Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1987 May;84(10):3344–3348. doi: 10.1073/pnas.84.10.3344

Expression of retrovirally transduced genes in primary cultures of adult rat hepatocytes.

J A Wolff, J K Yee, H F Skelly, J C Moores, J G Respess, T Friedmann, H Leffert
PMCID: PMC304866  PMID: 3033644

Abstract

Differentiated primary rat hepatocyte cultures have been infected with retroviral vectors expressing human hypoxanthine/guanine phosphoribosyltransferase or the transposon Tn5 neomycin-resistance gene. Expression of the markers was detected only after infection of the cells during a short period of cell replication and transient dedifferentiation from days 1 to 5 of culture. Provirus integrated during that period remains fully expressed during the entire subsequent stationary period of culture up to at least 25 days. Selection with the neomycin analogue G418 of cells infected with the neomycin vector led to the appearance of cells with hepatocyte morphology in which newly synthesized albumin was detectable by immunoprecipitation, indicating successful infection of hepatocytes.

Full text

PDF
3344

Images in this article

3346Image
on p.3346
3346Image
on p.3346
3346Image
on p.3346
3347Image
on p.3347
3347Image
on p.3347
3347Image
on p.3347

Selected References

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

  1. Anderson W. F. Prospects for human gene therapy. Science. 1984 Oct 26;226(4673):401–409. doi: 10.1126/science.6093246. [DOI] [PubMed] [Google Scholar]
  2. Brown J. W., Lad P. J., Skelly H., Koch K. S., Lin M., Leffert H. Expression of differentiated function by hepatocytes in primary culture: variable effects of glucagon and insulin on gluconeogenesis during cell growth. Differentiation. 1983;25(2):176–184. doi: 10.1111/j.1432-0436.1984.tb01353.x. [DOI] [PubMed] [Google Scholar]
  3. De Larco J. E., Rapp U. R., Todaro G. J. Cell surface receptors for ecotropic MuLV: detection and tissue distributions of free receptors in vivo. Int J Cancer. 1978 Mar 15;21(3):356–360. doi: 10.1002/ijc.2910210317. [DOI] [PubMed] [Google Scholar]
  4. Demetriou A. A., Whiting J. F., Feldman D., Levenson S. M., Chowdhury N. R., Moscioni A. D., Kram M., Chowdhury J. R. Replacement of liver function in rats by transplantation of microcarrier-attached hepatocytes. Science. 1986 Sep 12;233(4769):1190–1192. doi: 10.1126/science.2426782. [DOI] [PubMed] [Google Scholar]
  5. Eglitis M. A., Kantoff P., Gilboa E., Anderson W. F. Gene expression in mice after high efficiency retroviral-mediated gene transfer. Science. 1985 Dec 20;230(4732):1395–1398. doi: 10.1126/science.2999985. [DOI] [PubMed] [Google Scholar]
  6. Friedman J. M., Babiss L. E., Clayton D. F., Darnell J. E., Jr Cellular promoters incorporated into the adenovirus genome: cell specificity of albumin and immunoglobulin expression. Mol Cell Biol. 1986 Nov;6(11):3791–3797. doi: 10.1128/mcb.6.11.3791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Georgoff I., Secott T., Isom H. C. Effect of simian virus 40 infection on albumin production by hepatocytes cultured in chemically defined medium and plated on collagen and non-collagen attachment surfaces. J Biol Chem. 1984 Aug 10;259(15):9595–9602. [PubMed] [Google Scholar]
  8. Isom H. C., Georgoff I. Quantitative assay for albumin-producing liver cells after simian virus 40 transformation of rat hepatocytes maintained in chemically defined medium. Proc Natl Acad Sci U S A. 1984 Oct;81(20):6378–6382. doi: 10.1073/pnas.81.20.6378. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Isom H. C., Liao W. S., Taylor J. M., Willwerth G. E., Eadline T. S. Rapid and selective shutoff of plasma protein production in herpes simplex virus type 2-infected hepatoma cells. Virology. 1983 Apr 30;126(2):548–562. doi: 10.1016/s0042-6822(83)80012-9. [DOI] [PubMed] [Google Scholar]
  10. Jaenisch R., Fan H., Croker B. Infection of preimplantation mouse embryos and of newborn mice with leukemia virus: tissue distribution of viral DNA and RNA and leukemogenesis in the adult animal. Proc Natl Acad Sci U S A. 1975 Oct;72(10):4008–4012. doi: 10.1073/pnas.72.10.4008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Jaenisch R., Hoffmann E. Transcription of endogenous C-type viruses in resting and proliferating tissues of BALB/Mo mice. Virology. 1979 Oct 30;98(2):289–297. doi: 10.1016/0042-6822(79)90552-x. [DOI] [PubMed] [Google Scholar]
  12. Jaenisch R. Retroviruses and embryogenesis: microinjection of Moloney leukemia virus into midgestation mouse embryos. Cell. 1980 Jan;19(1):181–188. doi: 10.1016/0092-8674(80)90399-2. [DOI] [PubMed] [Google Scholar]
  13. Johnson G. G., Eisenberg L. R., Migeon B. R. Human and mouse hypoxanthine-guanine phosphoribosyltransferase: dimers and tetramers. Science. 1979 Jan 12;203(4376):174–176. doi: 10.1126/science.569362. [DOI] [PubMed] [Google Scholar]
  14. Joyner A., Keller G., Phillips R. A., Bernstein A. Retrovirus transfer of a bacterial gene into mouse haematopoietic progenitor cells. Nature. 1983 Oct 6;305(5934):556–558. doi: 10.1038/305556a0. [DOI] [PubMed] [Google Scholar]
  15. Kato N., Sahm H., Schütte H., Wagner F. Purification and properties of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase from a methanol-utilizing yeast, Candida boidinii. Biochim Biophys Acta. 1979 Jan 12;566(1):1–11. doi: 10.1016/0005-2744(79)90242-0. [DOI] [PubMed] [Google Scholar]
  16. Koch K. S., Leffert H. L. Growth control of differentiated adult rat hepatocytes in primary culture. Ann N Y Acad Sci. 1980;349:111–127. doi: 10.1111/j.1749-6632.1980.tb29520.x. [DOI] [PubMed] [Google Scholar]
  17. Koch K. S., Leffert H. L. Increased sodium ion influx is necessary to initiate rat hepatocyte proliferation. Cell. 1979 Sep;18(1):153–163. doi: 10.1016/0092-8674(79)90364-7. [DOI] [PubMed] [Google Scholar]
  18. Kruijer W., Skelly H., Botteri F., van der Putten H., Barber J. R., Verma I. M., Leffert H. L. Proto-oncogene expression in regenerating liver is simulated in cultures of primary adult rat hepatocytes. J Biol Chem. 1986 Jun 15;261(17):7929–7933. [PubMed] [Google Scholar]
  19. Lad P. J., Shier W. T., Skelly H., De Hemptinne B., Leffert H. L. Adult rat hepatocytes in primary culture. VI. Developmental changes in alcohol dehydrogenase activity and ethanol conversion during the growth cycle. Alcohol Clin Exp Res. 1982 Winter;6(1):64–71. doi: 10.1111/j.1530-0277.1982.tb05382.x. [DOI] [PubMed] [Google Scholar]
  20. Lad P. J., Shier W. T., Skelly H., de Hemptinne B., Leffert H. L. Adult rat hepatocytes in primary culture. VII. Proliferative and functional properties of cells from ethanol-intoxicated animals: evidence for a reversible albumin 'production defect'. Alcohol Clin Exp Res. 1982 Winter;6(1):72–79. doi: 10.1111/j.1530-0277.1982.tb05383.x. [DOI] [PubMed] [Google Scholar]
  21. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  22. Leffert H. L., Moran T., Boorstein R., Koch K. S. Procarcinogen activation and hormonal control of cell proliferation in differentiated primary adult rat liver cell cultures. Nature. 1977 May 5;267(5606):58–61. doi: 10.1038/267058a0. [DOI] [PubMed] [Google Scholar]
  23. Leffert H., Moran T., Sell S., Skelly H., Ibsen K., Mueller M., Arias I. Growth state-dependent phenotypes of adult hepatocytes in primary monolayer culture. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1834–1838. doi: 10.1073/pnas.75.4.1834. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Linney E., Davis B., Overhauser J., Chao E., Fan H. Non-function of a Moloney murine leukaemia virus regulatory sequence in F9 embryonal carcinoma cells. 1984 Mar 29-Apr 4Nature. 308(5958):470–472. doi: 10.1038/308470a0. [DOI] [PubMed] [Google Scholar]
  25. Miller A. D., Eckner R. J., Jolly D. J., Friedmann T., Verma I. M. Expression of a retrovirus encoding human HPRT in mice. Science. 1984 Aug 10;225(4662):630–632. doi: 10.1126/science.6377498. [DOI] [PubMed] [Google Scholar]
  26. Miller A. D., Law M. F., Verma I. M. Generation of helper-free amphotropic retroviruses that transduce a dominant-acting, methotrexate-resistant dihydrofolate reductase gene. Mol Cell Biol. 1985 Mar;5(3):431–437. doi: 10.1128/mcb.5.3.431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Miller A. D., Law M. F., Verma I. M. Generation of helper-free amphotropic retroviruses that transduce a dominant-acting, methotrexate-resistant dihydrofolate reductase gene. Mol Cell Biol. 1985 Mar;5(3):431–437. doi: 10.1128/mcb.5.3.431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Schlegel-Haueter S. E., Schlegel W., Chou J. Y. Establishment of a fetal rat liver cell line that retains differentiated liver functions. Proc Natl Acad Sci U S A. 1980 May;77(5):2731–2734. doi: 10.1073/pnas.77.5.2731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sherr C. J., Todaro G. J. Purification and assay of murine leukemia viruses. Methods Enzymol. 1979;58:412–424. doi: 10.1016/s0076-6879(79)58156-7. [DOI] [PubMed] [Google Scholar]
  30. Stuhlmann H., Cone R., Mulligan R. C., Jaenisch R. Introduction of a selectable gene into different animal tissue by a retrovirus recombinant vector. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7151–7155. doi: 10.1073/pnas.81.22.7151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Tur-Kaspa R., Teicher L., Levine B. J., Skoultchi A. I., Shafritz D. A. Use of electroporation to introduce biologically active foreign genes into primary rat hepatocytes. Mol Cell Biol. 1986 Feb;6(2):716–718. doi: 10.1128/mcb.6.2.716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Williams D. A., Lemischka I. R., Nathan D. G., Mulligan R. C. Introduction of new genetic material into pluripotent haematopoietic stem cells of the mouse. Nature. 1984 Aug 9;310(5977):476–480. doi: 10.1038/310476a0. [DOI] [PubMed] [Google Scholar]
  33. Willis R. C., Jolly D. J., Miller A. D., Plent M. M., Esty A. C., Anderson P. J., Chang H. C., Jones O. W., Seegmiller J. E., Friedmann T. Partial phenotypic correction of human Lesch-Nyhan (hypoxanthine-guanine phosphoribosyltransferase-deficient) lymphoblasts with a transmissible retroviral vector. J Biol Chem. 1984 Jun 25;259(12):7842–7849. [PubMed] [Google Scholar]
  34. Yee J. K., Jolly D. J., Miller A. D., Willis R., Wolff J., Friedmann T. Epitope insertion into the human hypoxanthine phosphoribosyltransferase protein and detection of the mutant protein by an anti-peptide antibody. Gene. 1987;53(1):97–104. doi: 10.1016/0378-1119(87)90096-5. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES