Eur. J. Immunol. 1992. 22: 1293-1297
Precursor cells that originate germinal centers
1293
Phyllis-Jean Linton., David Lo., Lily Lai., G. Jeanette Thorbecke+ and Norman R. Klinman.
Among naive precursor cell subpopulations only progenitors of memory B cells originate germinal centers*
Department of Immunology., The Scripps Research Institute, La Jolla and New York University Medical Center+, Department of Pathology, Kaplan Cancer Center, New York
Immunization leads to the generation of both antibody-forming cells (AFC) and memory B cells which are thought to arise in germinal centers within lymphoid follicles. The findings that the precursors to memory B cells reside in the J11D'O subpopulation of the spleens in non-immune mice and that this subpopulation is distinct from conventional AFC precursors, including CD5+ B cells, suggest that the precursors of germinal centers might also reside in the J11D'O subpopulation. To test this hypothesis, SCID mice were repopulated with CD4+ carrier-primed T cells and T-depleted JllD'O, J1lDhi or CD5+ B cells and immunized with a hapten-carrier conjugate. Only the J11D'O population was enriched for cells that produced germinal centers. Thus, the subpopulation of precursors that generates memory B cells also originates germinal centers.
1 Introduction Upon immunization, virgin precursor cells in peripheral lymphoid organs respond by the generation of plasma cells and the formation of germinal centers (GC) [l-lo]. G C have been shown to generate memory B cells [4] and they are thought to be the sites wherein recruited cells proliferate and undergo the isotype switching and somatic mutation characteristic of memory B cells [5-151. For example, murine G C cells have been shown to bind peanut agglutinin (PNA) [16] and PNA+ cells isolated at the peak of G C formation have been shown to give rise to secondary responses [4]. Cells obtained from G C and memory B cells appear able to regenerate GC [17, 181; however, the cells from which G C originate in non-immune mice have not been identified. Analyses of mice shortly after primary immunization have indicated that in the spleen antibody-forming cells (AFC) arise in clusters that are temporally, anatomically, and phenotypically distinct from newly arising G C cells [ l , 2, lO].These findings are consistent with the possibility that primary AFC and GC cells originate from separate precursor cells; however, they do not distinguish the possibilities that (a) the microenvironment wherein cells happen to reside at the time of immunization determines whether they will become plasma cells vs. memory cells, or (b) that distinct cell subpopulations give rise to AFC vs. memory cells and the latter localize to GC. Consistent with the latter hypothesis, we have previously demonstrated that, upon antigenic stimulation, Ia+ spleen cells of non-
[I 101801
* This work
was supported by grants A1 15797, A G 01743, and A1 29689 from the National Institutes of Health, IM577 from the American Cancer Society, and a Research and Development award from the American Diabetes Association.
immune mice enriched by virtue of their binding low levels of the J l l D monoclonal antibody (JllD'O) give rise to memory B cells and few AFC, whereas J1lDhi precursors including CD5+ B cells give rise to AFC and not memory B cells [19]. Capitalizing on the ability to enrich for these precursor cell subpopulations, we have tested the prediction that G C arise from a cell subpopulation separate from those that yield primary AFC.This has been accomplished by the transfer to SCID mice, which normally do not express GC [20], cells representative of each of the three subpopulations along with Th. Upon immunization, only the J11D'O population was enriched for cells that generate GC. Although memory B cells generated via stimulation with environmental antigens would be included in the JllD'O population, the generation of G C is likely due to J11D'O primary progenitors to memory B cells since the J11D'O precursors isolated from athymic nude mice also generate G C at a high frequency.
2 Materials and methods 2.1 Animals Male BALBk and C.Bl7-SCID mice were obtained from the breeding colony at The Scripps Research Institute. BALBk nu/nu mice were obtained from the National Cancer Institute. The SCID mice used were 6 weeks of age and were determined to have serum immunoglobulin concentrations of
Precursor cells that originate germinal centers
1293
Phyllis-Jean Linton., David Lo., Lily Lai., G. Jeanette Thorbecke+ and Norman R. Klinman.
Among naive precursor cell subpopulations only progenitors of memory B cells originate germinal centers*
Department of Immunology., The Scripps Research Institute, La Jolla and New York University Medical Center+, Department of Pathology, Kaplan Cancer Center, New York
Immunization leads to the generation of both antibody-forming cells (AFC) and memory B cells which are thought to arise in germinal centers within lymphoid follicles. The findings that the precursors to memory B cells reside in the J11D'O subpopulation of the spleens in non-immune mice and that this subpopulation is distinct from conventional AFC precursors, including CD5+ B cells, suggest that the precursors of germinal centers might also reside in the J11D'O subpopulation. To test this hypothesis, SCID mice were repopulated with CD4+ carrier-primed T cells and T-depleted JllD'O, J1lDhi or CD5+ B cells and immunized with a hapten-carrier conjugate. Only the J11D'O population was enriched for cells that produced germinal centers. Thus, the subpopulation of precursors that generates memory B cells also originates germinal centers.
1 Introduction Upon immunization, virgin precursor cells in peripheral lymphoid organs respond by the generation of plasma cells and the formation of germinal centers (GC) [l-lo]. G C have been shown to generate memory B cells [4] and they are thought to be the sites wherein recruited cells proliferate and undergo the isotype switching and somatic mutation characteristic of memory B cells [5-151. For example, murine G C cells have been shown to bind peanut agglutinin (PNA) [16] and PNA+ cells isolated at the peak of G C formation have been shown to give rise to secondary responses [4]. Cells obtained from G C and memory B cells appear able to regenerate GC [17, 181; however, the cells from which G C originate in non-immune mice have not been identified. Analyses of mice shortly after primary immunization have indicated that in the spleen antibody-forming cells (AFC) arise in clusters that are temporally, anatomically, and phenotypically distinct from newly arising G C cells [ l , 2, lO].These findings are consistent with the possibility that primary AFC and GC cells originate from separate precursor cells; however, they do not distinguish the possibilities that (a) the microenvironment wherein cells happen to reside at the time of immunization determines whether they will become plasma cells vs. memory cells, or (b) that distinct cell subpopulations give rise to AFC vs. memory cells and the latter localize to GC. Consistent with the latter hypothesis, we have previously demonstrated that, upon antigenic stimulation, Ia+ spleen cells of non-
[I 101801
* This work
was supported by grants A1 15797, A G 01743, and A1 29689 from the National Institutes of Health, IM577 from the American Cancer Society, and a Research and Development award from the American Diabetes Association.
immune mice enriched by virtue of their binding low levels of the J l l D monoclonal antibody (JllD'O) give rise to memory B cells and few AFC, whereas J1lDhi precursors including CD5+ B cells give rise to AFC and not memory B cells [19]. Capitalizing on the ability to enrich for these precursor cell subpopulations, we have tested the prediction that G C arise from a cell subpopulation separate from those that yield primary AFC.This has been accomplished by the transfer to SCID mice, which normally do not express GC [20], cells representative of each of the three subpopulations along with Th. Upon immunization, only the J11D'O population was enriched for cells that generate GC. Although memory B cells generated via stimulation with environmental antigens would be included in the JllD'O population, the generation of G C is likely due to J11D'O primary progenitors to memory B cells since the J11D'O precursors isolated from athymic nude mice also generate G C at a high frequency.
2 Materials and methods 2.1 Animals Male BALBk and C.Bl7-SCID mice were obtained from the breeding colony at The Scripps Research Institute. BALBk nu/nu mice were obtained from the National Cancer Institute. The SCID mice used were 6 weeks of age and were determined to have serum immunoglobulin concentrations of
