Targeted and untargeted lipidomic analyses of bioactive and
metabolic lipids in plasma in EAE mice
A: Scatter plots of regulated fatty acid associated genes
(microarray) in the spinal cord of C57BL6 EAE mice versus control mice
(CFA without MOG) 16 days after immunization (GSE GSE60847). Volcano
plot overviews in Suppl. Fig. 1.
B: Volcano plot show the log2 difference (= fold difference;
X-axis) of metabolic plasma lipids between sapropterin and vehicle
treated EAE mice versus the –log10 of the P-value of the t-test
(Y-axis). Lipids that were reduced in sapropterin-treated mice appear on
the left side of the Y-axis, increased lipids on the right side.
C: The box/scatter plot shows the log2 transformed levels of
fatty acids. The data represent the ratios of the mass spectrometry peak
areas normalized by the peak areas of the respective standard. The box
represents the IQR, whiskers show minimum to maximum, the line is the
median. Each scatter is a mouse (n = 10 per group). Data were compared
with two-way ANOVA for ”fatty acid X treatment” and subsequent posthoc
analysis for treatment using an adjustment of alpha according to Šidák.
D: Canonical discriminant score plots of the first discriminant
factors CanDis1 and CanDisc2 for plasma lipids using 28 lipid species of
five classes as input. The plasma concentrations were obtained from
SJL/J mice immunized with PLP and treated orally with vehicle (2%
DMSO), sapropterin (2 mg/d) or DHAP (4 mg/d) via the drinking water
starting at the day of immunization (n=7 per group). The final blood
sample for lipid analyses was obtained 22 days after immunization.
Lipids encompassed ceramides, hexosylceramides, sphingolipids,
endocannabinoids and lysophosphatidic acids (individually presented in
Suppl. Fig. 3). The dots show individual mice. The ellipses show the
95% confidence.
E: Box/scatter plots of normalized ceramides and
endocannabinoids in vehicle, sapropterin (BH4) or DAHP treated SJL/J-EAE
mice as in A. Lipids were normalized as percentages of the 90%-quantile
(raw concentrations shown in Suppl. Fig. 3). The box represents the
interquartile range, whiskers show minimum to maximum, the line is the
median. Each scatter is a mouse. Data were compared with two-way ANOVA
for ”lipid X treatment” and subsequent posthoc analysis for treatment
using an adjustment of alpha according to Šidák (n = 6-8 per group, * P
< 0.05, ** P < 0.001).
F: Polar plots show the mean normalized levels (percentages of
the 90% quantile) of multiple lipid species analyzed via targeted
LC-MS/MS analyses in plasma of SJL/J-EAE mice treated as in A. Most
ceramides and some lysophosphatidic acids (LPAs) were increased in
sapropterin-treated mice.