BCMA is essential for the survival of long-lived bone marrow plasma cells

Abstract

Long-lived humoral immunity is manifested by the ability of bone marrow plasma cells (PCs) to survive for extended periods of time. Recent studies have underscored the importance of BLyS and APRIL as factors that can support the survival of B lineage lymphocytes. We show that BLyS can sustain PC survival in vitro, and this survival can be further enhanced by interleukin 6. Selective up-regulation of Mcl-1 in PCs by BLyS suggests that this alpha-apoptotic gene product may play an important role in PC survival. Blockade of BLyS, via transmembrane activator and cyclophilin ligand interactor-immunoglobulin treatment, inhibited PC survival in vitro and in vivo. Heightened expression of B cell maturation antigen (BCMA), and lowered expression of transmembrane activator and cyclophilin ligand interactor and BAFF receptor in PCs relative to resting B cells suggests a vital role of BCMA in PC survival. Affirmation of the importance of BCMA in PC survival was provided by studies in BCMA-/- mice in which the survival of long-lived bone marrow PCs was impaired compared with wild-type controls. These findings offer new insights into the molecular basis for the long-term survival of PCs.

Figure 1.

BLyS and IL-6 enhances BM PC survival in vitro. After a 6–8-wk immunization with 10 μg PE, whole BM samples were isolated from BALB/c and cultured in vitro with the indicated stimuli for a period of 10 d. (A) After isolation of whole BM samples, 8 × 10⁶ BM cells were cultured in medium alone (♦), 200 ng/ml BLyS (▵), 500 pg/ml IL-6 (▪), IL-6 and BLyS (X), or 10 μg/ml TACI-Ig (•). At days 2, 4, 7, and 10 of in vitro culture, samples were isolated, and the number of PE-specific IgG BM ASCs was enumerated by ELISPOT. These data are representative of four experiments. (B) Immune BM was cultured as in A, and on day 7, samples were stained with αB220, αCD138, and αLy6C antibodies to estimate the number of total PCs (B220⁻CD138⁺Ly6C⁺) in culture. Two representative experiments out of four are shown.

Figure 2.

Blockade of BLyS via TACI-Ig inhibits PC survival in vivo. Approximately 6–8 wk after immunization with PE-CFA, immune mice (four mice/group) were treated with 100 μg TACI-Ig or human Ig every 3 d for a period of 2 wk. After the 2-wk period of treatment, BM, lymph node (LN), or spleen (SP) tissue samples were isolated from treated mice, and the number of αPE IgG ASCs was determined by ELISPOT analysis. Standard error was calculated between values from four independent mice for each treatment group. Data are representative of three independent experiments.

Figure 3.

(A) PCs express BLyS receptor. Immune BM and spleen (SP) was isolated and stained with αCD138, αB220, and biotinylated BLyS. Histograms of BLyS receptor expression are shown for B220⁺CD138⁻ cells (red); B220⁻CD138⁻ cells (green); and B220⁻CD138⁺ cells (blue). Data are representative of three independent experiments. (B) BM PCs express heightened levels of BCMA mRNA but not BAFF-R or TACI. RNA from splenic B cells (B cells), αCD40-induced B cell blasts (αCD40 B), or BM PCs (PC) was isolated via the TRIzol method. Real-time RT-PCR was performed with each sample to evaluate the relative levels of mRNA expression for BCMA, TACI, and BAFF-R. Data are representative of three experiments. (C) BLyS enhances Mcl-1 expression in PCs but not B cells. Purified CD138⁺ BM PCs or splenic B cells were cultured with the indicated stimulus, as described in Fig. 1. After 18 h, RNA was isolated, and the relative expression of antiapoptotic genes was determined relative to β-actin.

Figure 4.

BCMA^−/− mice have a reduced capacity to sustain long-lived BM PCs. BCMA^+/− (WT) and BCMA^−/− mice were immunized with 100 μg of alum-precipitated (4-hydroxy-3-nitrophenyl)acetyl chicken γ globulin for 7 wk. The number of NP-specific IgG ASCs in the BM was quantified by antigen-specific ELISPOT analysis. The determined p-value = 0.00094 and is representative of two experiments each with three mice/group.