Circulating plasmablasts/plasma cells: a potential biomarker for IgG4-related disease

The data derived from the present study indicate that CD19⁺CD24^?CD38^hi plasmablasts/plasma cells are significantly elevated in the peripheral blood of patients with IgG4-RD, and this B-cell population notably correlates with the number of organs involved, serum IgG4, IgG4-RD RI, and the ratio of IgG4/IgG. In addition, peripheral CD19⁺CD24^?CD38^hi plasmablasts/plasma cells decreased remarkably after treatment of glucocorticoids, and the clinical manifestations of patients improved accordingly. These findings are consistent with previous observations [17] and indicate that the level of CD19⁺CD24^?CD38^hi plasmablasts/plasma cells could be an important biomarker in IgG4-RD.

By gene expression analysis, we found that CD19⁺CD24^?CD38^hi plasmablasts/plasma cells differed greatly from the other three subsets of B cells, including naïve B cells, memory B cells, and Bregs, and expressed many genes typical of plasma cells. Considering that the gene expression of CD19⁺CD24^?CD38^hi plasmablasts/plasma cells differed from that of other known B-cell subsets, we attempted to use gene expression analysis to determine their stage of plasma cell maturation. A number of typical plasma cell genes were overexpressed, including XBP1, PRDM1, IRF4, and TNFRSF17, whereas BCL6, SPIB, and PAX5 were underexpressed, which is typical of mature plasma cells. However, expression of these genes does not permit a more detailed analysis of the stage of maturation of plasma cells. It is known, however, that expression of HLA-DR can be employed to identify newly generated cells from more mature plasma cells; in this regard, CD19⁺CD24^?CD38^hi plasmablasts/plasma cells from patients with IgG4-RD exhibited bright expression of HLA-DR protein, as is typical of newly formed plasma cells [27]. Notably, however, messenger RNA (mRNA) levels of various major histocompatibility complex class II molecules were decreased, as has been noted in systemic lupus erythematosus (SLE) [22], suggesting that these plasma cells were transitioning from newly formed to more mature plasma cells that had decreased mRNA expression but retained protein expression of HLA-DR. Consistent with this, IgG4-RD plasma cells greatly overexpressed Mki-67 mRNA, which encodes the Ki-67 protein, a marker of recently divided immunoglobulin-secreting plasma cells [28] and which is increased in plasmablasts compared with memory B cells [29]. Together, the increased expression of HLA-DR protein but not mRNA and the increase in Mki-67 mRNA identify IgG4-RD plasma cells as newly generated and similar to those found in the circulation of patients with SLE [30].

A number of other mRNAs and proteins were assessed to further understand the maturation status of the IgG4-RD plasma cells. The presence of mRNA for both lambda and kappa light chains suggests that these CD19⁺CD24^?CD38^hi plasmablasts/plasma cells are polyclonal, although more detailed analysis on a single-cell level would be required to confirm this. An interesting feature of CD19⁺CD24^?CD38^hi plasmablasts/plasma cells was their decreased expression of CXCR4. This chemokine receptor uniquely recognizes CXCL12 and plays an important role in plasma cell homing to bone marrow and other niches [31, 32]. The decreased expression of CXCR4 could contribute to their persistence in the circulation. A similar abnormality has been noted in SLE plasma cells [22].

To further identify the characteristics of CD19⁺CD24^?CD38^hi plasmablasts/plasma cells, we analyzed the surface markers by flow cytometry, and we found that CD19⁺CD24^?CD38^hi plasmablasts/plasma cells highly expressed CD86, CD62L, IL-6R, CD27, and CD95, which was consistent with those observed in gene levels. CD86 (B7-2) signaling plays a pivotal role in activating T cells [33]. Expression of CD86 on B-lineage cells has been shown to foster B-cell–T-cell collaboration and facilitate immunoglobulin production, including IgE and IgG4 [34–39]. In IgG4-RD, the CD19⁺CD24^?CD38^hi plasmablasts/plasma cell subset highly expressed the CD86 molecule, suggesting the intriguing possibility that this cell subset may maintain some features of earlier B cells and, by virtue of persistent expression of CD86, chronically stimulate T-cell help and in so doing enhance the likelihood of class-switching recombination to the downstream heavy-chain isotypes IgG4 and IgE. The physiologic role of other molecules expressed by CD19⁺CD24^?CD38^hi plasmablasts/plasma cells requires delineation.

As previously reported, circulating plasmablasts identified using CD19^lowCD38⁺CD20^?CD27⁺ phenotypic markers are significantly elevated in active IgG4-RD, even in patients with normal serum IgG4 concentrations, suggesting that plasmablast counts are a potentially useful biomarker for diagnosis of IgG4-RD as well as for assessing response to treatment [13]. In our study, we found that 76.46% of CD19⁺CD24^?CD38^hi cells expressed CD27, indicating that the CD19⁺CD24^?CD38^hi plasmablasts/plasma cell population is a larger one that contained CD19⁺CD20^?CD27⁺CD38^hi cells. Similarly, the ratio of CD19⁺CD24^?CD38^hi cells correlated positively with IgG4-RD RI, as did the absolute number of CD19⁺CD24^?CD38^hi plasmablasts/plasma cells [17]. Although there was no statistical correlation between the magnitude of change in circulating CD19⁺CD24^?CD38^hi plasmablasts/plasma cells and the change in the IgG4-RD RI, this may emerge when more patients are studied. Moreover, the significant correlation between disease activity and circulating plasmablasts/plasma cells, as well as the decrease in circulating plasma cells with therapy, suggests that this might be useful in assessing these patients and their response to treatment.