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. 1977 Nov 1;146(5):1420–1435. doi: 10.1084/jem.146.5.1420

Regulation of B-lymphocyte clonal proliferation by stimulatory and inhibitory macrophage-derived factors

JI Kurland, PW Kincade, MAS Moore
PMCID: PMC2180959  PMID: 303681

Abstract

A functional subpopulation of murine B lymphocytes proliferate in semisolid agar culture in the presence of 2-mercaptoethanol to form colonies. The effects of diffusible macrophage-derived factors on this focal proliferation was investigated using a two-layer culture system which prevented macrophage-lymphocyte contact and permitted B-cell activation to be critically assessed under conditions of extremely low cell densities. Adherent peritoneal macrophages incorporated within underlayers of spleen or lymph node cell cultures potentiated both the number and size of developing B-cell colonies. These effects were most striking when low numbers of spleen or lymph node cells, or macrophage- depleted lymphoid cell suspensions were used. Thus, macrophage-depleted lymph node ceils gave rise to virtually no colonies, but colony-forming ability was restored by the presence of an optimal number of macrophages. When the number of macrophages exceeded that required for optimal stimulation, colony formation was suppressed; an effect which was largely prevented by indomethacin, an inhibitor of prostaglandin synthesis. Under these conditions, stimulation and inhibition of B-cell activation by macrophages could be dissociated, indicating that each signal is selectively controlled by individual molecules elaborated by the macrophage. With an appropriate number of macrophages required for B-cell activation, and sufficient indomethacin to inhibit the accumulation of macrophage-derived prostaglandin, B-lymphocyte clonal proliferation was a linear function of the number of B cells placed in culture. In the absence of macrophages, B-cell colony formation was potentiated by both lipopolysaccharide and intact sheep erythrocytes through a mechanism different from that of the macrophage-derived stimulatory factor. In addition to their direct stimulatory effect on B-cell proliferation, lipopolysaccharide and sheep erythrocytes were each capable of modulating the production and/or release of B-cell stimulatory and inhibitory factors by the macrophage. Parallel studies of conventional mitogen- stimulated lymphocyte cultures did not show a requirement for macrophages and confirm that the semisolid assay is uniquely suited to studies on the regulatory role of the macrophage in B-cell activation.

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Selected References

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  1. Bach F. H., Alter B. J., Solliday S., Zoschke D. C., Janis M. Lymphocyte reactivity in vitro. II. Soluble reconstituting factor permitting response of purified lymphocyte. Cell Immunol. 1970 Jul;1(2):219–227. doi: 10.1016/0008-8749(70)90009-2. [DOI] [PubMed] [Google Scholar]
  2. Berlinger N. T., Lopez C., Good R. A. Facilitation or attenuation of mixed leukocyte culture responsiveness by adherent cells. Nature. 1976 Mar 11;260(5547):145–146. doi: 10.1038/260145a0. [DOI] [PubMed] [Google Scholar]
  3. Broder S., Humphrey R., Durm M., Blackman M., Meade B., Goldman C., Strober W., Waldmann T. Impaired synthesis of polyclonal (non-paraprotein) immunoglobulins by circulating lymphocytes from patients with multiple myeloma Role of suppressor cells. N Engl J Med. 1975 Oct 30;293(18):887–892. doi: 10.1056/NEJM197510302931801. [DOI] [PubMed] [Google Scholar]
  4. Calderon J., Kiely J. M., Lefko J. L., Unanue E. R. The modulation of lymphocyte functions by molecules secreted by macrophages. I. Description and partial biochemical analysis. J Exp Med. 1975 Jul 1;142(1):151–164. doi: 10.1084/jem.142.1.151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Diener E., Shortman K., Russell P. Induction of immunity and tolerance in vitro in the absence of phagocytic cells. Nature. 1970 Feb 21;225(5234):731–732. doi: 10.1038/225731a0. [DOI] [PubMed] [Google Scholar]
  6. Epstein L. B., Kreth H. W., Herzenberg L. A. Fluorescence-activated cell sorting of human T and B lymphocytes. II. Identification of the cell type responsible for interferon production and cell proliferation in response to mitogens. Cell Immunol. 1974 Jun;12(3):407–421. doi: 10.1016/0008-8749(74)90097-5. [DOI] [PubMed] [Google Scholar]
  7. Ferreira S. H., Moncada S., Vane J. R. Indomethacin and aspirin abolish prostaglandin release from the spleen. Nat New Biol. 1971 Jun 23;231(25):237–239. doi: 10.1038/newbio231237a0. [DOI] [PubMed] [Google Scholar]
  8. Gailani S., McLimans W. F., Nussbaum A., Robinson F., Roholt O. Studies on normal human bone marrow cultures in controlled environment thin-film culture systems. In Vitro. 1976 May;12(5):363–372. doi: 10.1007/BF02796314. [DOI] [PubMed] [Google Scholar]
  9. Gery I., Waksman B. H. Potentiation of the T-lymphocyte response to mitogens. II. The cellular source of potentiating mediator(s). J Exp Med. 1972 Jul 1;136(1):143–155. doi: 10.1084/jem.136.1.143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hamburger A. W., Salmon S. E. Primary bioassay of human tumor stem cells. Science. 1977 Jul 29;197(4302):461–463. doi: 10.1126/science.560061. [DOI] [PubMed] [Google Scholar]
  11. Hoffmann M., Dutton R. W. Immune response restoration with macrophage culture supernatants. Science. 1971 Jun 4;172(3987):1047–1048. doi: 10.1126/science.172.3987.1047. [DOI] [PubMed] [Google Scholar]
  12. Kincade P. W. Defective colony formation by B lymphocytes from CBA/N and C3H/HeJ mice. J Exp Med. 1977 Feb 1;145(2):249–263. doi: 10.1084/jem.145.2.249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kincade P. W., Ralph P., Moore M. A. Growth of B-lymphocytes clones in semisolid culture is mitogen dependent. J Exp Med. 1976 May 1;143(5):1265–1270. doi: 10.1084/jem.143.5.1265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kincade P. W., Ralph P. Regulation of clonal B-lymphocyte proliferation by anti-immunoglobulin or anti-Ia antibodies. Cold Spring Harb Symp Quant Biol. 1977;41(Pt 1):245–250. doi: 10.1101/sqb.1977.041.01.030. [DOI] [PubMed] [Google Scholar]
  15. King J. L. Progress in the neutral mutation - random drift controversy. Fed Proc. 1976 Aug;35(10):2087–2091. [PubMed] [Google Scholar]
  16. Lipsky P. E., Rosenthal A. S. The induction and regulation of guinea pig B-lymphocyte proliferation in vitro. J Immunol. 1976 Nov;117(5 Pt 1):1594–1602. [PubMed] [Google Scholar]
  17. Ly I. A., Mishell R. I. Separation of mouse spleen cells by passage through columns of sephadex G-10. J Immunol Methods. 1974 Aug;5(3):239–247. doi: 10.1016/0022-1759(74)90108-2. [DOI] [PubMed] [Google Scholar]
  18. Metcalf D. Colony formation in agar by murine plasmacytoma cells: potentiation by hemopoietic cells and serum. J Cell Physiol. 1973 Jun;81(3):397–410. doi: 10.1002/jcp.1040810312. [DOI] [PubMed] [Google Scholar]
  19. Metcalf D., Nossal G. J., Warner N. L., Miller J. F., Mandel T. E., Layton J. E., Gutman G. A. Growth of B-lymphocyte colonies in vitro. J Exp Med. 1975 Dec 1;142(6):1534–1549. doi: 10.1084/jem.142.6.1534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Metcalf D. Role of mercaptoethanol and endotoxin in stimulating B lymphocyte colony formation in vitro. J Immunol. 1976 Mar;116(3):635–638. [PubMed] [Google Scholar]
  21. Metcalf D., Warner N. L., Nossal G. J., Miller J. F., Shortman K., Rabellino E. Growth of B lymphocyte colonies in vitro from mouse lymphoid organs. Nature. 1975 Jun 19;255(5510):630–632. doi: 10.1038/255630a0. [DOI] [PubMed] [Google Scholar]
  22. Metcalf D., Wilson J. W., Shortman K., Miller J. F., Stocker J. The nature of the cells generating B-lymphocyte colonies in vitro. J Cell Physiol. 1976 May;88(1):107–116. doi: 10.1002/jcp.1040880112. [DOI] [PubMed] [Google Scholar]
  23. Namba Y., Hanaoka M. Immunocytology of cultured IgM-forming cells of mouse. II. Purification of phagocytic cell factor and its role in antibody formation. Cell Immunol. 1974 Apr;12(1):74–84. doi: 10.1016/0008-8749(74)90058-6. [DOI] [PubMed] [Google Scholar]
  24. Nelson D. S. Production by stimulated macrophages of factors depressing lymphocyte transformation. Nature. 1973 Nov 30;246(5431):306–307. doi: 10.1038/246306a0. [DOI] [PubMed] [Google Scholar]
  25. Nelson R. D., Leu R. W. Macrophage requirement for production of guinea pig migration inhibitory factor (MIF) in vitro. J Immunol. 1975 Feb;114(2 Pt 1):606–609. [PubMed] [Google Scholar]
  26. Oppenheim J. J., Leventhal B. G., Hersh E. M. The transformation of column-purified lymphocytes with nonspecific and specific antigenic stimuli. J Immunol. 1968 Aug;101(2):262–267. [PubMed] [Google Scholar]
  27. PERKINS E. H., MAKINODAN T. THE SUPPRESSIVE ROLE OF MOUSE PERITONEAL PHAGOCYTES IN AGGLUTININ RESPONSE. J Immunol. 1965 May;94:765–777. [PubMed] [Google Scholar]
  28. Parkhouse R. M., Dutton R. W. Inhibition of spleen cell DNA synthesis by autologous macrophages. J Immunol. 1966 Nov;97(5):663–669. [PubMed] [Google Scholar]
  29. Rosenstreich D. L., Blake J. T., Rosenthal A. S. The peritoneal exudate lymphocyte. I. Differences in antigen responsiveness between peritoneal exudate and lymph node lymphocytes from immunized guinea pigs. J Exp Med. 1971 Nov 1;134(5):1170–1186. doi: 10.1084/jem.134.5.1170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Schrader J. W. Mechanism of activation of the bone marrow-derived lymphocyte. 3. A distinction between a macrophage-produced triggering signal and the amplifying effect on triggered B lymphocytes of allogeneic interactions. J Exp Med. 1973 Dec 1;138(6):1466–1480. doi: 10.1084/jem.138.6.1466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Twomey J. J., Laughter A. H., Farrow S., Douglass C. C. Hodgkin's disease. An immunodepleting and immunosuppressive disorder. J Clin Invest. 1975 Aug;56(2):467–475. doi: 10.1172/JCI108113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Waldman S. R., Gottlieb A. A. Macrophage regulation of DNA synthesis in lymphoid cells: effects of a soluble factor from macrophages. Cell Immunol. 1973 Oct;9(1):142–156. doi: 10.1016/0008-8749(73)90175-5. [DOI] [PubMed] [Google Scholar]
  33. Wood D. D., Gaul S. L. Enhancement of the humoral response of T cell-depleted murine spleens by a factor derived from human monocytes in vitro. J Immunol. 1974 Sep;113(3):925–933. [PubMed] [Google Scholar]
  34. Yoshinaga M., Yoshinaga A., Waksman B. H. Regulation of lymphocyte responses in vitro. I. Regulatory effect of macrophages and thymus-dependent (T) cells on the response of thymus-independent (B) lymphocytes to endotoxin. J Exp Med. 1972 Oct 1;136(4):956–961. doi: 10.1084/jem.136.4.956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Zeevi A., Goldman I., Rozenszajn L. A. Stimulation and inhibition of human T-lymphocyte colony cell proliferation by hemopoietic cell factors. Cell Immunol. 1977 Feb;28(2):235–247. doi: 10.1016/0008-8749(77)90108-3. [DOI] [PubMed] [Google Scholar]

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