Recently, the role of B cells in atherosclerosis has gained more attention but studies have mainly focused on B1 and follicular B cell subsets. Therefore, the contribution of marginal zone (MZ) B cells in experimental atherosclerosis remains elusive. In the current study, we examined the MZ B cell compartment in atherosclerotic apoE-deficient (apoE) mice and found that hypercholesterolemia in these mice was associated with an increased number and percentage of MZ B cells. This aberrant accumulation of MZ B cells was not associated with alterations in their development or increased proliferation but was due to decreased apoptotic cell death. This decrease in MZ B cell death in apoE mice was associated with the reduced capacity of invariant NKT (iNKT) cells to produce IFN-γ and IL-4 after activation. Lowering cholesterol plasma levels with ezetimibe in apoE mice reversed iNKT function and MZ B cell accumulation. To elucidate the mechanism whereby iNKT cells control MZ B cell accumulation in apoE mice, we performed an adoptive transfer of iNKT cells and found that only wild-type iNKT cells but not IFN-γ iNKT cells reversed MZ B cell accumulation in apoE recipient mice. Our findings reveal that lipid changes associated with atherosclerotic disease induce decreased production of IFN-γ by iNKT, which in turn leads to aberrant accumulation of MZ B cells. This study further extends the importance of iNKT cells in regulating MZ B cell compartment.

Hypercholesterolemia associated with atherosclerotic disease is known to be associated with increased total and oxidized (ox) low-density lipoprotein (LDL)-specific IgM antibodies in circulation. However, the B-cell responses accounting for this increase remain to be elucidated. Here, we observed an association between total IgM and oxLDL-specific IgM autoantibodies with cholesterol in the plasma of hypercholesterolemic apolipoprotein E deficient (apoE −/− ) mice. Our findings also indicated that oxLDL-specific IgM autoantibodies production was restricted to the spleen, but not the lymph nodes. Further examination of the spleen revealed that the extrafollicular responses, but not germinal center reactions, were the dominant antibody-producing pathway. A quiescent population of IgM + plasma cells including oxLDL-specific IgM antibody secreting cells in BM also sustained the elevated IgM antibodies response in circulation. We determined that IgM + plasma cells in the BM were, at least in part, splenic derived by depleting CD11c + DCs and plasmablasts to disrupt the humoral responses. In addition, lowering hypercholesterolemia reduced IgM response by interfering with extrafollicular and BM responses. By elucidating the mechanism underlying the elevated IgM response observed in hypercholesterolemia, this study provides insight into novel immunotherapeutic avenues. Keywords: Antibodies r Extrafollicular responses r Hypercholesterolemia r Plasma cells r SpleenAdditional supporting information may be found in the online version of this article at the publisher's web-site IntroductionThe presence of increased modified lipid specific autoantibodies such as anti-oxLDL (where oxLDL is oxidized low-density lipoprotein) in hypercholesterolemia associated with atherosclerosis in both human and experimental models of the disease is indicative not sIgM −/− B1a (cannot secrete IgM), into splenectomized recipient mice confers protection [7]. Furthermore, the oxLDL-specific E06 antibodies cloned from apoE −/− mice, which is identical in the heavy chain variable region to the B1 origin, classic T15 antiphosphorylcholine idiotypic antibody [2,8], was demonstrated to be able to inhibit the uptake of oxLDL via scavenger receptors on macrophages [9] and also bind to oxidative epitopes on apoptotic cells, which in turn mediate their clearance via classical complement pathway [10,11]. However, despite the importance of IgM antibody production in conferring atheroprotection, little is known on how this IgM antibody response is generated and sustained during atherosclerosis development. Antigen-activated B cells, with the upregulation of chemokine receptor CCR7, migrate to T-B-cell boundaries to receive cognate T-cell help [12,13]. Thereafter, under the influence of B-cell receptor (BCR) affinity to antigen, activated B cells either migrate back into the follicles to participate in the formation of germinal center (GC), or migrate to the bridging channel of the follicle in the spleen or medullary cords in the lymph node (LN) to participate...