipoproteins and chylomicron remnants by macrophages is mediated by the low density lipoprotein receptor pathway. J Biol Chem. 1987 Feb 15;262(5):2316–2325. [PubMed] [Google Scholar]
  11. FOLCH J., LEES M., SLOANE STANLEY G. H. A simple method for the isolation and purification of total lipides from animal tissues. J Biol Chem. 1957 May;226(1):497–509. [PubMed] [Google Scholar]
  12. Faggiotto A., Ross R., Harker L. Studies of hypercholesterolemia in the nonhuman primate. I. Changes that lead to fatty streak formation. Arteriosclerosis. 1984 Jul-Aug;4(4):323–340. doi: 10.1161/01.atv.4.4.323. [DOI] [PubMed] [Google Scholar]
  13. Fogelman A. M., Shechter I., Seager J., Hokom M., Child J. S., Edwards P. A. Malondialdehyde alteration of low density lipoproteins leads to cholesteryl ester accumulation in human monocyte-macrophages. Proc Natl Acad Sci U S A. 1980 Apr;77(4):2214–2218. doi: 10.1073/pnas.77.4.2214. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fowler S., Shio H., Haley N. J. Characterization of lipid-laden aortic cells from cholesterol-fed rabbits. IV. Investigation of macrophage-like properties of aortic cell populations. Lab Invest. 1979 Oct;41(4):372–378. [PubMed] [Google Scholar]
  15. Goldstein J. L., Ho Y. K., Basu S. K., Brown M. S. Binding site on macrophages that mediates uptake and degradation of acetylated low density lipoprotein, producing massive cholesterol deposition. Proc Natl Acad Sci U S A. 1979 Jan;76(1):333–337. doi: 10.1073/pnas.76.1.333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Goldstein J. L., Ho Y. K., Brown M. S., Innerarity T. L., Mahley R. W. Cholesteryl ester accumulation in macrophages resulting from receptor-mediated uptake and degradation of hypercholesterolemic canine beta-very low density lipoproteins. J Biol Chem. 1980 Mar 10;255(5):1839–1848. [PubMed] [Google Scholar]
  17. Gown A. M., Tsukada T., Ross R. Human atherosclerosis. II. Immunocytochemical analysis of the cellular composition of human atherosclerotic lesions. Am J Pathol. 1986 Oct;125(1):191–207. [PMC free article] [PubMed] [Google Scholar]
  18. Hara A., Radin N. S. Lipid extraction of tissues with a low-toxicity solvent. Anal Biochem. 1978 Oct 1;90(1):420–426. doi: 10.1016/0003-2697(78)90046-5. [DOI] [PubMed] [Google Scholar]
  19. Heinecke J. W., Baker L., Rosen H., Chait A. Superoxide-mediated modification of low density lipoprotein by arterial smooth muscle cells. J Clin Invest. 1986 Mar;77(3):757–761. doi: 10.1172/JCI112371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Henriksen T., Mahoney E. M., Steinberg D. Enhanced macrophage degradation of biologically modified low density lipoprotein. Arteriosclerosis. 1983 Mar-Apr;3(2):149–159. doi: 10.1161/01.atv.3.2.149. [DOI] [PubMed] [Google Scholar]
  21. Hiramatsu K., Rosen H., Heinecke J. W., Wolfbauer G., Chait A. Superoxide initiates oxidation of low density lipoprotein by human monocytes. Arteriosclerosis. 1987 Jan-Feb;7(1):55–60. doi: 10.1161/01.atv.7.1.55. [DOI] [PubMed] [Google Scholar]
  22. Innerarity T. L., Arnold K. S., Weisgraber K. H., Mahley R. W. Apolipoprotein E is the determinant that mediates the receptor uptake of beta-very low density lipoproteins by mouse macrophages. Arteriosclerosis. 1986 Jan-Feb;6(1):114–122. doi: 10.1161/01.atv.6.1.114. [DOI] [PubMed] [Google Scholar]
  23. Ives H. E., Schultz G. S., Galardy R. E., Jamieson J. D. Preparation of functional smooth muscle cells from the rabbit aorta. J Exp Med. 1978 Nov 1;148(5):1400–1413. doi: 10.1084/jem.148.5.1400. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jaakkola O., Kallioniemi O. P., Nikkari T. Lipoprotein uptake in primary cell cultures of rabbit atherosclerotic lesions. A fluorescence microscopic and flow cytometric study. Atherosclerosis. 1988 Feb;69(2-3):257–268. doi: 10.1016/0021-9150(88)90022-6. [DOI] [PubMed] [Google Scholar]
  25. Kallioniemi O. P., Jaakkola O., Nikkari S. T., Nikkari T. Growth properties and composition of cytoskeletal and cytocontractile proteins in aortic cells isolated and cultured from normal and atherosclerotic rabbits. Atherosclerosis. 1984 Jul;52(1):13–26. doi: 10.1016/0021-9150(84)90153-9. [DOI] [PubMed] [Google Scholar]
  26. Kovanen P. T., Brown M. S., Basu S. K., Bilheimer D. W., Goldstein J. L. Saturation and suppression of hepatic lipoprotein receptors: a mechanism for the hypercholesterolemia of cholesterol-fed rabbits. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1396–1400. doi: 10.1073/pnas.78.3.1396. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  28. Mahley R. W., Innerarity T. L., Weisgraber K. B., Oh S. Y. Altered metabolism (in vivo and in vitro) of plasma lipoproteins after selective chemical modification of lysine residues of the apoproteins. J Clin Invest. 1979 Sep;64(3):743–750. doi: 10.1172/JCI109518. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Orekhov A. N., Tertov V. V., Smirnov V. N. Lipids in cells of atherosclerotic and uninvolved human aorta. II. Lipid metabolism in primary culture. Exp Mol Pathol. 1985 Oct;43(2):187–195. doi: 10.1016/0014-4800(85)90039-5. [DOI] [PubMed] [Google Scholar]
  30. Parthasarathy S., Printz D. J., Boyd D., Joy L., Steinberg D. Macrophage oxidation of low density lipoprotein generates a modified form recognized by the scavenger receptor. Arteriosclerosis. 1986 Sep-Oct;6(5):505–510. doi: 10.1161/01.atv.6.5.505. [DOI] [PubMed] [Google Scholar]
  31. Pitas R. E., Innerarity T. L., Mahley R. W. Foam cells in explants of atherosclerotic rabbit aortas have receptors for beta-very low density lipoproteins and modified low density lipoproteins. Arteriosclerosis. 1983 Jan-Feb;3(1):2–12. doi: 10.1161/01.atv.3.1.2. [DOI] [PubMed] [Google Scholar]
  32. Pitas R. E., Innerarity T. L., Weinstein J. N., Mahley R. W. Acetoacetylated lipoproteins used to distinguish fibroblasts from macrophages in vitro by fluorescence microscopy. Arteriosclerosis. 1981 May-Jun;1(3):177–185. doi: 10.1161/01.atv.1.3.177. [DOI] [PubMed] [Google Scholar]
  33. Portman O. W., O'Malley J. P., Alexander M. Metabolism of native and acetylated low density lipoproteins in squirrel monkeys with emphasis on aortas with varying severities of atherosclerosis. Atherosclerosis. 1987 Aug;66(3):227–235. doi: 10.1016/0021-9150(87)90066-9. [DOI] [PubMed] [Google Scholar]
  34. Schaffner T., Taylor K., Bartucci E. J., Fischer-Dzoga K., Beeson J. H., Glagov S., Wissler R. W. Arterial foam cells with distinctive immunomorphologic and histochemical features of macrophages. Am J Pathol. 1980 Jul;100(1):57–80. [PMC free article] [PubMed] [Google Scholar]
  35. Slater H. R., Shepherd J., Packard C. J. Receptor-mediated catabolism and tissue uptake of human low density lipoprotein in the cholesterol-fed, atherosclerotic rabbit. Biochim Biophys Acta. 1982 Nov 12;713(2):435–445. doi: 10.1016/0005-2760(82)90263-6. [DOI] [PubMed] [Google Scholar]
  36. Spector A. A., Hoak J. C. An improved method for the addition of long-chain free fatty acid to protein solutions. Anal Biochem. 1969 Nov;32(2):297–302. doi: 10.1016/0003-2697(69)90089-x. [DOI] [PubMed] [Google Scholar]
  37. Tsukada T., Rosenfeld M., Ross R., Gown A. M. Immunocytochemical analysis of cellular components in atherosclerotic lesions. Use of monoclonal antibodies with the Watanabe and fat-fed rabbit. Arteriosclerosis. 1986 Nov-Dec;6(6):601–613. doi: 10.1161/01.atv.6.6.601. [DOI] [PubMed] [Google Scholar]
  38. Wahlqvist M. L., Day A. J., Tume R. K. Incorporation of oleic acid into lipid by foam cells in human atherosclerotic lesions. Circ Res. 1969 Jan;24(1):123–130. doi: 10.1161/01.res.24.1.123. [DOI] [PubMed] [Google Scholar]
  39. Witte L. D., Cornicelli J. A. Platelet-derived growth factor stimulates low density lipoprotein receptor activity in cultured human fibroblasts. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5962–5966. doi: 10.1073/pnas.77.10.5962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Ylä-Herttuala S., Jaakkola O., Solakivi T., Kuivaniemi H., Nikkari T. The effect of proteoglycans, collagen and lysyl oxidase on the metabolism of low density lipoprotein by macrophages. Atherosclerosis. 1986 Oct;62(1):73–80. doi: 10.1016/0021-9150(86)90021-3. [DOI] [PubMed] [Google Scholar]
  41. Ylä-Herttuala S., Palinski W., Rosenfeld M. E., Parthasarathy S., Carew T. E., Butler S., Witztum J. L., Steinberg D. Evidence for the presence of oxidatively modified low density lipoprotein in atherosclerotic lesions of rabbit and man. J Clin Invest. 1989 Oct;84(4):1086–1095. doi: 10.1172/JCI114271. [DOI] [PMC free article] [PubMed] [Google Scholar]

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