A Comprehensive Profiling of Cellular Sphingolipids in Mammalian Endothelial and Microglial Cells Cultured in Normal and High-Glucose Conditions

Abstract

Sphingolipids (SPLs) play a diverse role in maintaining cellular homeostasis. Dysregulated SPL metabolism is associated with pathological changes in stressed and diseased cells. This study investigates differences in SPL metabolism between cultured human primary retinal endothelial (HREC) and murine microglial cells (BV2) in normal conditions (normal glucose, NG, 5 mM) and under high-glucose (HG, 25 mM)-induced stress by sphingolipidomics, immunohistochemistry, biochemical, and molecular assays. Measurable differences were observed in SPL profiles between HREC and BV2 cells. High-glucose treatment caused a >2.5-fold increase in the levels of Lactosyl-ceramide (LacCer) in HREC, but in BV2 cells, it induced Hexosyl-Ceramides (HexCer) by threefold and a significant increase in Sphingosine-1-phosphate (S1P) compared to NG. Altered SPL profiles coincided with changes in transcript levels of inflammatory and vascular permeability mediators in HREC and inflammatory mediators in BV2 cells. Differences in SPL profiles and differential responses to HG stress between endothelial and microglial cells suggest that SPL metabolism and signaling differ in mammalian cell types and, therefore, their pathological association with those cell types.