Background/Purpose: Systemic lupus erythematosus (SLE) is a recurrent autoimmune disease characterized by multiple B cell abnormalities, including the activation of naïve B cells. However, gaps in terms of our knowledge regarding the extent and mechanisms of activation remain. We therefore investigated the expression of activation markers on activated naïve (aN) and resting naïve (rN) B cells of SLE patients as well as the stimulatory pathways responsible for their expression. 

Methods: Multidimensional flow analysis was utilized to determine levels of B cell activation markers including CD21, CD23, CD24, CD25, CD69, CD80, CD83, CD86, and IgM on aN and rN B cells of SLE patients with active disease versus healthy controls (HC). In addition, total PBMCs from SLE and HC patients were stimulated ex vivo with B cell receptor (anti-kappa or anti-lambda), T cell (CD40L), or cytokine stimulation for 16 h, 48 h, and 4 days and compared via flow analysis. Lastly, we compared the epigenome of SLE aN and rN by analyzing their genome-wide DNA methylation status using MeDIP-Seq. 

Results: Compared to HC, aN B cells, globally defined by Mitotracker green retention, from SLE patients exhibited low levels of CD21, CD24, CD69, and CD83, while CD86 levels were up-regulated. Following stimulation, aN B cells of SLE patients exhibited increased levels of CD21, CD25, CD69, CD80, CD83, and CD86 and decreased levels of CD24 at 16 h post-stimulation. Levels of CD21 and CD25 decreased at 48 h, followed by CD83 levels on day 4 post-stimulation. Expression of CD69, CD80, and CD86 remained high at all time points. However, aN B cells of SLE patients exhibited differential expression of CD23 and IgM based on stimulation type. Global DNA methylation analysis of aN B cells from an SLE patient revealed several genes, including interferon-regulated genes, were hypomethylated when compared to the HC. Interestingly, the CD83 gene was hypermethylated in aN B cells of the SLE patient; a result consistent with low CD83 levels observed via flow staining and RNASeq transcriptional analysis performed on several SLE patients.

Conclusion: Activated naïve B cells of SLE patients differentially express activation markers including CD21, CD24, CD83, and CD86. The stimulation of total PBMCs through distinct pathways reveals that activation markers are temporally expressed on aN B cells of SLE patients only to decrease with prolonged stimulation, as would be expected in vivo.  Epigenetic analysis indicated hypermethylation of CD83, a result consistent with flow and transcriptional studies. Decreased expression of CD83 could help explain several phenotypic characteristics of lupus B cells including: decreased marginal zone maturation, decreased IL-10 production, and increased Ig secretion [1, 2]. Our results provide important clues regarding abnormal B cell function, highlighting the power of integrated experimental approaches to address this problem. 

1.         Kretschmer, B., et al., CD83 modulates B cell function in vitro: increased IL-10 and reduced Ig secretion by CD83Tg B cells. PLoS One, 2007. 2(8): p. e755.

2.         Luthje, K., et al., CD83 regulates splenic B cell maturation and peripheral B cell homeostasis. Int Immunol, 2008. 20(8): p. 949-60.

ACR Meeting Abstracts - https://acrabstracts.org/abstract/understanding-the-stimulatory-pathways-responsible-for-naive-b-cell-activation-in-systemic-lupus-erythematosus/