Supplementary data: Age, Diet, and Cyp2b status (Cyp2b-null) contribute to changes in lipidomic profiles

The effects of age (9 months old), high-fat diet (4.5 months old), and the loss of the xeno- and endobiotic metabolizing enzymes Cyp2b9, Cyp2b10, and Cyp2b13 (Cyp2b-null mice) on the male murine hepatic lipidome was compared. Hierarchical clustering and principal component analysis show that age perturbs phospholipid profiles and serum lipid markers the most compared to healthy, young mice; followed by a high-fat diet and then loss of Cyp2b. Lipid profiles from older mice followed by diet-induced obese mice contain greater n-6 fatty acids than normal diet (ND)-fed young mice that contain significantly more n-3 fatty acids. The lack of Cyp2b typically enhanced the adverse effects found in the older (9 mo) mice with increased liver injury, cluster of phospholipids, and weight gain combined with a deteriorating cholesterol profile.Treatment of experimental groups Animal care procedures were approved by Clemson University’s Institutional Animal Care and Use committee. Cyp2b-null mice were developed using CRISPR/Cas9 as previously described (Kumar, et al., 2017) and wildtype (WT) B6 mice were purchased from The Jackson Laboratory (Bar Harbor, ME, USA) at 3 weeks of age and acclimated for 6 weeks prior to treatment. WT and Cyp2b-null male (9 weeks old) mice were divided into groups (n=9) and fed either commercially available diets; either a normal chow diet (ND; 2018S-Envigo Teklad Diet, 3.1 Kcal/g: 18.6% protein, 6.2 % fat, 44.2% carbohydrates; Madison, WI USA) or a high-fat diet (HFD; Envigo TD.06414, 5.1 Kcal/g: 60.3% fat (36% saturated, 41% monounsaturated, 23% polyunsaturated fat), 18.3% protein, 21.4% carbohydrates; Madison, WI USA)(Table 1) for 10 weeks (Heintz, et al., 2019). Mice were 4.5 months old at the end of the HFD study and referred to as ND-fed young or HFD-fed young; WT or Cyp2b-null mice. An additional experimental group of WT (Jackson) and Cyp2b-null male mice (n= 5) were fed a ND until they reached 9 months (termed old WT and old Cyp2b-null mice). At the end of the studies, mice were weighed, anesthetized, and blood collected by heart puncture prior to euthanasia and serum prepared. Serum biomarkers and liver triglycerides (TAG) were measured as described (Heintz, et al., 2019). Liver and inguinal white adipose tissue (WAT) were excised, weighed, and divided by total body weight to determine the hepatosomatic index (HSI) and white adipose somatic index (WSI). Tissues were immediately snap frozen in liquid nitrogen and stored at -80°C. Mass SpectrometryLipids were extracted and mass spectrometry performed using a Sciex AC LC system and Sciex QTrap5500 mass spectrometer (Framingham, MA, USA). Phospholipid species were identified and quantified from the livers of mice (n=3 for young (4.5 mo) ND and HFD-fed mice; n=5 for old (9 mo.) mice) from each experimental group by LC-MS/MS at the Emory Integrated Lipidomics Core (EILC) as described in their online protocols.THIS DATASET IS ARCHIVED AT DANS/EASY, BUT NOT ACCESSIBLE HERE. TO VIEW A LIST OF FILES AND ACCESS THE FILES IN THIS DATASET CLICK ON THE DOI-LINK ABOV