Cholesterol and cholesteryl esters are increased in Plin2−/− adrenals. Adrenals collected from fed or 24 h fasted Plin2+/+ and Plin2−/− mice were dissected free from the surrounding fat capsule and analyzed with thin layer chromatography (TLC) or HPLC-qTOF/MS. A: Major neutral lipid species in adrenals of chow-fed 50-week-old female Plin2+/+ and Plin2−/− mice. Lipids normalized against adrenal weight (20 μg) were applied on the TLC plate and developed in heptane:diethyl ether:acetic acid (55:45:1, v/v) to separate the major lipid species. Lipids were visualized with copper sulfate-phosphate solution followed by heating to 150°C (n = 3). B: Cholesteryl ester (CE) and cholesterol content in adrenals of 15-week-old male Plin2+/+ and Plin2−/− mice. Lipids were normalized against protein content (2.5 μg) and separated by TLC. CE and cholesterol were visualized with copper sulfate-phosphate solution followed by heating to 60°C. One representative of three TLC plates is shown. C: Quantification of cholesteryl ester content relative to protein content based on TLC in B (n = 6). D: Quantification of unesterified cholesterol content relative to protein content based on TLC in B (n = 6). E: CE species normalized against adrenal weight of 15-week-old chow-fed female and male Plin2+/+ and Plin2−/− mice analyzed with HPLC-qTOF/MS (n = 3). Results in C-E are presented as means ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001 indicate difference between Plin2+/+ and Plin2−/− mice; #P < 0.05, ##P < 0.01 indicate difference between fed and fasted mice of the same genotype). CE, cholesteryl ester; Chol, cholesterol; DAG, diacylglycerol; MAG, monoacylglycerol; PL, phospholipid. Std., standard; TAG, triacylglycerol.