A critical role for microglial cholesterol 25-hydroxylase in regulating neuroinflammation following neonatal brain hypoxia-ischemia

Project summary/Abstract Our long-term objectives are to define the complex mechanisms responsible for brain damage and repair following neonatal hypoxia-ischemia (HI), as a model for neonatal hypoxic-ischemic encephalopathy (HIE) in term infants, and to search for novel and specific diagnostic and therapeutic targets for HIE. Inflammation is a hallmark of HI brain injury, during which brain resident microglia play critical roles in both induction and resolution of the inflammatory responses. We recently observed a striking buildup of cholesterol- containing lipid droplets in microglia (lipid-laden microglia) in the injured areas following HI in the neonatal mice, as well as in human HIE patients. In this proposal, we study how abnormal lipid accumulation and defective clearance of cholesterol in microglia affect their normal function and neuroinflammation after neonatal HI. We focus on cholesterol 25-hydroxylase (ch25h), which converts cholesterol to 25-hydroxycholesterol for clearance, and plays multiple roles in reducing intracellular cholesterol overload. We hypothesized that ch25h is critical for the maintenance of microglial homeostasis and attenuation of neuroinflammation following neonatal brain HI. In Aim1, we will assess how ch25h regulates microglial lipid accumulation and function in HI-injured neonatal brain. We will delete ch25h selectively in microglia and investigate the direct effects on lipid accumulation in microglia, microglial phagocytic ability, as well as the production of cytokines and reactive oxygen species. The contribution of microglial ch25h to brain injury are determined at early phase after HI. In Aim 2, to begin dissecting the mechanisms by which ch25h regulates microglia phenotype after neonatal HI, we will characterize the transcriptional profiles of microglia with unperturbed or deficient ch25h in the sham and HI-injured brain using translating ribosome affinity purification (TRAP) technology followed by RNA-sequencing. This is the first study of the crosstalk between cholesterol metabolism and microglial immunity using a clinically relevant mouse model of HIE. By addressing this important gap in knowledge in perinatal brain injury, we will improve our understanding of how lipid regulation/processing in microglia controls their innate immune and inflammatory responses, and contributes to brain damage/recovery. The results of this study could inform novel therapeutic opportunities for neonatal brain injury by targeting microglial lipid metabolism.