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IL-21 regulates germinal center B cell differentiation and proliferation through a B cell-intrinsic mechanism.

Zotos D, Coquet JM, Zhang Y, Light A, D'Costa K, Kallies A, Corcoran LM, Godfrey DI, Toellner KM, Smyth MJ, Nutt SL, Tarlinton DM - J. Exp. Med. (2010)

Bottom Line: Germinal centers (GCs) are sites of B cell proliferation, somatic hypermutation, and selection of variants with improved affinity for antigen.The absence of IL-21 or IL-21 receptor does not abrogate the appearance of T cells in GCs or the appearance of CD4 T cells with a follicular helper phenotype.IL-21 thus controls fate choices of GC B cells directly.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Walter and Eliza Hall Institute of Medical Research, Parkville 3052, Australia.

ABSTRACT
Germinal centers (GCs) are sites of B cell proliferation, somatic hypermutation, and selection of variants with improved affinity for antigen. Long-lived memory B cells and plasma cells are also generated in GCs, although how B cell differentiation in GCs is regulated is unclear. IL-21, secreted by T follicular helper cells, is important for adaptive immune responses, although there are conflicting reports on its target cells and mode of action in vivo. We show that the absence of IL-21 signaling profoundly affects the B cell response to protein antigen, reducing splenic and bone marrow plasma cell formation and GC persistence and function, influencing their proliferation, transition into memory B cells, and affinity maturation. Using bone marrow chimeras, we show that these activities are primarily a result of CD3-expressing cells producing IL-21 that acts directly on B cells. Molecularly, IL-21 maintains expression of Bcl-6 in GC B cells. The absence of IL-21 or IL-21 receptor does not abrogate the appearance of T cells in GCs or the appearance of CD4 T cells with a follicular helper phenotype. IL-21 thus controls fate choices of GC B cells directly.

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Radiation chimeras show IL-21 acts directly on B cells. (A) Production of chimeric mice with IL-21 or IL-21R deficiency restricted to the T cells. Irradiated recipients were reconstituted with a 1:4 ratio of IL-21R−/− and CD3−/−, IL-21−/− and CD3−/−, or WT and CD3−/− bone marrows and subsequently immunized with NP-KLH in alum. (B) Symbols depict the frequency of NP+IgG1+ B cells in individual chimeric mice at the indicated time in the presence of WT T cells, IL-21−/− T cells, and IL-21R−/− T cells. Mean is indicated by the bar and data are from two independent experiments. (C) The proportion of memory cells among the NP+IgG1+ B populations shown in B. (D) Production of chimeric mice reconstituted with a 1:1 ratio of C57BL/6 (Ly5.1) and IL-21R−/− (Ly5.2) bone marrow cells. (E) Frequency of NP+IgG1+ B cells in individual mice, partitioned according to Ly5 allotype (IL-21R status) at the indicated times with mean shown by the bar. Data are from two independent experiments. (F) The proportion of the NP-specific B cells shown in E with a CD38hi memory phenotype. Significant differences were determined by a Student’s t test and are indicated.
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fig5: Radiation chimeras show IL-21 acts directly on B cells. (A) Production of chimeric mice with IL-21 or IL-21R deficiency restricted to the T cells. Irradiated recipients were reconstituted with a 1:4 ratio of IL-21R−/− and CD3−/−, IL-21−/− and CD3−/−, or WT and CD3−/− bone marrows and subsequently immunized with NP-KLH in alum. (B) Symbols depict the frequency of NP+IgG1+ B cells in individual chimeric mice at the indicated time in the presence of WT T cells, IL-21−/− T cells, and IL-21R−/− T cells. Mean is indicated by the bar and data are from two independent experiments. (C) The proportion of memory cells among the NP+IgG1+ B populations shown in B. (D) Production of chimeric mice reconstituted with a 1:1 ratio of C57BL/6 (Ly5.1) and IL-21R−/− (Ly5.2) bone marrow cells. (E) Frequency of NP+IgG1+ B cells in individual mice, partitioned according to Ly5 allotype (IL-21R status) at the indicated times with mean shown by the bar. Data are from two independent experiments. (F) The proportion of the NP-specific B cells shown in E with a CD38hi memory phenotype. Significant differences were determined by a Student’s t test and are indicated.

Mentions: The abnormalities in the immune response of intact IL-21−/− and IL-21R−/− mice did not distinguish between IL-21 acting directly on the B cells or having an effect via, for example, Tfh or Th17 cells, both of which are known to respond to and produce IL-21 and both of which are implicated in GC development (Hsu et al., 2008; Nurieva et al., 2008; Vogelzang et al., 2008). To resolve the impact of IL-21 on B cells and T cells, a series of chimeric mice were made by reconstituting irradiated IL-21R−/− or IL-21−/− recipients with mixtures of donor bone marrow (Fig. 5, A and D). In the first series, IL-21−/− and IL-21R−/− recipients were reconstituted with a 4:1 mixture of CD3ε−/− and Ly5.1, CD3ε−/− and IL-21−/−, or CD3ε−/− and IL-21R−/− bone marrow such that CD3 T cells in these mice were Ly5.1 WT, unable to secrete IL-21, or lacked the IL-21R, whereas the vast majority of other leukocytes were WT. 8 wk after reconstitution, mice were immunized with NP-KLH in alum and the frequency of NP-specific B cells was determined at days 14 and 28 of the response (Fig. 5 B). Although the overall B cell response to NP was similar in all groups, declining by an equivalent degree between days 14 and 28, the distribution of NP-specific B cells into the GC and memory compartments was strikingly different. As in the intact IL-21–deficient mice (Fig. 2), there was a near threefold increase in the representation of memory B cells at day 28 compared with controls in the CD3ε−/−/IL-21−/− chimeras in which the T cells were unable to secrete IL-21. The CD3ε−/−/IL-21R−/− group, in which T cells lacked the IL-21R, showed a slight, albeit significant, increase over controls (Fig. 5 C). Despite the ability of NKT cells to secrete IL-21 (Harada et al., 2006), they were ruled out as the source of IL-21 through analysis of TCR Jα18−/− immunized mice (Fig. S5).


IL-21 regulates germinal center B cell differentiation and proliferation through a B cell-intrinsic mechanism.

Zotos D, Coquet JM, Zhang Y, Light A, D'Costa K, Kallies A, Corcoran LM, Godfrey DI, Toellner KM, Smyth MJ, Nutt SL, Tarlinton DM - J. Exp. Med. (2010)

Radiation chimeras show IL-21 acts directly on B cells. (A) Production of chimeric mice with IL-21 or IL-21R deficiency restricted to the T cells. Irradiated recipients were reconstituted with a 1:4 ratio of IL-21R−/− and CD3−/−, IL-21−/− and CD3−/−, or WT and CD3−/− bone marrows and subsequently immunized with NP-KLH in alum. (B) Symbols depict the frequency of NP+IgG1+ B cells in individual chimeric mice at the indicated time in the presence of WT T cells, IL-21−/− T cells, and IL-21R−/− T cells. Mean is indicated by the bar and data are from two independent experiments. (C) The proportion of memory cells among the NP+IgG1+ B populations shown in B. (D) Production of chimeric mice reconstituted with a 1:1 ratio of C57BL/6 (Ly5.1) and IL-21R−/− (Ly5.2) bone marrow cells. (E) Frequency of NP+IgG1+ B cells in individual mice, partitioned according to Ly5 allotype (IL-21R status) at the indicated times with mean shown by the bar. Data are from two independent experiments. (F) The proportion of the NP-specific B cells shown in E with a CD38hi memory phenotype. Significant differences were determined by a Student’s t test and are indicated.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2822601&req=5

fig5: Radiation chimeras show IL-21 acts directly on B cells. (A) Production of chimeric mice with IL-21 or IL-21R deficiency restricted to the T cells. Irradiated recipients were reconstituted with a 1:4 ratio of IL-21R−/− and CD3−/−, IL-21−/− and CD3−/−, or WT and CD3−/− bone marrows and subsequently immunized with NP-KLH in alum. (B) Symbols depict the frequency of NP+IgG1+ B cells in individual chimeric mice at the indicated time in the presence of WT T cells, IL-21−/− T cells, and IL-21R−/− T cells. Mean is indicated by the bar and data are from two independent experiments. (C) The proportion of memory cells among the NP+IgG1+ B populations shown in B. (D) Production of chimeric mice reconstituted with a 1:1 ratio of C57BL/6 (Ly5.1) and IL-21R−/− (Ly5.2) bone marrow cells. (E) Frequency of NP+IgG1+ B cells in individual mice, partitioned according to Ly5 allotype (IL-21R status) at the indicated times with mean shown by the bar. Data are from two independent experiments. (F) The proportion of the NP-specific B cells shown in E with a CD38hi memory phenotype. Significant differences were determined by a Student’s t test and are indicated.
Mentions: The abnormalities in the immune response of intact IL-21−/− and IL-21R−/− mice did not distinguish between IL-21 acting directly on the B cells or having an effect via, for example, Tfh or Th17 cells, both of which are known to respond to and produce IL-21 and both of which are implicated in GC development (Hsu et al., 2008; Nurieva et al., 2008; Vogelzang et al., 2008). To resolve the impact of IL-21 on B cells and T cells, a series of chimeric mice were made by reconstituting irradiated IL-21R−/− or IL-21−/− recipients with mixtures of donor bone marrow (Fig. 5, A and D). In the first series, IL-21−/− and IL-21R−/− recipients were reconstituted with a 4:1 mixture of CD3ε−/− and Ly5.1, CD3ε−/− and IL-21−/−, or CD3ε−/− and IL-21R−/− bone marrow such that CD3 T cells in these mice were Ly5.1 WT, unable to secrete IL-21, or lacked the IL-21R, whereas the vast majority of other leukocytes were WT. 8 wk after reconstitution, mice were immunized with NP-KLH in alum and the frequency of NP-specific B cells was determined at days 14 and 28 of the response (Fig. 5 B). Although the overall B cell response to NP was similar in all groups, declining by an equivalent degree between days 14 and 28, the distribution of NP-specific B cells into the GC and memory compartments was strikingly different. As in the intact IL-21–deficient mice (Fig. 2), there was a near threefold increase in the representation of memory B cells at day 28 compared with controls in the CD3ε−/−/IL-21−/− chimeras in which the T cells were unable to secrete IL-21. The CD3ε−/−/IL-21R−/− group, in which T cells lacked the IL-21R, showed a slight, albeit significant, increase over controls (Fig. 5 C). Despite the ability of NKT cells to secrete IL-21 (Harada et al., 2006), they were ruled out as the source of IL-21 through analysis of TCR Jα18−/− immunized mice (Fig. S5).

Bottom Line: Germinal centers (GCs) are sites of B cell proliferation, somatic hypermutation, and selection of variants with improved affinity for antigen.The absence of IL-21 or IL-21 receptor does not abrogate the appearance of T cells in GCs or the appearance of CD4 T cells with a follicular helper phenotype.IL-21 thus controls fate choices of GC B cells directly.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Walter and Eliza Hall Institute of Medical Research, Parkville 3052, Australia.

ABSTRACT
Germinal centers (GCs) are sites of B cell proliferation, somatic hypermutation, and selection of variants with improved affinity for antigen. Long-lived memory B cells and plasma cells are also generated in GCs, although how B cell differentiation in GCs is regulated is unclear. IL-21, secreted by T follicular helper cells, is important for adaptive immune responses, although there are conflicting reports on its target cells and mode of action in vivo. We show that the absence of IL-21 signaling profoundly affects the B cell response to protein antigen, reducing splenic and bone marrow plasma cell formation and GC persistence and function, influencing their proliferation, transition into memory B cells, and affinity maturation. Using bone marrow chimeras, we show that these activities are primarily a result of CD3-expressing cells producing IL-21 that acts directly on B cells. Molecularly, IL-21 maintains expression of Bcl-6 in GC B cells. The absence of IL-21 or IL-21 receptor does not abrogate the appearance of T cells in GCs or the appearance of CD4 T cells with a follicular helper phenotype. IL-21 thus controls fate choices of GC B cells directly.

Show MeSH
Related in: MedlinePlus