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Serum from patients with SLE instructs monocytes to promote IgG and IgA plasmablast differentiation.

Joo H, Coquery C, Xue Y, Gayet I, Dillon SR, Punaro M, Zurawski G, Banchereau J, Pascual V, Oh S - J. Exp. Med. (2012)

Bottom Line: In this study, we demonstrate that SLE-DCs can efficiently stimulate naive and memory B cells to differentiate into IgG- and IgA-plasmablasts (PBs) resembling those found in the blood of SLE patients.Importantly, SLE-DCs express CD138 and trans-present CD138-bound APRIL to B cells, leading to the induction of IgA switching and PB differentiation in an IFN-α-independent manner.Collectively, our study suggests that a direct myeloid DC-B cell interplay might contribute to the pathogenesis of SLE.

View Article: PubMed Central - HTML - PubMed

Affiliation: Baylor Institute for Immunology Research, Dallas, TX 75204, USA.

ABSTRACT
The development of autoantibodies is a hallmark of systemic lupus erythematosus (SLE). SLE serum can induce monocyte differentiation into dendritic cells (DCs) in a type I IFN-dependent manner. Such SLE-DCs activate T cells, but whether they promote B cell responses is not known. In this study, we demonstrate that SLE-DCs can efficiently stimulate naive and memory B cells to differentiate into IgG- and IgA-plasmablasts (PBs) resembling those found in the blood of SLE patients. SLE-DC-mediated IgG-PB differentiation is dependent on B cell-activating factor (BAFF) and IL-10, whereas IgA-PB differentiation is dependent on a proliferation-inducing ligand (APRIL). Importantly, SLE-DCs express CD138 and trans-present CD138-bound APRIL to B cells, leading to the induction of IgA switching and PB differentiation in an IFN-α-independent manner. We further found that this mechanism of providing B cell help is relevant in vivo, as CD138-bound APRIL is expressed on blood monocytes from active SLE patients. Collectively, our study suggests that a direct myeloid DC-B cell interplay might contribute to the pathogenesis of SLE.

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IFN-DCs and SLE-DCs induce naive B cell differentiation into PBs. DCs were co-cultured with CFSE-labeled naive B cells for 6 or 12 d in the presence of 50 nM CpG and 20 U/ml IL-2. (A) CD38 and CD20 expression as well as CFSE dilution were assessed after 6 d. Experiments using sera from 36 patients and monocytes from 12 healthy controls showed similar results. (B and C) Proliferation (B) and PB differentiation (C) on day 6. (D) Total Ig assayed by ELISA on day 12. In B–D, combined data (mean ± SD) from experiments using sera from 19 patients and cells from 8 healthy controls are presented. (E) Intracellular Ig staining on day 6. Experiments using sera from six patients and monocytes/B cells from four healthy controls showed similar results. (F) RNA was harvested after 6 d of DC and naive B cell co-culture in the presence of 50 nM CpG and 20 U/ml IL-2. RT-PCR was performed to measure AICDA expression. Representative data are from three independent experiments with triplicate assay using sera from eight patients and monocytes from three healthy controls. Individual experiments showed similar results. (G) RNA was harvested after 4 d of co-culture for γ switch circles and after 5 d of co-culture for α switch circles. RT-PCR was performed to measure the relative expression of each switch circle transcript. (F and G) Combined data (mean ± SD) from experiments using sera from four SLE patients and monocytes and B cells from three healthy controls are presented. (H) RNA was harvested after 6 d of DC and naive B cell co-cultures in the presence of 50 nM CpG and 20 U/ml IL-2. The products of an RT-PCR reaction were run on an agarose gel. Three independent experiments using sera from five patients and monocytes and B cells from five healthy controls showed similar results. Student’s t test: *, P < 0.05; **, P < 0.01.
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fig2: IFN-DCs and SLE-DCs induce naive B cell differentiation into PBs. DCs were co-cultured with CFSE-labeled naive B cells for 6 or 12 d in the presence of 50 nM CpG and 20 U/ml IL-2. (A) CD38 and CD20 expression as well as CFSE dilution were assessed after 6 d. Experiments using sera from 36 patients and monocytes from 12 healthy controls showed similar results. (B and C) Proliferation (B) and PB differentiation (C) on day 6. (D) Total Ig assayed by ELISA on day 12. In B–D, combined data (mean ± SD) from experiments using sera from 19 patients and cells from 8 healthy controls are presented. (E) Intracellular Ig staining on day 6. Experiments using sera from six patients and monocytes/B cells from four healthy controls showed similar results. (F) RNA was harvested after 6 d of DC and naive B cell co-culture in the presence of 50 nM CpG and 20 U/ml IL-2. RT-PCR was performed to measure AICDA expression. Representative data are from three independent experiments with triplicate assay using sera from eight patients and monocytes from three healthy controls. Individual experiments showed similar results. (G) RNA was harvested after 4 d of co-culture for γ switch circles and after 5 d of co-culture for α switch circles. RT-PCR was performed to measure the relative expression of each switch circle transcript. (F and G) Combined data (mean ± SD) from experiments using sera from four SLE patients and monocytes and B cells from three healthy controls are presented. (H) RNA was harvested after 6 d of DC and naive B cell co-cultures in the presence of 50 nM CpG and 20 U/ml IL-2. The products of an RT-PCR reaction were run on an agarose gel. Three independent experiments using sera from five patients and monocytes and B cells from five healthy controls showed similar results. Student’s t test: *, P < 0.05; **, P < 0.01.

Mentions: SLE-DCs and IFN-DCs were equally effective at enhancing the proliferation of naive B cells (Fig. 2, A and B). They also equally supported the differentiation of PBs (Fig. 2, A and C), as measured by up-regulation of CD38 and down-regulation of CD20 expression. Although IFN-DCs and SLE-DCs were equally potent at inducing IgM secretion, SLE-DCs were more potent than IFN-DCs at inducing naive B cells to secrete switched isotypes, especially IgA (Fig. 2 D). This was further confirmed by intracellular Ig staining (Fig. 2 E). The lower capability of IFN-DCs to promote IgG and IgA was not caused by a defect in AID (activation-induced cytidine deaminase) expression, as AICDA expression levels were similar in B cells co-cultured with either type of DCs (Fig. 2 F). RT-PCR analysis of switch circle transcripts revealed that naive B cells co-cultured with either IFN-DCs or SLE-DCs had significantly increased levels of Iγ-Cμ (Fig. 2 G). Naive B cells co-cultured with SLE-DCs but not IFN-DCs showed increased levels of IgA class switching as measured by Iα-Cμ transcripts. Additionally, B cells co-cultured with SLE-DCs showed higher levels of the mature transcripts VHDJH-CHγ3 and VHDJH-CHα1 than B cells co-cultured with IFN-DCs (Fig. 2 H). Compared with B cells alone, IFN-DCs also increased expression of VHDJH-CHμ and VHDJH-CHγ3 transcripts. Collectively, SLE-DCs display a potent capability to enhance naive B cell differentiation into IgG- and IgA-PBs. Although SLE sera contain elevated levels of IL-21 (0.05–2 ng/ml; Kang et al., 2011), 1–100 ng/ml of exogenous IL-21 did not further enhance the SLE-DC–mediated naive B cell responses in the presence of IL-2 and CpG (not depicted).


Serum from patients with SLE instructs monocytes to promote IgG and IgA plasmablast differentiation.

Joo H, Coquery C, Xue Y, Gayet I, Dillon SR, Punaro M, Zurawski G, Banchereau J, Pascual V, Oh S - J. Exp. Med. (2012)

IFN-DCs and SLE-DCs induce naive B cell differentiation into PBs. DCs were co-cultured with CFSE-labeled naive B cells for 6 or 12 d in the presence of 50 nM CpG and 20 U/ml IL-2. (A) CD38 and CD20 expression as well as CFSE dilution were assessed after 6 d. Experiments using sera from 36 patients and monocytes from 12 healthy controls showed similar results. (B and C) Proliferation (B) and PB differentiation (C) on day 6. (D) Total Ig assayed by ELISA on day 12. In B–D, combined data (mean ± SD) from experiments using sera from 19 patients and cells from 8 healthy controls are presented. (E) Intracellular Ig staining on day 6. Experiments using sera from six patients and monocytes/B cells from four healthy controls showed similar results. (F) RNA was harvested after 6 d of DC and naive B cell co-culture in the presence of 50 nM CpG and 20 U/ml IL-2. RT-PCR was performed to measure AICDA expression. Representative data are from three independent experiments with triplicate assay using sera from eight patients and monocytes from three healthy controls. Individual experiments showed similar results. (G) RNA was harvested after 4 d of co-culture for γ switch circles and after 5 d of co-culture for α switch circles. RT-PCR was performed to measure the relative expression of each switch circle transcript. (F and G) Combined data (mean ± SD) from experiments using sera from four SLE patients and monocytes and B cells from three healthy controls are presented. (H) RNA was harvested after 6 d of DC and naive B cell co-cultures in the presence of 50 nM CpG and 20 U/ml IL-2. The products of an RT-PCR reaction were run on an agarose gel. Three independent experiments using sera from five patients and monocytes and B cells from five healthy controls showed similar results. Student’s t test: *, P < 0.05; **, P < 0.01.
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fig2: IFN-DCs and SLE-DCs induce naive B cell differentiation into PBs. DCs were co-cultured with CFSE-labeled naive B cells for 6 or 12 d in the presence of 50 nM CpG and 20 U/ml IL-2. (A) CD38 and CD20 expression as well as CFSE dilution were assessed after 6 d. Experiments using sera from 36 patients and monocytes from 12 healthy controls showed similar results. (B and C) Proliferation (B) and PB differentiation (C) on day 6. (D) Total Ig assayed by ELISA on day 12. In B–D, combined data (mean ± SD) from experiments using sera from 19 patients and cells from 8 healthy controls are presented. (E) Intracellular Ig staining on day 6. Experiments using sera from six patients and monocytes/B cells from four healthy controls showed similar results. (F) RNA was harvested after 6 d of DC and naive B cell co-culture in the presence of 50 nM CpG and 20 U/ml IL-2. RT-PCR was performed to measure AICDA expression. Representative data are from three independent experiments with triplicate assay using sera from eight patients and monocytes from three healthy controls. Individual experiments showed similar results. (G) RNA was harvested after 4 d of co-culture for γ switch circles and after 5 d of co-culture for α switch circles. RT-PCR was performed to measure the relative expression of each switch circle transcript. (F and G) Combined data (mean ± SD) from experiments using sera from four SLE patients and monocytes and B cells from three healthy controls are presented. (H) RNA was harvested after 6 d of DC and naive B cell co-cultures in the presence of 50 nM CpG and 20 U/ml IL-2. The products of an RT-PCR reaction were run on an agarose gel. Three independent experiments using sera from five patients and monocytes and B cells from five healthy controls showed similar results. Student’s t test: *, P < 0.05; **, P < 0.01.
Mentions: SLE-DCs and IFN-DCs were equally effective at enhancing the proliferation of naive B cells (Fig. 2, A and B). They also equally supported the differentiation of PBs (Fig. 2, A and C), as measured by up-regulation of CD38 and down-regulation of CD20 expression. Although IFN-DCs and SLE-DCs were equally potent at inducing IgM secretion, SLE-DCs were more potent than IFN-DCs at inducing naive B cells to secrete switched isotypes, especially IgA (Fig. 2 D). This was further confirmed by intracellular Ig staining (Fig. 2 E). The lower capability of IFN-DCs to promote IgG and IgA was not caused by a defect in AID (activation-induced cytidine deaminase) expression, as AICDA expression levels were similar in B cells co-cultured with either type of DCs (Fig. 2 F). RT-PCR analysis of switch circle transcripts revealed that naive B cells co-cultured with either IFN-DCs or SLE-DCs had significantly increased levels of Iγ-Cμ (Fig. 2 G). Naive B cells co-cultured with SLE-DCs but not IFN-DCs showed increased levels of IgA class switching as measured by Iα-Cμ transcripts. Additionally, B cells co-cultured with SLE-DCs showed higher levels of the mature transcripts VHDJH-CHγ3 and VHDJH-CHα1 than B cells co-cultured with IFN-DCs (Fig. 2 H). Compared with B cells alone, IFN-DCs also increased expression of VHDJH-CHμ and VHDJH-CHγ3 transcripts. Collectively, SLE-DCs display a potent capability to enhance naive B cell differentiation into IgG- and IgA-PBs. Although SLE sera contain elevated levels of IL-21 (0.05–2 ng/ml; Kang et al., 2011), 1–100 ng/ml of exogenous IL-21 did not further enhance the SLE-DC–mediated naive B cell responses in the presence of IL-2 and CpG (not depicted).

Bottom Line: In this study, we demonstrate that SLE-DCs can efficiently stimulate naive and memory B cells to differentiate into IgG- and IgA-plasmablasts (PBs) resembling those found in the blood of SLE patients.Importantly, SLE-DCs express CD138 and trans-present CD138-bound APRIL to B cells, leading to the induction of IgA switching and PB differentiation in an IFN-α-independent manner.Collectively, our study suggests that a direct myeloid DC-B cell interplay might contribute to the pathogenesis of SLE.

View Article: PubMed Central - HTML - PubMed

Affiliation: Baylor Institute for Immunology Research, Dallas, TX 75204, USA.

ABSTRACT
The development of autoantibodies is a hallmark of systemic lupus erythematosus (SLE). SLE serum can induce monocyte differentiation into dendritic cells (DCs) in a type I IFN-dependent manner. Such SLE-DCs activate T cells, but whether they promote B cell responses is not known. In this study, we demonstrate that SLE-DCs can efficiently stimulate naive and memory B cells to differentiate into IgG- and IgA-plasmablasts (PBs) resembling those found in the blood of SLE patients. SLE-DC-mediated IgG-PB differentiation is dependent on B cell-activating factor (BAFF) and IL-10, whereas IgA-PB differentiation is dependent on a proliferation-inducing ligand (APRIL). Importantly, SLE-DCs express CD138 and trans-present CD138-bound APRIL to B cells, leading to the induction of IgA switching and PB differentiation in an IFN-α-independent manner. We further found that this mechanism of providing B cell help is relevant in vivo, as CD138-bound APRIL is expressed on blood monocytes from active SLE patients. Collectively, our study suggests that a direct myeloid DC-B cell interplay might contribute to the pathogenesis of SLE.

Show MeSH
Related in: MedlinePlus