In vitro stimulation of immune functions by lipids derived from macrophages exposed to bacterial peptidoglycan

It has previously been established that the processing of Bacillus subtilis cell walls by the macrophage-like cell line RAW264 leads to the formation of peptidoglycolipids containing glycopeptides of bacterial origin. In view of the immunologic activities associated with lipophilic muramyl peptide derivatives, lipid extracts derived from macrophages exposed to bacterial cell walls were assayed for mitogenicity. The crude lipid extract derived from RAW264 cells exposed to bacterial cell walls gave a strong mitogenic response when delivered as liposomes to murine splenocytes, whereas lipids derived from macrophages not exposed to bacterial cell walls did not give a mitogenic response. Fractionation of the RAW264 lipids into neutral lipids, glycolipids, and phospholipids demonstrated that mitogenicity was associated primarily with the phospholipid fraction and thus correlated with the presence of the aforementioned peptidoglycolipids. Mitogenicity was specific to the lipids derived from macrophages exposed to bacterial cell walls and was not observed in lipids derived from macrophages that had either been nonspecifically activated by exposure to Con A or had been allowed to phagocytize latex beads. The mitogenic response was found to be dependent on the presence of macrophages in the splenocyte population and to, at least in part, involve macrophage activation.