B cell depletion in immune thrombocytopenia reveals splenic long-lived plasma cells
Matthieu Mahévas ... Jean-Claude Weill, Claude-Agnès Reynaud
Matthieu Mahévas ... Jean-Claude Weill, Claude-Agnès Reynaud
Published January 2, 2013
Citation Information: J Clin Invest. 2013;123(1):432-442. doi:10.1172/JCI65689.
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Categories: Research Article Autoimmunity

B cell depletion in immune thrombocytopenia reveals splenic long-lived plasma cells

Abstract

Primary immune thrombocytopenia (ITP) is a disorder caused by autoantibody-mediated platelet destruction and decreased platelet production. Rituximab, a B cell–depleting agent, has become the first-line treatment for ITP; however, patients with refractory disease usually require splenectomy. We identified antibody-secreting cells as the major splenic B cell population that is resistant to rituximab. The phenotype, antibody specificity, and gene expression profile of these cells were characterized and compared to those of antibody-secreting cells from untreated ITP spleens and from healthy tissues. Antiplatelet-specific plasma cells (PC) were detected in the spleens of patients with ITP up to 6 months after rituximab treatment, and the PC population displayed a long-lived program similar to the one of bone marrow PC, thus explaining for most of these patients the absence of response to rituximab and the response to splenectomy. When analyzed by multiplex PCR at the single-cell level, normal splenic PC showed a markedly different gene expression profile, with an intermediate signature, including genes characteristic of both long-lived PC and proliferating plasmablasts. Surprisingly, long-lived PC were not detected in untreated ITP spleens. These results suggest that the milieu generated by B cell depletion promotes the differentiation and settlement of long-lived PC in the spleen.

Authors

Matthieu Mahévas, Pauline Patin, François Huetz, Marc Descatoire, Nicolas Cagnard, Christine Bole-Feysot, Simon Le Gallou, Mehdi Khellaf, Olivier Fain, David Boutboul, Lionel Galicier, Mikael Ebbo, Olivier Lambotte, Mohamed Hamidou, Philippe Bierling, Bertrand Godeau, Marc Michel, Jean-Claude Weill, Claude-Agnès Reynaud

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Figure 1

Spleen in ITP is the site of an active B cell response producing anti-GpIIbIIIa–specific ASCs.

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Spleen in ITP is the site of an active B cell response producing anti-Gp...
(A) Flow cytometry analysis of splenic mononuclear cells labeled with antibodies to CD19, CD38, CD20, CD24, HLA-DR, and Ki67. After gating on CD19+ cells, ASCs were identified as CD20–, CD24–, CD38hi (blue); PB were identified as CD20–, CD24–, CD38hi, HLA-DR+, Ki67+ (red); and GC B cells were identified as CD20+, CD24–, CD38+, Ki67+ (green). (B–D) Numbers of splenic (B) PB, (C) PCs (identified as CD19+CD20–CD24–CD27+CD38hiKi67–), and (D) GC B cells in patients with ITP (n = 5) compared with HD (n = 8). (E–J) Analysis by confocal microscopy of spleen sections from a patient with ITP (ITP1) and a HD (17 years old) using anti-CD3, anti-CD20, anti-Ki67, and anti-IgL (anti-κ/λ light chains) antibodies. (E) Secondary follicles with proliferating GC B cells, (G, box enlarged in H) PB (CD20–, Ig κ/λ+, Ki67+), and (I, box enlarged in J) T cells (CD3+Ki67+) in patient with ITP and (F) primary follicles in HD. Scale bars: 100 μm (E–G and I); 5 μm (H and J). Data are representative of 3 ITP and HD samples. (K) ELISPOT quantification of anti-GpIIbIIIa IgG-secreting cells in 3 ITP (top) and 1 HD (bottom left) among total IgG-secreting cells (bottom middle), with KLH as antigen control (bottom right). (L) Frequency of anti-GPIIbIIIa IgG-secreting cells in patients with ITP (n = 5) compared with HD (n = 4). Two-tailed Mann-Whitney tests (***P < 0.001; *P = 0.044). Symbols indicate individual samples; horizontal bars represent mean values (± SEM).