Cindy Nguyen, Kenneth Cheung, Xiaoli Chen, Julien Saint‐Pol, Fumitaka Shimizu, Takashi Kanda, Caroline Pot, Maxime Culot, Kannie W. Y. Chan, Lu Wang, Aiping Lü, Wei Jia, Fabien Gosselet
The blood-brain barrier (BBB) is essential for central nervous system homeostasis, and its dysfunction is implicated in neuroinflammatory disorders such as Alzheimer's disease and multiple sclerosis. Oxysterols, cholesterol-derived metabolites, modulate lipid metabolism, immune responses, and BBB physiology, but their specific metabolic effects remain poorly defined. Using an in vitro human BBB model, we examined the impact of tumor necrosis factor-α (TNFα) and selected oxysterols (24S-hydroxycholesterol, 25-hydroxycholesterol, 7α,25-dihydroxycholesterol) on metabolomic profiles of brain-like endothelial cells (hBLEC) and pericytes (hBP) via targeted LC-MS/MS analysis of over 300 metabolites. TNFα markedly increased phenylpyruvic acid, indicating dysregulated phenylalanine metabolism. Oxysterols elicited minimal effects in hBLEC, but significantly altered glycolysis and fatty acid metabolism in hBP, notably reducing fructose 6-phosphate, glucose 6-phosphate, and myristoleic acid, while increasing specific fatty acids under inflammatory conditions. These results identify phenylpyruvic acid as a candidate biomarker of BBB inflammation and highlight pericyte metabolic reprogramming as a key mechanism of oxysterol action.