Bis(monoacylglycero)phosphate protects murine RAW 264.7 macrophages against 7-Ketocholesterol-induced apoptosis and impaired autophagy

Abstract

Atherosclerosis, a leading cause of cardiovascular disease, is driven by the accumulation of oxidized low-density lipoproteins (oxLDL) in arterial walls. 7-Ketocholesterol (7KC), a major oxysterol found in oxLDL and atherosclerotic plaques, triggers multiple cell injuries including loss of lysosomal integrity, oxidative stress, apoptosis, and impaired autophagy in vascular cells. Bis(monoacylglycero)phosphate (BMP), also known as lysobisphosphatidic acid, is a unique phospholipid concentrated in the endolysosomal compartment, known to regulate vesicle dynamics, lysosomal enzyme activities, intracellular cholesterol trafficking and its oxidative metabolism. Using a validated model of BMP enrichment in murine RAW 264.7 macrophages, we investigated whether BMP could exert protective activity against 7KC-induced damage. Our findings revealed that BMP enrichment provides comprehensive protection against 7KC at the cellular level by preserving cell viability, morphology, and neutral lipid balance. Mechanistically, BMP enrichment prevented apoptosis by maintaining mitochondrial integrity and blocking caspase activation. This was demonstrated by normalized BAX/BCL2 ratios, preserved pro-Caspase-3 levels, and reduced PARP cleavage. Remarkably, BMP enrichment also restored autophagic flux, thereby preventing the pathological accumulation of LC3-II and p62 that characterizes autophagy dysfunction. Enhanced colocalization between LC3 and BMP suggests direct functional interactions in the stress response. Gene expression analysis confirmed that BMP enrichment normalized the transcriptional dysregulation of key autophagy regulators, including Sqstm1, Becn1, and Pink1. Taken together, these results suggest that BMP is an endogenous protective factor that counteracts 7KC-induced cellular damage at multiple steps by regulating cell death and autophagy pathways in a coordinated manner.