A genetic lesion that arrests plasma cell homing to the bone marrow

Abstract

The coordinated regulation of chemokine responsiveness plays a critical role in the development of humoral immunity. After antigen challenge and B cell activation, the emerging plasma cells (PCs) undergo CXCL12-induced chemotaxis to the bone marrow, where they produce Ab and persist. Here we show that PCs, but not B cells or T cells from lupus-prone NZM mice, are deficient in CXCL12-induced migration. PC unresponsiveness to CXCL12 results in a marked accumulation of PCs in the spleen of mice, and a concordant decrease in bone marrow PCs. Unlike normal mice, in NZM mice, a majority of the splenic PCs are long-lived. This deficiency is a consequence of the genetic interactions of multiple systemic lupus erythematosus susceptibility loci.

Fig. 1.

PCs accumulate in the spleen but are absent in the BM of NZM mice. Spleen- and BM-derived lymphocytes from autoimmune mice were assessed for the generation of anti-DNA IgG PCs (A and C) or NP-specific IgG PCs (B and D) from mice that were immunized with NP₃₆-KLH CFA (immune) or CFA alone (naïve) for 30 days.

Fig. 2.

PCs are absent in the BM of NZM mice on secondary immunization. NP-specific ELISPOT was performed on spleen and BM cells from naïve NZM and B6 mice and those that were immunized for 30 days (1°) or rechallenged with Ags for 30 days (2°). Total Ag-specific IgG PCs were quantified from individual organs, with the mean (±SD) values from three mice per group shown.

Fig. 3.

The deficit of BM PCs in NZM mice tracks with the NZW parental strain and show aberrant chemokine responsiveness. (A) Immunohistological analysis was performed by staining spleen cryosections with allophycocyanin (APC)-CD4, FITC-B220, and PE-CD138. The localization of CD4⁺ T cells (blue), follicular B cells (green), and PCs (red) are shown from naïve (Left) and immune (Right) B6 and NZB mice. The accumulation of CD138⁺B220^lo PCs in the splenic red pulp of NZM mice is evidenced by areas of yellow (Left). The exclusion of NZM PCs from B cell follicles is shown by CD4 and CD138 staining only (Right). (B) Spleen (SP) or BM cells from TD immune NZM, NZB, and B6 mice were adoptively transferred into Rag1^–/– recipients. Cells from TI immune B6 mice were also transferred. As indicated, recipients also received splenic B cells that were treated with Mito-C. Affinity maturation and long-lived nature of the transferred PCs was measured after 30 days by performing ELISPOT on splenocytes from recipients. (C) Ex vivo chemokine migration of PCs from immune mice to the chemokines, CXCL12 and CXCL13, was examined by flow cytofluorimetric analysis. Migrating cells were collected and stained with fluorochrome-coupled Abs to B220 and CD138. (D) The percentage of input NP-specific IgG PCs that migrated in response to chemokine was measured from the indicated mice by performing ELISPOT on nonmigrating cells (inserts) and those that migrated (wells).

Fig. 4.

Deficient chemokine responsiveness to CXCL12 is restricted to B cells differentiating to a PC fate. (A) Ex vivo flow cytofluorimetric analysis was performed on input spleen cells from immune B6 and NZM before chemotaxis assays. Cells were stained with mAbs to B220, CD138, CXCR4, and VLA4. Fluorescence intensity of CXCR4 and VLA4 are shown on gated early differentiating PCs (B220⁺CD138⁺) and mature PCs (B220^loCD138⁺). (B) Migration of early and mature PCs in response to CXCL12 and CXCL13 were quantified, based on their respective phenotype. (C and D) The selective distortion in CXCL12-induced migration of NZM PCs was further shown by measuring the chemotaxis of mature B and CD4⁺ T cells from identical assays. Profiles shown represent input B (B220⁺) and T (CD4⁺) cells and those that migrated, in response to medium alone or chemokines. The percentage of input B and T cells that migrated in response to chemokine was quantified by performing flow cytofluorimetric analysis on nonmigrating cells (inserts) and those that migrated (wells).

Fig. 5.

Multiple autoimmune susceptibility loci are required for distorted chemokine responsiveness of NZM PCs. (A–C) Spleen and BM cells from B6 congenic mice that singly express the SLE susceptibility locus Sle1, Sle2, and Sle3, or in combination, were assessed for the production of anti-DNA, total NP, and high-affinity NP IgG-secreting PCs by ELISPOT. (D) The percentage of input autoreactive PCs that migrated in response to chemokine was measured from the indicated mice by performing ELISPOT as described (21).