Skip to main content
PMC home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1995 Sep;96(3):1612–1620. doi: 10.1172/JCI118200

Apolipoprotein E deficiency in mice: gene replacement and prevention of atherosclerosis using adenovirus vectors.

V S Kashyap 1, S Santamarina-Fojo 1, D R Brown 1, C L Parrott 1, D Applebaum-Bowden 1, S Meyn 1, G Talley 1, B Paigen 1, N Maeda 1, H B Brewer Jr 1
PMCID: PMC185787  PMID: 7657831

Abstract

Apolipoprotein E (apoE)-deficient mice develop marked hyperlipidemia as well as atherosclerosis and thus are an excellent animal model for evaluating the potential for gene therapy in human genetic dyslipoproteinemias. Recombinant adenovirus containing either human apoE (rAdv.apoE) or the reporter gene luciferase (rAdv.luc) were generated and infused intravenously in apoE-deficient mice with preinfusion plasma total cholesterol of 644 +/- 149 mg/dl an cholesterol rich VLDL/IDL. After a single infusion of rAdv.apoE, plasma concentrations of human apoE ranging from 1.5 to 650 mg/dl were achieved. Adenovirus-mediated apoE replacement resulted in normalization of the lipid and lipoprotein profile with markedly decreased total cholesterol (103 +/- 18mg/dl), VLDL, IDL, and LDL, as well as increased HDL. Measurement of aortic atherosclerosis 1 mo after adenoviral infusion demonstrated a marked reduction in the mean lesion area of mice infused with rAdv.apoE (58 +/- 8 x 10(3) microns2) when compared with control mice infused with rAdv.luc (161 +/- 10 x 10(3) microns2; P < 0.0001). Thus, apoE expression for 4 wk was sufficient to markedly reduce atherosclerosis, demonstrating the feasibility of gene therapy for correction of genetic hyperlipidemias resulting in atherosclerosis. The combined use of adenovirus vectors and the apoE-deficient mouse represents a new in vivo approach that will permit rapid screening of candidate genes for the prevention of atherosclerosis.

Full text

PDF
1616

Images in this article

1614Image
on p.1614
1617Image
on p.1617
1617Image
on p.1617
1617Image
on p.1617
1617Image
on p.1617

Selected References

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

  1. Bennett J., Wilson J., Sun D., Forbes B., Maguire A. Adenovirus vector-mediated in vivo gene transfer into adult murine retina. Invest Ophthalmol Vis Sci. 1994 Apr;35(5):2535–2542. [PubMed] [Google Scholar]
  2. Berkner K. L. Development of adenovirus vectors for the expression of heterologous genes. Biotechniques. 1988 Jul-Aug;6(7):616–629. [PubMed] [Google Scholar]
  3. Breslow J. L., McPherson J., Nussbaum A. L., Williams H. W., Lofquist-Kahl F., Karathanasis S. K., Zannis V. I. Identification and DNA sequence of a human apolipoprotein E cDNA clone. J Biol Chem. 1982 Dec 25;257(24):14639–14641. [PubMed] [Google Scholar]
  4. Breslow J. L. Transgenic mouse models of lipoprotein metabolism and atherosclerosis. Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8314–8318. doi: 10.1073/pnas.90.18.8314. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brewer H. B., Jr, Zech L. A., Gregg R. E., Schwartz D., Schaefer E. J. NIH conference. Type III hyperlipoproteinemia: diagnosis, molecular defects, pathology, and treatment. Ann Intern Med. 1983 May;98(5 Pt 1):623–640. doi: 10.7326/0003-4819-98-5-623. [DOI] [PubMed] [Google Scholar]
  6. Chen C., Okayama H. High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol. 1987 Aug;7(8):2745–2752. doi: 10.1128/mcb.7.8.2745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Descamps V., Blumenfeld N., Villeval J. L., Vainchenker W., Perricaudet M., Beuzard Y. Erythropoietin gene transfer and expression in adult normal mice: use of an adenovirus vector. Hum Gene Ther. 1994 Aug;5(8):979–985. doi: 10.1089/hum.1994.5.8-979. [DOI] [PubMed] [Google Scholar]
  8. Engelhardt J. F., Simon R. H., Yang Y., Zepeda M., Weber-Pendleton S., Doranz B., Grossman M., Wilson J. M. Adenovirus-mediated transfer of the CFTR gene to lung of nonhuman primates: biological efficacy study. Hum Gene Ther. 1993 Dec;4(6):759–769. doi: 10.1089/hum.1993.4.6-759. [DOI] [PubMed] [Google Scholar]
  9. Ghiselli G., Schaefer E. J., Gascon P., Breser H. B., Jr Type III hyperlipoproteinemia associated with apolipoprotein E deficiency. Science. 1981 Dec 11;214(4526):1239–1241. doi: 10.1126/science.6795720. [DOI] [PubMed] [Google Scholar]
  10. Gould S. J., Subramani S. Firefly luciferase as a tool in molecular and cell biology. Anal Biochem. 1988 Nov 15;175(1):5–13. doi: 10.1016/0003-2697(88)90353-3. [DOI] [PubMed] [Google Scholar]
  11. Graham F. L., Smiley J., Russell W. C., Nairn R. Characteristics of a human cell line transformed by DNA from human adenovirus type 5. J Gen Virol. 1977 Jul;36(1):59–74. doi: 10.1099/0022-1317-36-1-59. [DOI] [PubMed] [Google Scholar]
  12. Ishibashi S., Brown M. S., Goldstein J. L., Gerard R. D., Hammer R. E., Herz J. Hypercholesterolemia in low density lipoprotein receptor knockout mice and its reversal by adenovirus-mediated gene delivery. J Clin Invest. 1993 Aug;92(2):883–893. doi: 10.1172/JCI116663. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jaffe H. A., Danel C., Longenecker G., Metzger M., Setoguchi Y., Rosenfeld M. A., Gant T. W., Thorgeirsson S. S., Stratford-Perricaudet L. D., Perricaudet M. Adenovirus-mediated in vivo gene transfer and expression in normal rat liver. Nat Genet. 1992 Aug;1(5):372–378. doi: 10.1038/ng0892-372. [DOI] [PubMed] [Google Scholar]
  14. Jiao S., Cole T. G., Kitchens R. T., Pfleger B., Schonfeld G. Genetic heterogeneity of lipoproteins in inbred strains of mice: analysis by gel-permeation chromatography. Metabolism. 1990 Feb;39(2):155–160. doi: 10.1016/0026-0495(90)90069-o. [DOI] [PubMed] [Google Scholar]
  15. Kopfler W. P., Willard M., Betz T., Willard J. E., Gerard R. D., Meidell R. S. Adenovirus-mediated transfer of a gene encoding human apolipoprotein A-I into normal mice increases circulating high-density lipoprotein cholesterol. Circulation. 1994 Sep;90(3):1319–1327. doi: 10.1161/01.cir.90.3.1319. [DOI] [PubMed] [Google Scholar]
  16. Kurosaka D., Teramoto T., Matsushima T., Yokoyama T., Yamada A., Aikawa T., Miyamoto Y., Kurokawa K. Apolipoprotein E deficiency with a depressed mRNA of normal size. Atherosclerosis. 1991 May;88(1):15–20. doi: 10.1016/0021-9150(91)90252-x. [DOI] [PubMed] [Google Scholar]
  17. Le Gal La Salle G., Robert J. J., Berrard S., Ridoux V., Stratford-Perricaudet L. D., Perricaudet M., Mallet J. An adenovirus vector for gene transfer into neurons and glia in the brain. Science. 1993 Feb 12;259(5097):988–990. doi: 10.1126/science.8382374. [DOI] [PubMed] [Google Scholar]
  18. Lemarchand P., Jaffe H. A., Danel C., Cid M. C., Kleinman H. K., Stratford-Perricaudet L. D., Perricaudet M., Pavirani A., Lecocq J. P., Crystal R. G. Adenovirus-mediated transfer of a recombinant human alpha 1-antitrypsin cDNA to human endothelial cells. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6482–6486. doi: 10.1073/pnas.89.14.6482. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Li Q., Kay M. A., Finegold M., Stratford-Perricaudet L. D., Woo S. L. Assessment of recombinant adenoviral vectors for hepatic gene therapy. Hum Gene Ther. 1993 Aug;4(4):403–409. doi: 10.1089/hum.1993.4.4-403. [DOI] [PubMed] [Google Scholar]
  20. Linton M. F., Atkinson J. B., Fazio S. Prevention of atherosclerosis in apolipoprotein E-deficient mice by bone marrow transplantation. Science. 1995 Feb 17;267(5200):1034–1037. doi: 10.1126/science.7863332. [DOI] [PubMed] [Google Scholar]
  21. Lohse P., Brewer H. B., 3rd, Meng M. S., Skarlatos S. I., LaRosa J. C., Brewer H. B., Jr Familial apolipoprotein E deficiency and type III hyperlipoproteinemia due to a premature stop codon in the apolipoprotein E gene. J Lipid Res. 1992 Nov;33(11):1583–1590. [PubMed] [Google Scholar]
  22. Mabuchi H., Itoh H., Takeda M., Kajinami K., Wakasugi T., Koizumi J., Takeda R., Asagami C. A young type III hyperlipoproteinemic patient associated with apolipoprotein E deficiency. Metabolism. 1989 Feb;38(2):115–119. doi: 10.1016/0026-0495(89)90249-7. [DOI] [PubMed] [Google Scholar]
  23. Mahley R. W. Apolipoprotein E: cholesterol transport protein with expanding role in cell biology. Science. 1988 Apr 29;240(4852):622–630. doi: 10.1126/science.3283935. [DOI] [PubMed] [Google Scholar]
  24. Mahley R. W., Innerarity T. L., Weisgraber K. H., Rall S. C., Jr, Hui D. Y., Lalazar A., Boyles J. K., Taylor J. M., Levy-Wilson B. Cellular and molecular biology of lipoprotein metabolism: characterization of lipoprotein receptor-ligand interactions. Cold Spring Harb Symp Quant Biol. 1986;51(Pt 2):821–828. doi: 10.1101/sqb.1986.051.01.095. [DOI] [PubMed] [Google Scholar]
  25. Mahley R. W., Ji Z. S., Brecht W. J., Miranda R. D., He D. Role of heparan sulfate proteoglycans and the LDL receptor-related protein in remnant lipoprotein metabolism. Ann N Y Acad Sci. 1994 Sep 10;737:39–52. doi: 10.1111/j.1749-6632.1994.tb44300.x. [DOI] [PubMed] [Google Scholar]
  26. McGrory W. J., Bautista D. S., Graham F. L. A simple technique for the rescue of early region I mutations into infectious human adenovirus type 5. Virology. 1988 Apr;163(2):614–617. doi: 10.1016/0042-6822(88)90302-9. [DOI] [PubMed] [Google Scholar]
  27. Mittal S. K., McDermott M. R., Johnson D. C., Prevec L., Graham F. L. Monitoring foreign gene expression by a human adenovirus-based vector using the firefly luciferase gene as a reporter. Virus Res. 1993 Apr;28(1):67–90. doi: 10.1016/0168-1702(93)90090-a. [DOI] [PubMed] [Google Scholar]
  28. Mulder M., Lombardi P., Jansen H., van Berkel T. J., Frants R. R., Havekes L. M. Low density lipoprotein receptor internalizes low density and very low density lipoproteins that are bound to heparan sulfate proteoglycans via lipoprotein lipase. J Biol Chem. 1993 May 5;268(13):9369–9375. [PubMed] [Google Scholar]
  29. Nishina P. M., Verstuyft J., Paigen B. Synthetic low and high fat diets for the study of atherosclerosis in the mouse. J Lipid Res. 1990 May;31(5):859–869. [PubMed] [Google Scholar]
  30. Nykjaer A., Bengtsson-Olivecrona G., Lookene A., Moestrup S. K., Petersen C. M., Weber W., Beisiegel U., Gliemann J. The alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein binds lipoprotein lipase and beta-migrating very low density lipoprotein associated with the lipase. J Biol Chem. 1993 Jul 15;268(20):15048–15055. [PubMed] [Google Scholar]
  31. Paigen B., Ishida B. Y., Verstuyft J., Winters R. B., Albee D. Atherosclerosis susceptibility differences among progenitors of recombinant inbred strains of mice. Arteriosclerosis. 1990 Mar-Apr;10(2):316–323. doi: 10.1161/01.atv.10.2.316. [DOI] [PubMed] [Google Scholar]
  32. Paigen B., Morrow A., Holmes P. A., Mitchell D., Williams R. A. Quantitative assessment of atherosclerotic lesions in mice. Atherosclerosis. 1987 Dec;68(3):231–240. doi: 10.1016/0021-9150(87)90202-4. [DOI] [PubMed] [Google Scholar]
  33. Paik Y. K., Chang D. J., Reardon C. A., Davies G. E., Mahley R. W., Taylor J. M. Nucleotide sequence and structure of the human apolipoprotein E gene. Proc Natl Acad Sci U S A. 1985 May;82(10):3445–3449. doi: 10.1073/pnas.82.10.3445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Piedrahita J. A., Zhang S. H., Hagaman J. R., Oliver P. M., Maeda N. Generation of mice carrying a mutant apolipoprotein E gene inactivated by gene targeting in embryonic stem cells. Proc Natl Acad Sci U S A. 1992 May 15;89(10):4471–4475. doi: 10.1073/pnas.89.10.4471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Plump A. S., Smith J. D., Hayek T., Aalto-Setälä K., Walsh A., Verstuyft J. G., Rubin E. M., Breslow J. L. Severe hypercholesterolemia and atherosclerosis in apolipoprotein E-deficient mice created by homologous recombination in ES cells. Cell. 1992 Oct 16;71(2):343–353. doi: 10.1016/0092-8674(92)90362-g. [DOI] [PubMed] [Google Scholar]
  36. Quantin B., Perricaudet L. D., Tajbakhsh S., Mandel J. L. Adenovirus as an expression vector in muscle cells in vivo. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2581–2584. doi: 10.1073/pnas.89.7.2581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Reue K. L., Quon D. H., O'Donnell K. A., Dizikes G. J., Fareed G. C., Lusis A. J. Cloning and regulation of messenger RNA for mouse apolipoprotein E. J Biol Chem. 1984 Feb 25;259(4):2100–2107. [PubMed] [Google Scholar]
  38. Roessler B. J., Allen E. D., Wilson J. M., Hartman J. W., Davidson B. L. Adenoviral-mediated gene transfer to rabbit synovium in vivo. J Clin Invest. 1993 Aug;92(2):1085–1092. doi: 10.1172/JCI116614. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Rosenfeld M. A., Siegfried W., Yoshimura K., Yoneyama K., Fukayama M., Stier L. E., Päkkö P. K., Gilardi P., Stratford-Perricaudet L. D., Perricaudet M. Adenovirus-mediated transfer of a recombinant alpha 1-antitrypsin gene to the lung epithelium in vivo. Science. 1991 Apr 19;252(5004):431–434. doi: 10.1126/science.2017680. [DOI] [PubMed] [Google Scholar]
  40. Sanger F., Coulson A. R. A rapid method for determining sequences in DNA by primed synthesis with DNA polymerase. J Mol Biol. 1975 May 25;94(3):441–448. doi: 10.1016/0022-2836(75)90213-2. [DOI] [PubMed] [Google Scholar]
  41. Santamarina-Fojo S., Dugi K. A. Structure, function and role of lipoprotein lipase in lipoprotein metabolism. Curr Opin Lipidol. 1994 Apr;5(2):117–125. doi: 10.1097/00041433-199404000-00008. [DOI] [PubMed] [Google Scholar]
  42. Schaefer E. J., Gregg R. E., Ghiselli G., Forte T. M., Ordovas J. M., Zech L. A., Brewer H. B., Jr Familial apolipoprotein E deficiency. J Clin Invest. 1986 Nov;78(5):1206–1219. doi: 10.1172/JCI112704. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Simon R. H., Engelhardt J. F., Yang Y., Zepeda M., Weber-Pendleton S., Grossman M., Wilson J. M. Adenovirus-mediated transfer of the CFTR gene to lung of nonhuman primates: toxicity study. Hum Gene Ther. 1993 Dec;4(6):771–780. doi: 10.1089/hum.1993.4.6-771. [DOI] [PubMed] [Google Scholar]
  44. Smith T. A., Mehaffey M. G., Kayda D. B., Saunders J. M., Yei S., Trapnell B. C., McClelland A., Kaleko M. Adenovirus mediated expression of therapeutic plasma levels of human factor IX in mice. Nat Genet. 1993 Dec;5(4):397–402. doi: 10.1038/ng1293-397. [DOI] [PubMed] [Google Scholar]
  45. Stevenson S. C., Marshall-Neff J., Teng B., Lee C. B., Roy S., McClelland A. Phenotypic correction of hypercholesterolemia in apoE-deficient mice by adenovirus-mediated in vivo gene transfer. Arterioscler Thromb Vasc Biol. 1995 Apr;15(4):479–484. doi: 10.1161/01.atv.15.4.479. [DOI] [PubMed] [Google Scholar]
  46. Stratford-Perricaudet L. D., Levrero M., Chasse J. F., Perricaudet M., Briand P. Evaluation of the transfer and expression in mice of an enzyme-encoding gene using a human adenovirus vector. Hum Gene Ther. 1990 Fall;1(3):241–256. doi: 10.1089/hum.1990.1.3-241. [DOI] [PubMed] [Google Scholar]
  47. Takahashi S., Kawarabayasi Y., Nakai T., Sakai J., Yamamoto T. Rabbit very low density lipoprotein receptor: a low density lipoprotein receptor-like protein with distinct ligand specificity. Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):9252–9256. doi: 10.1073/pnas.89.19.9252. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Thomas K. R., Capecchi M. R. Site-directed mutagenesis by gene targeting in mouse embryo-derived stem cells. Cell. 1987 Nov 6;51(3):503–512. doi: 10.1016/0092-8674(87)90646-5. [DOI] [PubMed] [Google Scholar]
  49. Yang Y., Nunes F. A., Berencsi K., Furth E. E., Gönczöl E., Wilson J. M. Cellular immunity to viral antigens limits E1-deleted adenoviruses for gene therapy. Proc Natl Acad Sci U S A. 1994 May 10;91(10):4407–4411. doi: 10.1073/pnas.91.10.4407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Zannis V. I., Breslow J. L. Characterization of a unique human apolipoprotein E variant associated with type III hyperlipoproteinemia. J Biol Chem. 1980 Mar 10;255(5):1759–1762. [PubMed] [Google Scholar]
  51. Zhang S. H., Reddick R. L., Piedrahita J. A., Maeda N. Spontaneous hypercholesterolemia and arterial lesions in mice lacking apolipoprotein E. Science. 1992 Oct 16;258(5081):468–471. doi: 10.1126/science.1411543. [DOI] [PubMed] [Google Scholar]
  52. de Wet J. R., Wood K. V., DeLuca M., Helinski D. R., Subramani S. Firefly luciferase gene: structure and expression in mammalian cells. Mol Cell Biol. 1987 Feb;7(2):725–737. doi: 10.1128/mcb.7.2.725. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES