B cells from NOD mice can suppress antigen-specific CD8 T cells via an IL-10- mediated contact dependent DC mechanism

B cells are key contributors to the pathology of T1D, demonstrated by numerous studies in both mouse and man. However, B cell subsets, which include IL-10 producing Bregs, can regulate disease development. In the NOD mouse model, although female mice have higher incidence of spontaneous diabetes, over 20% remain diabetes-free. Our aim was to understand the role of B cells in the ‘protection’ of NOD mice that do not develop diabetes.

We studied B cells from NOD mice, bone marrow-derived dendritic cells (BMDC) from NOD.PI2^tg mice expressing proinsulin2 driven by the MHC class II promoter in antigen presenting cells (APC) and insulinB15-23-specific CD8 T cells from G9Ca-/-CD8 TCR transgenic mice. Stimulated splenic B cells were cultured with NOD.PI2^tg BMDC, and purified G9Ca-/- CD8 T cells. We also used transwell culture system in some experiments to investigate the cell-cell interaction via direct contact or soluble factors. All assays were then analysed by flow cytometry and supernatants were analysed via Meso Scale Discovery (MSD).

B cells from ‘protected’ NOD mice had the potential to produce more IL-10 when stimulated with LPS and anti-CD40, compared to B cells from diabetic NOD mice. In functional assays, LPS stimulated B cells from both diabetic and protected NOD mice significantly down-regulated DC activation markers and had increased IL-10 secretion in supernatants. Furthermore, only LPS-stimulated B cells from both diabetic and protected mice regulated insulin-specific CD8 T cells, demonstrated by a lack of proliferation and decreased pro-inflammatory cytokines. This suppression was contact dependent between NOD B cells and DCs.

We suggest that B cells from protected mice have a regulatory role by producing higher levels of IL-10 and down regulating DC activation in response to innate (LPS) stimulation. This may contribute to the protection from diabetes.