Figure 2
Dose-dependent action of thyroid hormone on DAG (A), choline and Peth (B) formation in hepatocytes. [14C]oleic acid pre-labeled cells were treated with 100 nM NaOH (control) or thyroid hormones for 2 min (A). The DAG level in control cells: 4707 ± 544 cpm/107 cells. To determine Peth formation in intact hepatocytes(B), 1.5% ethanol was added 15 min before hormone addition. Cells were treated with hormones for 2 min. Reaction was stopped by addition of ice-cold methanol to the culture dishes. Lipid extraction was performed as described in [17]. Peth was separated by silica gel plates in a solvent system of upper phase of ethyl acetate/2,2,4-trimethylpentane/acetic acid/water (13:2:3:10, v/v). The Peth area identified by co-migration with Peth standard was scraped off the plates and the radioactivity was determined by a liquid scintillation counter. The intensity of the lipid spots was estimated by densitometry using a LKB Ultroscan laser densitometer. Free choline content was determined in intact control and hormone-treated cells by the method described in [18]. Results are mean ± S.E. of five individual experiments. * P < 0.05 vs.control.