Abstract
Lipoprotein lipase (LPL) is an important enzyme in lipid metabolism, and adipose LPL activity is increased in rats that are deficient in thyroid hormone. To examine the mechanism of thyroid hormone's effect on LPL, LPL gene expression was assessed in the epididymal fat pads of hypothyroid rats. When compared to control rats, LPL activity, mass, and synthetic rate in hypothyroid rats were increased; heparin-releasable LPL activity and mass were increased to 448% and 300% of control, respectively, and [35S]methionine incorporation into LPL was increased to 250% of control. The increases in LPL activity and mass were reversed by treatment of hypothyroid rats with triiodothyronine (T3). However, there was no change in the level of LPL mRNA when compared to the level of gamma-actin mRNA and no effect on LPL transcription using run-off assays. Isolated adipocytes were prepared from normal rats and exposed to 2 nM T3 in vitro for 24 h. The addition of T3 to cultures of adipocytes resulted in a decrease in LPL activity, mass, and [35S]methionine incorporation, but still no change in LPL mRNA level. To determine whether thyroid hormone regulated catecholamine responsiveness, adipocytes were prepared from hypothyroid and control rats, and the responses to epinephrine were compared. Although epinephrine inhibited [35S]methionine incorporation into LPL in control rat adipocytes, there was essentially no effect in hypothyroid rat cells. In addition, T3 treatment of the hypothyroid rats restored the responsiveness to epinephrine. Thus, thyroid hormone regulates LPL in rat adipose tissue posttranscriptionally, resulting in parallel changes in LPL synthetic rate, immunoreactive mass, and activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Highlights
Lipoprotein lipase (LPL) is an important enzyme in lipid metabolism, and adipose LPL activity is increased in rats that are deficient in thyroid hormone
LPL activity and immunoreactive mass were measured in epididymal fat pads from control and hypothyroid rats
Because the changes in LPL activity and mass occurred in parallel, there was no difference in LPL specific activity between control and hypothyroid rats
Summary
Both normal and hypothyroid male Sprague-Dawley rats were purchased from Harland Laboratories (Gilmore, CA), and weighed between 210 and 230 g. Cells were cultured for 24 h in Medium 199 (Imine Scientific, Santa Ana, CA) without serum, and were washed in the same medium, followed by the addition of the indicated concentration of Ts. In some experiments, LPL was measured in the heparin-releasable and extractable fractions of whole epididymal adipose tissue, as described previously ( 14). Adipose tissue pieces were incubated in phosphate-buffered saline (PBS) containing 13 pg/ml heparin (Fisher Scientific Co.) for 30 min at 37°C An aliquot of this solution was assayed as described below. Equal amounts of total RNA were resolved on a 2.2 M formaldehyde-1% agarose gel, transferred to a nylon membrane, and blotted with the 92P-labeled [21] cDNA probes for human LPL [22] and gamma-actin [23] as described previously [14, 19]. All data were expressed as the mean+SEM, and were analyzed using the Student’s t-test
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