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. 1990 Sep 1;172(3):901–909. doi: 10.1084/jem.172.3.901

Cloning of self-major histocompatibility complex antigen-specific suppressor cells from adult bone marrow

PMCID: PMC2188562  PMID: 2143775

Abstract

We examined if suppressor cell clones may be established from adult bone marrow that contains a population of cells capable of specifically downregulating the immune response directed toward self-major histocompatibility complex (MHC) antigens. Freshly prepared adult C3H (H-2k) marrow cells were cultured in medium containing interleukin 2 (IL-2), IL-3, or a mixture of IL-2 and IL-3. After 7-10 d, cells grown in IL-3-containing medium were screened for their capacity to suppress cytotoxic T lymphocyte (CTL) generation against self-MHC antigens in allogeneic mixed lymphocyte cultures. Cells capable of suppressing anti- C3H CTL generation were cloned by limiting dilution. Several suppressor clones were established that exhibited strong suppression of anti-H-2k, anti-H-2Kk/Ik, and anti-H-2Dk CTL generation, but failed to suppress anti-H-2d and anti-H-2b responses. When tested in a skin allograft model, intravenous injections of these bone marrow-derived anti-self suppressor cells (2.5 x 10(7) cells) together with IL-3 induced prolongation of C3H skin allografts in anti-mouse lymphocyte serum- treated B6AF1 mice. Injection of IL-3 alone had no effect on allograft survival. Moreover, these cells failed to prolong B10.AKM skin allografts on B6AF1 recipients. Northern blot analysis showed that these cells express full-length transcripts of the T cell receptor (TCR) gamma gene, but not those of TCR alpha, beta, or delta genes. However, no rearrangement of gamma gene was observed by Southern blot analysis. Flow cytometric analysis revealed that bone marrow-derived suppressor cells are strongly positive for Thy-1 antigen but negative for CD3, CD4 (L3T4), and CD8 (Lyt-2) surface markers, and express only class I MHC antigens. Suppressor cells derived from adult bone marrow may play an important role in extrathymic induction of self-tolerance.

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Selected References

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  1. Chien Y. H., Iwashima M., Wettstein D. A., Kaplan K. B., Elliott J. F., Born W., Davis M. M. T-cell receptor delta gene rearrangements in early thymocytes. Nature. 1987 Dec 24;330(6150):722–727. doi: 10.1038/330722a0. [DOI] [PubMed] [Google Scholar]
  2. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  3. Djeu J. Y., Lanza E., Pastore S., Hapel A. J. Selective growth of natural cytotoxic but not natural killer effector cells in interleukin-3. Nature. 1983 Dec 22;306(5945):788–791. doi: 10.1038/306788a0. [DOI] [PubMed] [Google Scholar]
  4. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  5. Fink P. J., Rammensee H. G., Benedetto J. D., Staerz U. D., Lefrancois L., Bevan M. J. Studies on the mechanism of suppression of primary cytotoxic responses by cloned cytotoxic T lymphocytes. J Immunol. 1984 Oct;133(4):1769–1774. [PubMed] [Google Scholar]
  6. Fink P. J., Rammensee H. G., Bevan M. J. Cloned cytolytic T cells can suppress primary cytotoxic responses directed against them. J Immunol. 1984 Oct;133(4):1775–1781. [PubMed] [Google Scholar]
  7. Fink P. J., Shimonkevitz R. P., Bevan M. J. Veto cells. Annu Rev Immunol. 1988;6:115–137. doi: 10.1146/annurev.iy.06.040188.000555. [DOI] [PubMed] [Google Scholar]
  8. Glisin V., Crkvenjakov R., Byus C. Ribonucleic acid isolated by cesium chloride centrifugation. Biochemistry. 1974 Jun 4;13(12):2633–2637. doi: 10.1021/bi00709a025. [DOI] [PubMed] [Google Scholar]
  9. Hurwitz J. L., Samaridis J., Pelkonen J. Progression of rearrangements at T cell receptor beta and gamma gene loci during athymic differentiation of bone marrow cells in vitro. Cell. 1988 Mar 25;52(6):821–829. doi: 10.1016/0092-8674(88)90424-2. [DOI] [PubMed] [Google Scholar]
  10. Ihle J. N., Keller J., Oroszlan S., Henderson L. E., Copeland T. D., Fitch F., Prystowsky M. B., Goldwasser E., Schrader J. W., Palaszynski E. Biologic properties of homogeneous interleukin 3. I. Demonstration of WEHI-3 growth factor activity, mast cell growth factor activity, p cell-stimulating factor activity, colony-stimulating factor activity, and histamine-producing cell-stimulating factor activity. J Immunol. 1983 Jul;131(1):282–287. [PubMed] [Google Scholar]
  11. Ihle J. N., Pepersack L., Rebar L. Regulation of T cell differentiation: in vitro induction of 20 alpha-hydroxysteroid dehydrogenase in splenic lymphocytes from athymic mice by a unique lymphokine. J Immunol. 1981 Jun;126(6):2184–2189. [PubMed] [Google Scholar]
  12. Itohara S., Nakanishi N., Kanagawa O., Kubo R., Tonegawa S. Monoclonal antibodies specific to native murine T-cell receptor gamma delta: analysis of gamma delta T cells during thymic ontogeny and in peripheral lymphoid organs. Proc Natl Acad Sci U S A. 1989 Jul;86(13):5094–5098. doi: 10.1073/pnas.86.13.5094. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jadus M. R., Parkman R. The selective growth of murine newborn-derived suppressor cells and their probable mode of action. J Immunol. 1986 Feb 1;136(3):783–792. [PubMed] [Google Scholar]
  14. Jadus M. R., Schmunk G., Djeu J. Y., Parkman R. Morphology and lytic mechanisms of interleukin 3-dependent natural cytotoxic cells: tumor necrosis factor as a possible mediator. J Immunol. 1986 Nov 1;137(9):2774–2783. [PubMed] [Google Scholar]
  15. Kappler J. W., Roehm N., Marrack P. T cell tolerance by clonal elimination in the thymus. Cell. 1987 Apr 24;49(2):273–280. doi: 10.1016/0092-8674(87)90568-x. [DOI] [PubMed] [Google Scholar]
  16. Kisielow P., Blüthmann H., Staerz U. D., Steinmetz M., von Boehmer H. Tolerance in T-cell-receptor transgenic mice involves deletion of nonmature CD4+8+ thymocytes. Nature. 1988 Jun 23;333(6175):742–746. doi: 10.1038/333742a0. [DOI] [PubMed] [Google Scholar]
  17. Kronenberg M., Siu G., Hood L. E., Shastri N. The molecular genetics of the T-cell antigen receptor and T-cell antigen recognition. Annu Rev Immunol. 1986;4:529–591. doi: 10.1146/annurev.iy.04.040186.002525. [DOI] [PubMed] [Google Scholar]
  18. Maki T., Gottschalk R., Wood M. L., Monaco A. P. Specific unresponsiveness to skin allografts in anti-lymphocyte serum-treated, marrow-injected mice: participation of donor marrow-derived suppressor T cells. J Immunol. 1981 Oct;127(4):1433–1438. [PubMed] [Google Scholar]
  19. Maki T., Satomi S., Gotoh M., Monaco A. P. Contra-IL 2; a suppressor lymphokine that inhibits IL 2 activity. J Immunol. 1986 May 1;136(9):3298–3303. [PubMed] [Google Scholar]
  20. Miller R. G. An immunological suppressor cell inactivating cytotoxic T-lymphocyte precursor cells recognizing it. Nature. 1980 Oct 9;287(5782):544–546. doi: 10.1038/287544a0. [DOI] [PubMed] [Google Scholar]
  21. Miller R. G., Derry H. A cell population in nu/nu spleen can prevent generation of cytotoxic lymphocytes by normal spleen cells against self antigens of the nu/nu spleen. J Immunol. 1979 Apr;122(4):1502–1509. [PubMed] [Google Scholar]
  22. Miller R. G., Muraoka S., Claesson M. H., Reimann J., Benveniste P. The veto phenomenon in T-cell regulation. Ann N Y Acad Sci. 1988;532:170–176. doi: 10.1111/j.1749-6632.1988.tb36336.x. [DOI] [PubMed] [Google Scholar]
  23. Monaco A. P., Wood M. L. Studies on heterologous antilymphocyte serum in mice. VII. Optimal cellular antigen for induction of immunologic tolerance with antilymphocyte serum. Transplant Proc. 1970 Dec;2(4):489–496. [PubMed] [Google Scholar]
  24. Muraoka S., Miller R. G. Cells in bone marrow and in T cell colonies grown from bone marrow can suppress generation of cytotoxic T lymphocytes directed against their self antigens. J Exp Med. 1980 Jul 1;152(1):54–71. doi: 10.1084/jem.152.1.54. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pope B. L. Secretion of a suppressor cell-inducing factor by an interleukin 3-dependent cell line with natural cytotoxic activity. II. Range, potency, and kinetics of suppressive activity. Immunobiology. 1987 Jan;174(1):107–118. doi: 10.1016/S0171-2985(87)80088-8. [DOI] [PubMed] [Google Scholar]
  26. Rammensee H. G., Fink P. J., Bevan M. J. Functional clonal deletion of class I-specific cytotoxic T lymphocytes by veto cells that express antigen. J Immunol. 1984 Nov;133(5):2390–2396. [PubMed] [Google Scholar]
  27. Saito H., Kranz D. M., Takagaki Y., Hayday A. C., Eisen H. N., Tonegawa S. Complete primary structure of a heterodimeric T-cell receptor deduced from cDNA sequences. 1984 Jun 28-Jul 4Nature. 309(5971):757–762. doi: 10.1038/309757a0. [DOI] [PubMed] [Google Scholar]
  28. Sprent J., Webb S. R. Function and specificity of T cell subsets in the mouse. Adv Immunol. 1987;41:39–133. doi: 10.1016/s0065-2776(08)60030-9. [DOI] [PubMed] [Google Scholar]
  29. Takagaki Y., Nakanishi N., Ishida I., Kanagawa O., Tonegawa S. T cell receptor-gamma and -delta genes preferentially utilized by adult thymocytes for the surface expression. J Immunol. 1989 Mar 15;142(6):2112–2121. [PubMed] [Google Scholar]
  30. Weinstein Y., Morishita K., Cleveland J. L., Ihle J. N. Interleukin 3 (IL-3) induces transcription from nonrearranged T cell receptor gamma loci in IL-3-dependent cell lines. J Exp Med. 1989 Jun 1;169(6):2059–2071. doi: 10.1084/jem.169.6.2059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wood M. L., Monaco A. P. Suppressor cells in specific unresponsiveness to skin allografts in ALS-treated, marrow-injected mice. Transplantation. 1980 Mar;29(3):196–200. doi: 10.1097/00007890-198003000-00006. [DOI] [PubMed] [Google Scholar]
  32. Yoshikai Y., Takeda Y., Ohga S., Kishihara K., Matsuzaki G., Nomoto K. Rearrangements of T-cell antigen receptor gamma and delta chain genes are detected in the long-term cultured bone marrow cells of athymic nude mice but not in those of euthymic mice. Immunology. 1989 Apr;66(4):512–516. [PMC free article] [PubMed] [Google Scholar]

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