Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2007 Dec 26;104(52):20878-83.
doi: 10.1073/pnas.0709205105. Epub 2007 Dec 19.

Therapeutic B cell depletion impairs adaptive and autoreactive CD4+ T cell activation in mice

Jean-David Bouaziz  1 , Koichi Yanaba, Guglielmo M Venturi, Yaming Wang, Roland M Tisch, Jonathan C Poe, Thomas F Tedder
Affiliations

Therapeutic B cell depletion impairs adaptive and autoreactive CD4+ T cell activation in mice

Jean-David Bouaziz et al. Proc Natl Acad Sci U S A. .

Abstract

CD20 antibody depletion of B lymphocytes effectively ameliorates multiple T cell-mediated autoimmune diseases through mechanisms that remain unclear. To address this, a mouse CD20 antibody that depletes >95% of mature B cells in mice with otherwise intact immune systems was used to assess the role of B cells in CD4(+) and CD8(+) T cell activation and expansion in vivo. B cell depletion had no direct effect on T cell subsets or the activation status of CD4(+) and CD8(+) T cells in naive mice. However, B cell depletion impaired CD4(+) T cell activation and clonal expansion in response to protein antigens and pathogen challenge, whereas CD8(+) T cell activation was not affected. In vivo dendritic cell ablation, along with CD20 immunotherapy, revealed that optimal antigen-specific CD4(+) T cell priming required both B cells and dendritic cells. Most importantly, B cell depletion inhibited antigen-specific CD4(+) T cell expansion in both collagen-induced arthritis and autoimmune diabetes mouse models. These results provide direct evidence that B cells contribute to T cell activation and expansion in vivo and offer insights into the mechanism of action for B cell depletion therapy in the treatment of autoimmunity.

PubMed Disclaimer

Conflict of interest statement

Conflict of interest statement: T.F.T. is a paid consultant for MedImmune, Inc. and a consultant and shareholder for Angelica Therapeutics, Inc. J.C.P is a paid consultant for Angelica Therapeutics, Inc.

Figures

Fig. 1.
Fig. 1.
B cell depletion inhibits antigen-specific CD4+ but not CD8+ T cell expansion. (A) Impaired CD4+ T cell proliferation in B cell-depleted mice after KLH restimulation in vitro. CD20 (open circles) or control (CTL) (filled circles) mAb-treated mice were immunized with KLH. Seven days later, CD4+ (Left) or CD8+ (Right) T cells were purified from draining lymph nodes and cultured in vitro with KLH and mitomycin C-treated B cells from control mice. The values represent mean (±SEM) [3H]thymidine (TdR) uptake from triplicate cultures. (B) Impaired CD4+ T cell proliferation in vivo in B cell-depleted mice after low-dose OVA immunization. CD4+ cells from OT-II Thy1.1+ mice or CD8+ cells from OT-I Thy1.1+ mice were CFSE-labeled and transferred into Thy1.2+ recipients that had been treated with CD20 or control mAb 7 days earlier. After adoptive transfers, the mice were immunized with graded OVA doses (1, 10, and 100 μg). Three days later, splenocytes were harvested and stained to analyze divisions of donor cells. The bar graphs show CFSE geometric mean fluorescence of the whole histogram, which is inversely proportional to cell divisions (CD20 mAb, open bars; control mAb, closed bars). (C) Impaired CD4+ T cell proliferation in B cell-depleted mice after Listeria infection. The same experimental protocols were used as in B except that mice were infected 1 day after T cell adoptive transfers with Listeria that secretes OVA peptide and that splenocytes were harvested 7 days after infection. All data are representative of two independent experiments with at least three mice in each group. Significant differences between sample means are indicated: *, P < 0.05; **, P < 0.01.
Fig. 2.
Fig. 2.
Impaired CD4+ T cell activation after B cell depletion. WT mice treated with CD20 or control (CTL) mAb were infected with Listeria 7 days later. Splenocytes were harvested 7 days after infection. (A) CD44 and CD62L expression by CD4+ and CD8+ T cells. Flow cytometric histograms from a representative mouse in each group are shown. (Left and Center) Numbers denote the percentage of CD4+ (Left) and CD8+ (Center) T cells that were CD44high or CD62Llow. (Right) Bar graphs show CD44 and CD62L geometric mean fluorescence intensities for CD4+ and CD8+ T cells (CD20 mAb, open bars; control mAb, closed bars). (B) (Left and Center) Intracellular IL-2 production by CD4+ T cells (Left) and IFN-γ production by CD8+ T cells (Center) after Listeria infection. Flow cytometric histograms from a representative mouse in each group are shown. (Right) The bar graph denotes the percentage of CD4+ and CD8+ T cells that produce IL-2 and IFN-γ, respectively (CD20 mAb, open bars; CTL mAb, closed bars). All data are representative of two independent experiments with three mice in each group. Significant differences between sample means are indicated: *, P < 0.05; **, P < 0.01.
Fig. 3.
Fig. 3.
Both B cells and DCs contribute to CD4+ T cell expansion in vivo. (A) DT depletion of CD11c+ DCs in CD11c–DTR mice. WT or CD11c–DTR mice were given DT or PBS. Splenocytes were harvested 10 h later and stained for CD11c and B220 expression. Frequencies of DCs are indicated by the gated subpopulations. (B and C) CD4+ and CD8+ T cell proliferation in vivo after B cell and/or DC depletion and Listeria infection. OT-II Thy1.1+ CD4+ cells or OT-I Thy1.1+ CD8+ cells were CFSE-labeled and transferred into Thy1.2+ recipients. WT recipients were treated with control or CD20 mAb, whereas CD11c–DTR mice were treated with DT (DC-depleted) or DT and CD20 mAb (DC- and B cell-depleted). One day after T cell adoptive transfer, mice were infected with Listeria. At day 7 after infection, splenocytes were harvested to analyze divisions of donor cells. The thin lines represent adoptively transferred T cells without Listeria infection. The bar graphs show CFSE geometric (Geo) mean fluorescence of the whole histogram, which is inversely proportional to cell divisions. The data are representative of two experiments with three mice in each group. Significant differences between means are indicated: *, P < 0.05; **, P < 0.01.
Fig. 4.
Fig. 4.
B cell depletion inhibits autoantigen-driven CD4+ T cell expansion in vivo. (A) B cell depletion impairs β cell-specific CD4+ T cell expansion in pancreatic lymph nodes of NOD mice. CFSE-labeled BDC2.5 CD4+ cells were transferred into NOD recipients that had been treated with CD20 or control (CTL) mAb 7 days earlier. Four days later, pancreatic and superficial lymph node cells were stained for Vβ4/CD4 expression and analyzed for CFSE flourescence intensity by flow cytometric analysis. Representative CFSE profiles are shown. (B) Mean CFSE geometric mean fluorescence intensities (± SEM) for CD20 (open bars) and control (closed bars) mAb-treated mice shown in A. (C) B cell depletion impairs CD4+ T cell proliferation in response to collagen immunization. CD20 (open circles) or control (filled circles) mAb-treated DBA/1J mice were immunized with collagen to induce arthritis. Fourteen days later, CD4+ T cells were purified from draining lymph nodes and incubated with collagen plus mitomycin C-treated B cells from control mice (Left) or stimulated with CD3 mAb (Right). Values represent mean (± SEM) [3H]thymidine uptake from triplicate cultures. (A–C) All data represent results obtained in at least four mice for each group. Significant differences between samplemeans are indicated: **, P < 0.01.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Ermann J, Fathman CG. Nat Immunol. 2001;2:759–761. - PubMed
    1. Edwards JCW, Cambridge G. Rheumatology. 2001;40:205–211. - PubMed
    1. Stasi R, Del Poeta G, Stipa E, Evangelista ML, Trawinska MM, Cooper N, Amadori S. Blood. 2007;110:2924–2930. - PubMed
    1. Edwards JCW, Cambridge G. Rheumatology. 2005;44:151–156. - PubMed
    1. Lipsky PE. Nat Immunol. 2001;2:764–766. - PubMed

Publication types