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. 1988 Sep;8(9):3864–3871. doi: 10.1128/mcb.8.9.3864

Generation of murine stromal cell lines supporting hematopoietic stem cell proliferation by use of recombinant retrovirus vectors encoding simian virus 40 large T antigen.

D A Williams 1, M F Rosenblatt 1, D R Beier 1, R D Cone 1
PMCID: PMC365445  PMID: 2851729

Abstract

The bone marrow is a complex microenvironment made up of multiple cell types which appears to play an important role in the maintenance of hematopoietic stem cell self-renewal and proliferation. We used murine long-term marrow cultures and a defective recombinant retrovirus vector containing the simian virus 40 large T antigen to immortalize marrow stromal cells which can support hematopoiesis in vitro for up to 5 weeks. Such cloned cell lines differentially supported stem cells which, when transplanted, allowed survival of lethally irradiated mice, formed hematopoietic spleen colonies in vivo, and stimulated lymphocyte proliferation in vitro. Molecular and functional analyses of these cell lines did not demonstrate the production of any growth factors known to support the proliferation of primitive hematopoietic stem cells. All cell lines examined produced macrophage colony-stimulating factor. The use of immortalizing retrovirus vectors may allow determination of unique cellular proteins important in hematopoietic stem cell proliferation by the systematic comparison of stromal cells derived from a variety of murine tissues.

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Selected References

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

  1. Anklesaria P., Kase K., Glowacki J., Holland C. A., Sakakeeny M. A., Wright J. A., FitzGerald T. J., Lee C. Y., Greenberger J. S. Engraftment of a clonal bone marrow stromal cell line in vivo stimulates hematopoietic recovery from total body irradiation. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7681–7685. doi: 10.1073/pnas.84.21.7681. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bentley S. A. The role and composition of the adherent layer in long-term bone marrow culture. Kroc Found Ser. 1984;18:141–156. [PubMed] [Google Scholar]
  3. Bradley T. R., Metcalf D. The growth of mouse bone marrow cells in vitro. Aust J Exp Biol Med Sci. 1966 Jun;44(3):287–299. doi: 10.1038/icb.1966.28. [DOI] [PubMed] [Google Scholar]
  4. Campbell A. D., Long M. W., Wicha M. S. Haemonectin, a bone marrow adhesion protein specific for cells of granulocyte lineage. Nature. 1987 Oct 22;329(6141):744–746. doi: 10.1038/329744a0. [DOI] [PubMed] [Google Scholar]
  5. Cepko C. L., Roberts B. E., Mulligan R. C. Construction and applications of a highly transmissible murine retrovirus shuttle vector. Cell. 1984 Jul;37(3):1053–1062. doi: 10.1016/0092-8674(84)90440-9. [DOI] [PubMed] [Google Scholar]
  6. Chang S. E. In vitro transformation of human epithelial cells. Biochim Biophys Acta. 1986;823(3):161–194. doi: 10.1016/0304-419x(86)90001-6. [DOI] [PubMed] [Google Scholar]
  7. Cone R. D., Grodzicker T., Jaramillo M. A retrovirus expressing the 12S adenoviral E1A gene product can immortalize epithelial cells from a broad range of rat tissues. Mol Cell Biol. 1988 Mar;8(3):1036–1044. doi: 10.1128/mcb.8.3.1036. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dexter T. M., Allen T. D., Lajtha L. G., Schofield R., Lord B. I. Stimulation of differentiation and proliferation of haemopoietic cells in vitro. J Cell Physiol. 1973 Dec;82(3):461–473. doi: 10.1002/jcp.1040820315. [DOI] [PubMed] [Google Scholar]
  9. Dexter T. M., Moore M. A. In vitro duplication and "cure" of haemopoietic defects in genetically anaemic mice. Nature. 1977 Sep 29;269(5627):412–414. doi: 10.1038/269412a0. [DOI] [PubMed] [Google Scholar]
  10. Dick J. E., Magli M. C., Huszar D., Phillips R. A., Bernstein A. Introduction of a selectable gene into primitive stem cells capable of long-term reconstitution of the hemopoietic system of W/Wv mice. Cell. 1985 Aug;42(1):71–79. doi: 10.1016/s0092-8674(85)80102-1. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Ferrara J. L., Mauch P., McIntyre J., Michaelson J., Burakoff S. J. Engraftment following T-cell-depleted bone marrow transplantation. II. Stability of mixed chimerism in semiallogeneic recipients after total-body irradiation. Transplantation. 1987 Oct;44(4):495–499. doi: 10.1097/00007890-198710000-00007. [DOI] [PubMed] [Google Scholar]
  13. Fung M. C., Hapel A. J., Ymer S., Cohen D. R., Johnson R. M., Campbell H. D., Young I. G. Molecular cloning of cDNA for murine interleukin-3. Nature. 1984 Jan 19;307(5948):233–237. doi: 10.1038/307233a0. [DOI] [PubMed] [Google Scholar]
  14. Gluzman Y. SV40-transformed simian cells support the replication of early SV40 mutants. Cell. 1981 Jan;23(1):175–182. doi: 10.1016/0092-8674(81)90282-8. [DOI] [PubMed] [Google Scholar]
  15. Golde D. W., Hocking W. G., Quan S. G., Sparkes R. S., Gale R. P. Origin of human bone marrow fibroblasts. Br J Haematol. 1980 Feb;44(2):183–187. doi: 10.1111/j.1365-2141.1980.tb01200.x. [DOI] [PubMed] [Google Scholar]
  16. Gordon M. Y., Dowding C. R., Riley G. P., Goldman J. M., Greaves M. F. Altered adhesive interactions with marrow stroma of haematopoietic progenitor cells in chronic myeloid leukaemia. Nature. 1987 Jul 23;328(6128):342–344. doi: 10.1038/328342a0. [DOI] [PubMed] [Google Scholar]
  17. Gordon M. Y., Riley G. P., Watt S. M., Greaves M. F. Compartmentalization of a haematopoietic growth factor (GM-CSF) by glycosaminoglycans in the bone marrow microenvironment. 1987 Mar 26-Apr 1Nature. 326(6111):403–405. doi: 10.1038/326403a0. [DOI] [PubMed] [Google Scholar]
  18. Gough N. M., Gough J., Metcalf D., Kelso A., Grail D., Nicola N. A., Burgess A. W., Dunn A. R. Molecular cloning of cDNA encoding a murine haematopoietic growth regulator, granulocyte-macrophage colony stimulating factor. 1984 Jun 28-Jul 4Nature. 309(5971):763–767. doi: 10.1038/309763a0. [DOI] [PubMed] [Google Scholar]
  19. Gray P. W., Glaister D., Chen E., Goeddel D. V., Pennica D. Two interleukin 1 genes in the mouse: cloning and expression of the cDNA for murine interleukin 1 beta. J Immunol. 1986 Dec 1;137(11):3644–3648. [PubMed] [Google Scholar]
  20. Greenberger J. S., Sakakeeny M., Parker L. M. In vitro proliferation of hemopoietic stem cells in long-term marrow cultures: principles in mouse applied to man. Exp Hematol. 1979;7 (Suppl 5):135–148. [PubMed] [Google Scholar]
  21. Harlow E., Crawford L. V., Pim D. C., Williamson N. M. Monoclonal antibodies specific for simian virus 40 tumor antigens. J Virol. 1981 Sep;39(3):861–869. doi: 10.1128/jvi.39.3.861-869.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Harlow E., Pim D. C., Crawford L. V. Complex of simian virus 40 large-T antigen and host 53,000-molecular-weight protein in monkey cells. J Virol. 1981 Feb;37(2):564–573. doi: 10.1128/jvi.37.2.564-573.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hunt P., Robertson D., Weiss D., Rennick D., Lee F., Witte O. N. A single bone marrow-derived stromal cell type supports the in vitro growth of early lymphoid and myeloid cells. Cell. 1987 Mar 27;48(6):997–1007. doi: 10.1016/0092-8674(87)90708-2. [DOI] [PubMed] [Google Scholar]
  24. Iscove N. N., Sieber F., Winterhalter K. H. Erythroid colony formation in cultures of mouse and human bone marrow: analysis of the requirement for erythropoietin by gel filtration and affinity chromatography on agarose-concanavalin A. J Cell Physiol. 1974 Apr;83(2):309–320. doi: 10.1002/jcp.1040830218. [DOI] [PubMed] [Google Scholar]
  25. Jat P. S., Cepko C. L., Mulligan R. C., Sharp P. A. Recombinant retroviruses encoding simian virus 40 large T antigen and polyomavirus large and middle T antigens. Mol Cell Biol. 1986 Apr;6(4):1204–1217. doi: 10.1128/mcb.6.4.1204. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Jat P. S., Sharp P. A. Large T antigens of simian virus 40 and polyomavirus efficiently establish primary fibroblasts. J Virol. 1986 Sep;59(3):746–750. doi: 10.1128/jvi.59.3.746-750.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kawasaki E. S., Ladner M. B., Wang A. M., Van Arsdell J., Warren M. K., Coyne M. Y., Schweickart V. L., Lee M. T., Wilson K. J., Boosman A. Molecular cloning of a complementary DNA encoding human macrophage-specific colony-stimulating factor (CSF-1). Science. 1985 Oct 18;230(4723):291–296. doi: 10.1126/science.2996129. [DOI] [PubMed] [Google Scholar]
  28. Keller G., Paige C., Gilboa E., Wagner E. F. Expression of a foreign gene in myeloid and lymphoid cells derived from multipotent haematopoietic precursors. Nature. 1985 Nov 14;318(6042):149–154. doi: 10.1038/318149a0. [DOI] [PubMed] [Google Scholar]
  29. Kodama H., Sudo H., Koyama H., Kasai S., Yamamoto S. In vitro hemopoiesis within a microenvironment created by MC3T3-G2/PA6 preadipocytes. J Cell Physiol. 1984 Mar;118(3):233–240. doi: 10.1002/jcp.1041180303. [DOI] [PubMed] [Google Scholar]
  30. Lane D. P., Crawford L. V. T antigen is bound to a host protein in SV40-transformed cells. Nature. 1979 Mar 15;278(5701):261–263. doi: 10.1038/278261a0. [DOI] [PubMed] [Google Scholar]
  31. Lanotte M., Metcalf D., Dexter T. M. Production of monocyte/macrophage colony-stimulating factor by preadipocyte cell lines derived from murine marrow stroma. J Cell Physiol. 1982 Jul;112(1):123–127. doi: 10.1002/jcp.1041120118. [DOI] [PubMed] [Google Scholar]
  32. Lemischka I. R., Raulet D. H., Mulligan R. C. Developmental potential and dynamic behavior of hematopoietic stem cells. Cell. 1986 Jun 20;45(6):917–927. doi: 10.1016/0092-8674(86)90566-0. [DOI] [PubMed] [Google Scholar]
  33. Lennon J. E., Micklem H. S. Stromal cells in long-term murine bone marrow culture: FACS studies and origin of stromal cells in radiation chimeras. Exp Hematol. 1986 May;14(4):287–292. [PubMed] [Google Scholar]
  34. Li C. L., Johnson G. R. Stimulation of multipotential, erythroid and other murine haematopoietic progenitor cells by adherent cell lines in the absence of detectable multi-CSF (IL-3). Nature. 1985 Aug 15;316(6029):633–636. doi: 10.1038/316633a0. [DOI] [PubMed] [Google Scholar]
  35. Lim B., Williams D. A., Orkin S. H. Retrovirus-mediated gene transfer of human adenosine deaminase: expression of functional enzyme in murine hematopoietic stem cells in vivo. Mol Cell Biol. 1987 Oct;7(10):3459–3465. doi: 10.1128/mcb.7.10.3459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Linzer D. I., Levine A. J. Characterization of a 54K dalton cellular SV40 tumor antigen present in SV40-transformed cells and uninfected embryonal carcinoma cells. Cell. 1979 May;17(1):43–52. doi: 10.1016/0092-8674(79)90293-9. [DOI] [PubMed] [Google Scholar]
  37. Lomedico P. T., Gubler U., Hellmann C. P., Dukovich M., Giri J. G., Pan Y. C., Collier K., Semionow R., Chua A. O., Mizel S. B. Cloning and expression of murine interleukin-1 cDNA in Escherichia coli. 1984 Nov 29-Dec 5Nature. 312(5993):458–462. doi: 10.1038/312458a0. [DOI] [PubMed] [Google Scholar]
  38. McCulloch E. A., Siminovitch L., Till J. E., Russell E. S., Bernstein S. E. The cellular basis of the genetically determined hemopoietic defect in anemic mice of genotype Sl-Sld. Blood. 1965 Oct;26(4):399–410. [PubMed] [Google Scholar]
  39. Miller A. D., Ong E. S., Rosenfeld M. G., Verma I. M., Evans R. M. Infectious and selectable retrovirus containing an inducible rat growth hormone minigene. Science. 1984 Sep 7;225(4666):993–998. doi: 10.1126/science.6089340. [DOI] [PubMed] [Google Scholar]
  40. Paucha E., Kalderon D., Harvey R. W., Smith A. E. Simian virus 40 origin DNA-binding domain on large T antigen. J Virol. 1986 Jan;57(1):50–64. doi: 10.1128/jvi.57.1.50-64.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Perkins S., Fleischman R. A. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice. J Clin Invest. 1988 Apr;81(4):1072–1080. doi: 10.1172/JCI113419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Piersma A. H., Ploemacher R. E., Brockbank K. G. Transplantation of bone marrow fibroblastoid stromal cells in mice via the intravenous route. Br J Haematol. 1983 Jun;54(2):285–290. doi: 10.1111/j.1365-2141.1983.tb02097.x. [DOI] [PubMed] [Google Scholar]
  43. Rennick D., Yang G., Gemmell L., Lee F. Control of hemopoiesis by a bone marrow stromal cell clone: lipopolysaccharide- and interleukin-1-inducible production of colony-stimulating factors. Blood. 1987 Feb;69(2):682–691. [PubMed] [Google Scholar]
  44. Roberts R. A., Spooncer E., Parkinson E. K., Lord B. I., Allen T. D., Dexter T. M. Metabolically inactive 3T3 cells can substitute for marrow stromal cells to promote the proliferation and development of multipotent haemopoietic stem cells. J Cell Physiol. 1987 Aug;132(2):203–214. doi: 10.1002/jcp.1041320204. [DOI] [PubMed] [Google Scholar]
  45. Spooncer E., Heyworth C. M., Dunn A., Dexter T. M. Self-renewal and differentiation of interleukin-3-dependent multipotent stem cells are modulated by stromal cells and serum factors. Differentiation. 1986;31(2):111–118. doi: 10.1111/j.1432-0436.1986.tb00391.x. [DOI] [PubMed] [Google Scholar]
  46. Tsuchiya M., Asano S., Kaziro Y., Nagata S. Isolation and characterization of the cDNA for murine granulocyte colony-stimulating factor. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7633–7637. doi: 10.1073/pnas.83.20.7633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Werts E. D., Gibson D. P., Knapp S. A., DeGowin R. L. Stromal cell migration precedes hemopoietic repopulation of the bone marrow after irradiation. Radiat Res. 1980 Jan;81(1):20–30. [PubMed] [Google Scholar]
  48. Whitlock C. A., Robertson D., Witte O. N. Murine B cell lymphopoiesis in long term culture. J Immunol Methods. 1984 Mar 16;67(2):353–369. doi: 10.1016/0022-1759(84)90475-7. [DOI] [PubMed] [Google Scholar]
  49. Whitlock C. A., Tidmarsh G. F., Muller-Sieburg C., Weissman I. L. Bone marrow stromal cell lines with lymphopoietic activity express high levels of a pre-B neoplasia-associated molecule. Cell. 1987 Mar 27;48(6):1009–1021. doi: 10.1016/0092-8674(87)90709-4. [DOI] [PubMed] [Google Scholar]
  50. 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]
  51. Yasukawa K., Hirano T., Watanabe Y., Muratani K., Matsuda T., Nakai S., Kishimoto T. Structure and expression of human B cell stimulatory factor-2 (BSF-2/IL-6) gene. EMBO J. 1987 Oct;6(10):2939–2945. doi: 10.1002/j.1460-2075.1987.tb02598.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Zipori D., Sasson T., Frenkel A. Myelopoiesis in the presence of stromal cells from mouse bone marrow: I. Monosaccharides regulate colony formation. Exp Hematol. 1981 Jul;9(6):656–663. [PubMed] [Google Scholar]
  53. Zuckerman K. S., Prince C. W., Ribadeneira M. Sl/Sld mouse bone marrow stroma in vitro contains an active radiation-sensitive inhibitor of normal hemopoiesis. Blood. 1986 Dec;68(6):1201–1206. [PubMed] [Google Scholar]

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