Activation of the innate immune system promotes polyclonal antibody secretion to eliminate invading pathogens. Inherent in this process is the potential to activate autoreactive B cells and induce autoimmunity. We showed previously that TLR-stimulated dendritic cells and macrophages regulate B cell tolerance to Smith antigen, in part through the secretion of interleukin-6 (IL-6). In this manuscript, we show that neutralization of IL-6 fails to abrogate macrophage-mediated repression and identify soluble CD40 ligand (CD40L) as a second repressive factor secreted by macrophages. CD40L selectively repressed Ig secretion by chronically antigen-experienced (anergic) immunoglobulin transgenic and nontransgenic B cells but not by transiently stimulated B cells. The importance of macrophages in maintaining B cell tolerance was apparent in lupus-prone MRL/lpr mice. Compared with C57BL/6 mice, macrophages from MRL/lpr mice were significantly less efficient at repressing immunoglobulin secretion coincident with diminished IL-6 and CD40 ligand production. These data indicate that macrophages regulate autoreactive B cells by secreting repressive factors that prohibit terminal differentiation of B cells. The regulation of autoreactive B cells by macrophages is diminished in lupus-prone mice suggesting a role in autoimmunity.