Myelin lipid analysis of Fa2h-/-/ Cgt-/- mice. (A) TLC analysis of lipids extracted from purified myelin confirmed the low concentration of NFA-GlcCer in Fa2h-/-/Cgt-/- mice and showed that myelin from Fa2h-/-/Cgt-/- mice contained larger amounts of NFA-sphingomyelin (lane 3) compared to wild-type (lane 1) or Cgt-/- mice (lane 2). Lipid samples were subjected to alkaline hydrolysis before TLC. LPE, lyso-phosphatidylethanolamine; SM, sphingomyelin. (B) Total lipids were isolated from sciatic nerves of 4-week-old mice of the indicated genotypes and separated by HPTLC. Shown are 2 representative chromatograms (Note that lanes 5 to 8 are from the same TLC plate, however, 2 lanes containing lipids from mice with other genotypes have been removed from the Figure). PE, phosphatidylethanolamine. Hexosylceramide (HexCer) (C, D) and sphingomyelin levels (E, F) in CNS myelin (C, E) and sciatic nerves (D, F) were determined by densitometric scanning of HPTLC plates. HexCer levels were normalized to cholesterol. Data were combined from three independent experiments, and lipids from 3-6 animals per genotype were analyzed. Shown are the mean ± SD (CNS myelin: n = 3-5; sciatic nerves: n = 4-6); asterisks indicate statistically significant differences (ANOVA with post hoc Fisher's LSD test, *p < 0.05).