The effect of chemically synthetic mycobacterial mycolates on phospholipidome immunomodulation of murine macrophages

Active pulmonary tuberculosis (TB) is characterized by infection of Mycobacterium tuberculosis (Mtb) living on a lipid diet within lung granulomas. Mtb cell wall mycolic acids (MAs) provide a hydrophobic interface for lipid nutrition and are virulence factors directing macrophages into foam cells. Chemically synthetic MAs, each with a defined stereochemistry, representing the major classes of natural Mtb MA, differentially steer host macrophages. While methoxy-MA (mMA) induces vacuolation and keto-MA (kMA) stimulates cholesterol rich lipid droplet accumulation, alpha-MA (αMA) has no notable effect on macrophage physiology. Here we investigated how the peritoneal macrophage phospholipidome is affected by the individual MAs, employing ESI tandem MS (ESI-MS/MS). Compared to placebo and αMA, oxygenated MA treatment elicited significantly more glycerophospholipid (PL), lysophospholipid, and sphingolipid synthesis. Global PL synthesis and induction of eicosanoid potential was pronounced in phosphatidylethanolamine and phosphatidylserine for kMA macrophages, but only in phosphatidylcholine (PC) for mMA macrophages. All lysoPC species were strongly upregulated by both oxygenated MAs. Finally, enrichment of ceramide and sphingomyelin with saturated acyl chains was associated with mMA treatment, whereas unsaturated acyl chains and dihydrosphingomyelin were mostly associated with kMA treatment. Selective secretion of various ratios of oxygenated MAs may thus steer innate immunity through the phospholipidome of macrophages to establish persistent TB.