Cholesterol Quantification in Brain and Spinal Cord During Development, Demyelination, and Remyelination

Cholesterol in the central nervous system (CNS) is largely unesterified (>99%) and is predominantly present in the myelin sheath (~70% of total CNS cholesterol). However, cholesterol ester accumulation has been observed during neurological disease. In this cross-sectional study, we measured free cholesterol and cholesterol ester levels in the brains and the spinal cords of Plp1-iCKO-Myrf mouse model during postnatal myelination, demyelination, and remyelination using gas chromatography-mass spectrometry with single ion monitoring (GC-MS-SIM) and liquid chromatography-mass spectrometry (LC-MS). Cholesterol levels in healthy mouse brains increased steadily up to 38 weeks of age. In contrast, cholesterol in the healthy spinal cord increased until P42, and then remained steady out to 38 weeks of age. During demyelination, both brain and spinal cord cholesterol levels were significantly reduced compared to healthy mice and did not return to normal cholesterol levels even during remyelination. Semi-quantitative quantification of cholesterol esters during peak demyelination revealed that cholesterol esters comprise 19% of the measured cholesterol pool in the brain and 66% in the spinal cord. Robust methods for internal standard-based quantification of CNS cholesterol and cholesterol esters were developed, which are critical for revealing mechanisms of cholesterol regulation during disease.