DOI: 10.5061/dryad.70rxwdbv0
title: Serum lipidomics in low-density lipoprotein receptor (LDLr) deficient mice on chow diet or Western-type Diet (WTD)

description: Aims: A hallmark of advanced atherosclerosis is inadequate
 immunosuppression by regulatory T (Treg) cells inside atherosclerotic
 lesions. Dyslipidemia has been suggested to alter Treg cell migration by
 affecting the expression of specific membrane proteins, thereby decreasing
 Treg cell migration towards atherosclerotic lesions. Besides membrane
 proteins, cellular metabolism has been shown to be a crucial factor in
 Treg cell migration. We aimed to determine whether dyslipidemia
 contributes to altered migration of Treg cells, in part, by affecting
 cellular metabolism. Methods and results: Dyslipidemia was induced by
 feeding Ldlr-/- mice a Western-type diet for 16-20 weeks and intrinsic
 changes in Treg cells affecting their migration and metabolism were
 examined. Dyslipidemia was associated with altered mTORC2 signaling in
 Treg cells, decreased expression of membrane proteins involved in
 migration, including CD62L, CCR7 and S1Pr1, and decreased Treg cell
 migration towards lymph nodes. Furthermore, we discovered that
 diet-induced dyslipidemia inhibited mTORC1 signaling, induced PPARδ
 activation and increased fatty acid (FA) oxidation in Treg cells.
 Moreover, mass-spectrometry analysis of serum from Ldlr-/- mice with
 normolipidemia or dyslipidemia showed increases in multiple PPARδ ligands
 during dyslipidemia. Treatment with a synthetic PPARδ agonist increased
 the migratory capacity of Treg cells in vitro and in vivo in an FA
 oxidation dependent manner. Furthermore, diet-induced dyslipidemia
 actually enhanced Treg cell migration into the inflamed peritoneum and
 into atherosclerotic lesions in vitro. Conclusions: Altogether, our
 findings implicate that dyslipidemia does not contribute to
 atherosclerosis by impairing Treg cell migration as dyslipidemia
 associated with an effector-like migratory phenotype in Treg cells.
description: <i>Ldlr</i><sup>-/-</sup> mice were fed a
 WTD or maintained on an NCD for 8 weeks and upon sacrifice, serum samples
 were collected and frozen at -80˚C until use. Polar lipids are extracted
 using methanol to precipitate proteins from serum samples and this method
 covers low abundance lipid species, including free fatty acids and
 lysophospholipids—lysophosphatidylcholines (LPCs) and
 lysophosphatidylethanolamines (LPEs). Chromatographic separation was
 achieved on an ACQUITY UPLC™ with a HSS T3 column (1.8 μm, 2.1 * 100 mm)
 coupled to a Q-TOF (Agilent 6530) high resolution mass spectrometer using
 reference mass correction. Lipids were detected in full scan in the
 negative ion mode. The leukotrienes, hydroxyl-fatty acids, epoxy-fatty
 acids and lipoxins were analysed using a fully targeted method as as
 previously described (Strassburg et al.). Oxylipid enrichment was achieved
 using a hydrophilic-lipophilic balance (HLB) SPE cartridge (Oasis).
 Oxylipid analysis used high-performance liquid chromatography (Agilent
 1260) coupled to a triple-quadrupole mass spectrometer (Agilent 6460),
 using an Ascentis® Express column (2.7 μm, 2.1 × 150 mm). Signals from
 each lipid species were normalized for the response of an internal
 standard control, resulting in the relative response values indicated in
 the dataset.