Isolation of immunosuppressive phospholipids from marine bacteria

Lipid extracts of marine bacterial strains isolated from seawater and sediments from Japanese coasts were screened for inhibition of mouse mixed lymphocyte reaction (MLR). Two strains, Bacillus sp. SP22 and Alteromonas sp. RS103, were found to contain appreciable MLR-inhibitory activity. Two active components, 22I and 103I, were purified from the extracts of Bacillus sp. SP22 and Alteromonas sp. RS103, respectively, by successive silica gel column chromatographies. The structure of 22I was estimated to be phosphatidic acid containing saturated isofatty acids (C_14:0-C_17:0). 103I consisted of phosphatidylglycerol with saturated isofatty acids (C_15:0-C_18:0). 22I (0.01-1 μg/ml)and 103I (1-10 μg/ml) significantly inhibited MLR in a dose-dependent manner but did not inhibit mitogen-induced proliferation of lymphocytes, unlike cyclosporin A. They did not show any cytotoxicity against mouse splenocytes. L-α-Phosphatidic acid dipalmitoyl, L-α-phosphatidic acid prepared from egg yolk lecithin, and L-α-phosphatidylglycerol dipalmitoyl showed no MLR-inhibitory activity. Isofatty acid-containing phospholipids decreased plasma membrane fluidity of adherent splenocytes and lymphoid neoplasma cell line P388-D1 cells at concentrations with MLR-inhibitory activity, while nonadherent splenocytes were not affected. Daily subcutaneous injection of 221 or 1031 (1 mg/kg/day) resulted in significant suppression of skin-graft rejection between mice with histocompatibility differences. The results suggest that isofatty acid-containing phospholipids suppress the immune responses via the direct action on adherent splenocytes such as macrophages.