Abstract 429: Loss Of Ten-Eleven Translocation 2 (TET2) Reduces RNA Expression Of Chemokine Receptor CCR6 And Increases Peritoneal B1 B Cell Number And Total Plasma IgM Level

Abstract

TET2 is an epigenetic regulator with an emerging role in regulating CVD severity. TET2 deficiency in mice (TET2 -/- ) was shown to increase atherosclerosis via upregulation of proinflammatory pathways in macrophages. While TET2 has been reported to regulate germinal center formation and class switch recombination in atherogenic B2 cells, its role in atheroprotective B1 cell subsets remains largely unexplored. To investigate the role of TET2 in B1 subsets, flow cytometry of cells from the peritoneal cavity (PEC), spleen and bone marrow of TET2 -/- mice and wildtype littermate controls (n = 14/group) was performed. Results demonstrated increased B1a (p = 0.0162) and B1b (p = 0.0056) cells in TET2 -/- mice in the PEC, their primary niche, but not in the spleen, while in the bone marrow only TET2 -/- B1a cells were increased (p = 0.0021). An enzyme linked immunosorbent assay using the plasma from these mice demonstrated a significant increase in total IgM in the TET2 -/- mice compared to the wildtype mice (p = 0.0287). In addition, sort purified PEC B1a, B1b and B2 cells from TET2 -/- mice and wildtype littermate controls were analyzed for methylation status and RNA expression (n = 24). RNAseq analysis of TET2 -/- PEC B1a, B1b, and B2 cells revealed significantly reduced chemokine receptor CCR6 expression in B1a (-4.88 fold change, 8.03E-08 padj.) and a trending reduction of CCR6 in B1b cells (-2.17 fold change, 0.3477 padj.) compared to wildtype. Further, methylation analysis showed significant hypermethylation of CCR6 in B1a and B1b TET2 -/- cells compared to wildtype. This was corroborated by gene pathway analysis of the RNAseq data which showed significant downregulation of chemotaxis and migration pathways in B1a and B1b TET2 -/- cells. We conclude that loss of TET2 increases hypermethylation of CCR6 and suppresses CCR6 RNA expression in B1 cells which may impair trafficking out of the PEC to the spleen, but not necessarily the bone marrow, leading to PEC accumulation of B1 subtypes and an increase in the total plasma level of atheroprotective IgM. Further study is needed to identify mechanisms of potential B1 cell-mediated atheroprotection in the case of TET2 loss.