Skip to main content
. 2018 Mar 9;7:e35074. doi: 10.7554/eLife.35074

Figure 7. IgD-only cells have intact germinal center responses but impaired IgG1+ SLPC responses.

(A) Splenic (CD19+) B cells from WT, IgM−/−, and IgD−/− mice unimmunized or 5 days after i.p. immunization with 200 μL of 10% SRBCs. (B) Quantification of germinal center (Fashi GL-7hi) cells in (A). (C) Splenocytes from mice in (A). (D) Quantification of CD138+ IgG1+ plasma cells in (C). (E) WT (IgMb+) and IgM-null (IgDa+) germinal center B cells as a percentage of live splenocytes in unimmunized and IgM+/− mice 5 days after i.p. immunization with 200 μL of 10% SRBCs. (F) WT (IgG1b+) and IgM-null (IgG1a+) switched plasma cells (CD138 +IgG1+) as a percentage of live splenocytes in IgM+/− mice unimmunized or 5 days after i.p. immunization with 200 μL of 10% SRBCs. (G) Fraction of unswitched NP-specific germinal center cells (CD19+ Fashi GL-7hi IgM/IgD+) from the IgHa locus in the spleens of Balb/c-B6 F1 and IgM+/− mice 7–8 days after i.p. immunization with 100 μg NP-RSA. (H) Fraction of IgG1+CD138+ plasma cells from the IgHa locus in Balb/c-B6 F1 and IgM+/− mice 7–8 days after i.p. immunization with 100 μg NP-RSA. (I) NP-specific IgG1a and IgG1b titers at OD = 0.2 were calculated for the mice in (G–H) by ELISA. The IgG1a to IgG1b titer ratio was calculated for each mouse, and all ratios were normalized such that the average IgG1a/IgG1b ratio in Balb/c-B6 F1 samples = 1.0. For (A-D), statistics from n = 4 unimmunized mice of each genotype and n = 3 WT, n = 6 IgM−/−, and n = 7 IgD−/− immunized mice were pooled. For (E-F), n = 5 unimmunized and n = 5 immunized mice are shown. For (G-I), n = 5 Balb/c-B6 F1 mice and n = 3 IgM+/− mice are shown. One-way ANOVA with Tukey’s multiple comparisons test (B and D), a paired t test (E-F), and Welch’s t test (G-I) were used to calculate p values, and mean +SEM is displayed. *p<0.05, **p<0.01, ***p<0.001.

Figure 7—source data 1. Numerical data corresponding to germinal center and plasma cell responses in Figure 7B-I,Figure 7—figure supplement 1A–D.
Numerical data corresponding to germinal center and plasma cell responses in Figure 7B-I, Figure 7—figure supplement 1A–B. Numerical data corresponding to unswitched plasma cell responses in Figure 7—figure supplement 1C–D.
elife-35074-fig7-data1.zip (150.6KB, zip)
DOI: 10.7554/eLife.35074.038

Figure 7.

Figure 7—figure supplement 1. Role of BCR allotype and generation of unswitched plasma cells in SRBC-immunized mice.

Figure 7—figure supplement 1.

(A) Quantification of IgMa+ and IgMb+ germinal center B cells (CD19+FashiGL-7hi) as a percentage of live splenocytes in IgHa/b mice 5 days after i.p. immunization with 200 μL of 10% SRBCs. (B) Quantification of IgMa+ and IgMb+ plasma cells (CD138+) as a percentage of live splenocytes in IgHa/b mice 5 days after i.p. immunization with 200 μL of 10% SRBCs. (C) Quantification of splenic unswitched (IgM +or IgD+) CD138+ plasma cells from WT, IgM−/−, and IgD−/− mice unimmunized or 5 days after i.p. immunization with 200 μL of 10% SRBCs. (D) WT (IgMb+) and IgM-null (IgDa+) unswitched plasma cells (CD138+) as a percentage of live splenocytes in IgM+/− mice unimmunized or 5 days after i.p. immunization with 200 μL of 10% SRBCs. For (A) and (B), n = 3 mice were analyzed. Paired t tests (A-B and D) and one-way ANOVA with Tukey’s multiple comparisons test were used to calculate p values, and mean +SEM is displayed. For (C), cell numbers from n = 4 umimmunized mice of each genotype and n = 3 WT, n = 6 IgM−/−, and n = 7 IgD−/− immunized mice were analyzed. For (D), n = 5 unimmunized and n = 5 immunized were analyzed. **p<0.01, ***p<0.001.
Figure 7—figure supplement 2. Role of IgM and IgD in regulating rapid antibody responses.

Figure 7—figure supplement 2.

(A) Model: Control of peripheral B cell tolerance by IgM and IgD. Selective downregulation of IgM is a well-described feature of autoreactive B cells. This study demonstrates how loss of IgM could desensitize autoreactive B cells towards self-antigens. High and invariant IgD expression provides survival signals to allow these autoreactive cells to be maintained in the repertoire. Upon activation by foreign antigen, the least autoreactive cells receive strong signals through IgM and can quickly differentiate into plasma cells with little danger of autoimmunity. In contrast, the most autoreactive B cells are activated through IgD, which is less efficient at transducing signals in vivo. This weak signaling shunts IgDhi IgMlo cells into the germinal center where they can be ‘redeemed’ via somatic hypermutation, as proposed by Goodnow and colleagues. In this way, dual expression of IgM and IgD expression facilitates repertoire diversity and balances immune responsiveness with autoimmune potential.