Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1997 Sep 15;100(6):1493–1500. doi: 10.1172/JCI119671

Lipoprotein(a) vascular accumulation in mice. In vivo analysis of the role of lysine binding sites using recombinant adenovirus.

S D Hughes 1, X J Lou 1, S Ighani 1, J Verstuyft 1, D J Grainger 1, R M Lawn 1, E M Rubin 1
PMCID: PMC508329  PMID: 9294116

Abstract

Although the mechanism by which lipoprotein(a) [Lp(a)] contributes to vascular disease remains unclear, consequences of its binding to the vessel surface are commonly cited in postulated atherogenic pathways. Because of the presence of plasminogen-like lysine binding sites (LBS) in apo(a), fibrin binding has been proposed to play an important role in Lp(a)'s vascular accumulation. Indeed, LBS are known to facilitate Lp(a) fibrin binding in vitro. To examine the importance of apo(a) LBS in Lp(a) vascular accumulation in vivo, we generated three different apo(a) cDNAs: (a) mini apo(a), based on wild-type human apo(a); (b) mini apo(a) containing a naturally occurring LBS defect associated with a point mutation in kringle 4-10; and (c) human- rhesus monkey chimeric mini apo(a), which contains the same LBS defect in the context of several additional changes. Recombinant adenovirus vectors were constructed with the various apo(a) cDNAs and injected into human apoB transgenic mice. At the viral dosage used in these experiments, all three forms of apo(a) were found exclusively within the lipoprotein fractions, and peak Lp(a) plasma levels were nearly identical (approximately 45 mg/dl). In vitro analysis of Lp(a) isolated from the various groups of mice confirmed that putative LBS defective apo(a) yielded Lp(a) unable to bind lysine-Sepharose. Quantitation of in vivo Lp(a) vascular accumulation in mice treated with the various adenovirus vectors revealed significantly less accumulation of both types of LBS defective Lp(a), relative to wild-type Lp(a). These results indicate a correlation between lysine binding properties of Lp(a) and vascular accumulation, supporting the postulated role of apo(a) LBS in this potentially atherogenic characteristic of Lp(a).

Full Text

The Full Text of this article is available as a PDF (408.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bett A. J., Haddara W., Prevec L., Graham F. L. An efficient and flexible system for construction of adenovirus vectors with insertions or deletions in early regions 1 and 3. Proc Natl Acad Sci U S A. 1994 Sep 13;91(19):8802–8806. doi: 10.1073/pnas.91.19.8802. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Boonmark N. W., Lou X. J., Yang Z. J., Schwartz K., Zhang J. L., Rubin E. M., Lawn R. M. Modification of apolipoprotein(a) lysine binding site reduces atherosclerosis in transgenic mice. J Clin Invest. 1997 Aug 1;100(3):558–564. doi: 10.1172/JCI119565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brunner C., Kraft H. G., Utermann G., Müller H. J. Cys4057 of apolipoprotein(a) is essential for lipoprotein(a) assembly. Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):11643–11647. doi: 10.1073/pnas.90.24.11643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Callow M. J., Stoltzfus L. J., Lawn R. M., Rubin E. M. Expression of human apolipoprotein B and assembly of lipoprotein(a) in transgenic mice. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2130–2134. doi: 10.1073/pnas.91.6.2130. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Callow M. J., Verstuyft J., Tangirala R., Palinski W., Rubin E. M. Atherogenesis in transgenic mice with human apolipoprotein B and lipoprotein (a). J Clin Invest. 1995 Sep;96(3):1639–1646. doi: 10.1172/JCI118203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cushing G. L., Gaubatz J. W., Nava M. L., Burdick B. J., Bocan T. M., Guyton J. R., Weilbaecher D., DeBakey M. E., Lawrie G. M., Morrisett J. D. Quantitation and localization of apolipoproteins [a] and B in coronary artery bypass vein grafts resected at re-operation. Arteriosclerosis. 1989 Sep-Oct;9(5):593–603. doi: 10.1161/01.atv.9.5.593. [DOI] [PubMed] [Google Scholar]
  7. Ernst A., Helmhold M., Brunner C., Pethö-Schramm A., Armstrong V. W., Müller H. J. Identification of two functionally distinct lysine-binding sites in kringle 37 and in kringles 32-36 of human apolipoprotein(a). J Biol Chem. 1995 Mar 17;270(11):6227–6234. doi: 10.1074/jbc.270.11.6227. [DOI] [PubMed] [Google Scholar]
  8. Frank S., Durovic S., Kostner G. M. Structural requirements of apo-a for the lipoprotein-a assembly. Biochem J. 1994 Nov 15;304(Pt 1):27–30. doi: 10.1042/bj3040027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gabel B. R., May L. F., Marcovina S. M., Koschinsky M. L. Lipoprotein(a) assembly. Quantitative assessment of the role of apo(a) kringle IV types 2-10 in particle formation. Arterioscler Thromb Vasc Biol. 1996 Dec;16(12):1559–1567. doi: 10.1161/01.atv.16.12.1559. [DOI] [PubMed] [Google Scholar]
  10. Grainger D. J., Kemp P. R., Liu A. C., Lawn R. M., Metcalfe J. C. Activation of transforming growth factor-beta is inhibited in transgenic apolipoprotein(a) mice. Nature. 1994 Aug 11;370(6489):460–462. doi: 10.1038/370460a0. [DOI] [PubMed] [Google Scholar]
  11. Guevara J., Jr, Jan A. Y., Knapp R., Tulinsky A., Morrisett J. D. Comparison of ligand-binding sites of modeled apo[a] kringle-like sequences in human lipoprotein[a]. Arterioscler Thromb. 1993 May;13(5):758–770. doi: 10.1161/01.atv.13.5.758. [DOI] [PubMed] [Google Scholar]
  12. Hajjar K. A., Gavish D., Breslow J. L., Nachman R. L. Lipoprotein(a) modulation of endothelial cell surface fibrinolysis and its potential role in atherosclerosis. Nature. 1989 May 25;339(6222):303–305. doi: 10.1038/339303a0. [DOI] [PubMed] [Google Scholar]
  13. Harpel P. C., Gordon B. R., Parker T. S. Plasmin catalyzes binding of lipoprotein (a) to immobilized fibrinogen and fibrin. Proc Natl Acad Sci U S A. 1989 May;86(10):3847–3851. doi: 10.1073/pnas.86.10.3847. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hoover-Plow J. L., Boonmark N., Skocir P., Lawn R., Plow E. F. A quantitative immunoassay for the lysine-binding function of lipoprotein(a). Application to recombinant apo(a) and lipoprotein(a) in plasma. Arterioscler Thromb Vasc Biol. 1996 May;16(5):656–664. doi: 10.1161/01.atv.16.5.656. [DOI] [PubMed] [Google Scholar]
  15. Huby T., Schröder W., Doucet C., Chapman J., Thillet J. Characterization of the N-terminal and C-terminal domains of human apolipoprotein(a): relevance to fibrin binding. Biochemistry. 1995 Jun 6;34(22):7385–7393. doi: 10.1021/bi00022a011. [DOI] [PubMed] [Google Scholar]
  16. Hughes S. D., Rouy D., Navaratnam N., Scott J., Rubin E. M. Gene transfer of cytidine deaminase apoBEC-1 lowers lipoprotein(a) in transgenic mice and induces apolipoprotein B editing in rabbits. Hum Gene Ther. 1996 Jan;7(1):39–49. doi: 10.1089/hum.1996.7.1-39. [DOI] [PubMed] [Google Scholar]
  17. Klezovitch O., Edelstein C., Scanu A. M. Evidence that the fibrinogen binding domain of Apo(a) is outside the lysine binding site of kringle IV-10: a study involving naturally occurring lysine binding defective lipoprotein(a) phenotypes. J Clin Invest. 1996 Jul 1;98(1):185–191. doi: 10.1172/JCI118765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Koschinsky M. L., Côté G. P., Gabel B., van der Hoek Y. Y. Identification of the cysteine residue in apolipoprotein(a) that mediates extracellular coupling with apolipoprotein B-100. J Biol Chem. 1993 Sep 15;268(26):19819–19825. [PubMed] [Google Scholar]
  19. Kreuzer J., Lloyd M. B., Bok D., Fless G. M., Scanu A. M., Lusis A. J., Haberland M. E. Lipoprotein (a) displays increased accumulation compared with low-density lipoprotein in the murine arterial wall. Chem Phys Lipids. 1994 Jan;67-68:175–190. doi: 10.1016/0009-3084(94)90137-6. [DOI] [PubMed] [Google Scholar]
  20. Lawn R. M., Pearle A. D., Kunz L. L., Rubin E. M., Reckless J., Metcalfe J. C., Grainger D. J. Feedback mechanism of focal vascular lesion formation in transgenic apolipoprotein(a) mice. J Biol Chem. 1996 Dec 6;271(49):31367–31371. doi: 10.1074/jbc.271.49.31367. [DOI] [PubMed] [Google Scholar]
  21. Lawn R. M., Wade D. P., Hammer R. E., Chiesa G., Verstuyft J. G., Rubin E. M. Atherogenesis in transgenic mice expressing human apolipoprotein(a) Nature. 1992 Dec 17;360(6405):670–672. doi: 10.1038/360670a0. [DOI] [PubMed] [Google Scholar]
  22. Maher V. M., Brown B. G. Lipoprotein (a) and coronary heart disease. Curr Opin Lipidol. 1995 Aug;6(4):229–235. doi: 10.1097/00041433-199508000-00007. [DOI] [PubMed] [Google Scholar]
  23. McLean J. W., Tomlinson J. E., Kuang W. J., Eaton D. L., Chen E. Y., Fless G. M., Scanu A. M., Lawn R. M. cDNA sequence of human apolipoprotein(a) is homologous to plasminogen. Nature. 1987 Nov 12;330(6144):132–137. doi: 10.1038/330132a0. [DOI] [PubMed] [Google Scholar]
  24. Miles L. A., Fless G. M., Levin E. G., Scanu A. M., Plow E. F. A potential basis for the thrombotic risks associated with lipoprotein(a). Nature. 1989 May 25;339(6222):301–303. doi: 10.1038/339301a0. [DOI] [PubMed] [Google Scholar]
  25. Mosedale D. E., Metcalfe J. C., Grainger D. J. Optimization of immunofluorescence methods by quantitative image analysis. J Histochem Cytochem. 1996 Sep;44(9):1043–1050. doi: 10.1177/44.9.8773570. [DOI] [PubMed] [Google Scholar]
  26. Mulichak A. M., Tulinsky A., Ravichandran K. G. Crystal and molecular structure of human plasminogen kringle 4 refined at 1.9-A resolution. Biochemistry. 1991 Oct 29;30(43):10576–10588. doi: 10.1021/bi00107a029. [DOI] [PubMed] [Google Scholar]
  27. Nachman R. L. Review: Stratton Lecture. Thrombosis and atherogenesis: molecular connections. Blood. 1992 Apr 15;79(8):1897–1906. [PubMed] [Google Scholar]
  28. Nielsen L. B., Stender S., Jauhiainen M., Nordestgaard B. G. Preferential influx and decreased fractional loss of lipoprotein(a) in atherosclerotic compared with nonlesioned rabbit aorta. J Clin Invest. 1996 Jul 15;98(2):563–571. doi: 10.1172/JCI118824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Palabrica T. M., Liu A. C., Aronovitz M. J., Furie B., Lawn R. M., Furie B. C. Antifibrinolytic activity of apolipoprotein(a) in vivo: human apolipoprotein(a) transgenic mice are resistant to tissue plasminogen activator-mediated thrombolysis. Nat Med. 1995 Mar;1(3):256–259. doi: 10.1038/nm0395-256. [DOI] [PubMed] [Google Scholar]
  30. Pepin J. M., O'Neil J. A., Hoff H. F. Quantification of apo[a] and apoB in human atherosclerotic lesions. J Lipid Res. 1991 Feb;32(2):317–327. [PubMed] [Google Scholar]
  31. Rath M., Niendorf A., Reblin T., Dietel M., Krebber H. J., Beisiegel U. Detection and quantification of lipoprotein(a) in the arterial wall of 107 coronary bypass patients. Arteriosclerosis. 1989 Sep-Oct;9(5):579–592. doi: 10.1161/01.atv.9.5.579. [DOI] [PubMed] [Google Scholar]
  32. Scanu A. M., Miles L. A., Fless G. M., Pfaffinger D., Eisenbart J., Jackson E., Hoover-Plow J. L., Brunck T., Plow E. F. Rhesus monkey lipoprotein(a) binds to lysine Sepharose and U937 monocytoid cells less efficiently than human lipoprotein(a). Evidence for the dominant role of kringle 4(37). J Clin Invest. 1993 Jan;91(1):283–291. doi: 10.1172/JCI116182. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Scanu A. M., Pfaffinger D., Lee J. C., Hinman J. A single point mutation (Trp72-->Arg) in human apo(a) kringle 4-37 associated with a lysine binding defect in Lp(a). Biochim Biophys Acta. 1994 Oct 21;1227(1-2):41–45. doi: 10.1016/0925-4439(94)90104-x. [DOI] [PubMed] [Google Scholar]
  34. Tomlinson J. E., McLean J. W., Lawn R. M. Rhesus monkey apolipoprotein(a). Sequence, evolution, and sites of synthesis. J Biol Chem. 1989 Apr 5;264(10):5957–5965. [PubMed] [Google Scholar]
  35. Trieu V. N., McConathy W. J. A two-step model for lipoprotein(a) formation. J Biol Chem. 1995 Jun 30;270(26):15471–15474. doi: 10.1074/jbc.270.26.15471. [DOI] [PubMed] [Google Scholar]
  36. Utermann G. The mysteries of lipoprotein(a). Science. 1989 Nov 17;246(4932):904–910. doi: 10.1126/science.2530631. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES