Dimorphic expression of uncoupling protein-3 in golden hamster harderian gland: effects of castration and testosterone administration.

Hamster (Mesocricetus auratus) harderian gland (HG) is a dimorphic orbital gland producing a copious lipid secretion. Two cell-types are present in hamster HG, type I in both sexes, type II only in males. In hamster HGs, we found a marked sexual dichotomy in the expression of uncoupling protein-3 (UCP3), a mitochondrial protein carrier, that probably exports fatty acid anions and fatty acid peroxides from the mitochondrial matrix. Following castration and/or testosterone treatment: (1) UCP3 levels correlated with the type II-cell percentage, not with testosterone levels, (2) in male HGs, UCP3 was comparable to female levels at 30 days post-castration (when the type II-cell percentage had fallen from 50 to 5%), although testosterone was already near zero at 15 days (when neither the type II-cell percentage nor the UCP3 level had fallen), and testosterone-replacement therapy prevented these changes. Testosterone-treated females possessed type II cells and a UCP3 level about twofold higher than in control females. Males displayed more intense UCP3 immunohistochemical positivity in type I HG cells than females. Hence, testosterone may indirectly control UCP3 expression by regulating the gland's morphological and lipid dimorphism. Straight-chain fatty acids [found in alkyl diacylglycerols (ADGs) in males] are oxidized predominantly in mitochondria, branched-chain fatty acids (abundant in ADGs in females) predominantly in peroxisomes, so we speculate that the higher UCP3 expression in males reflects greater fatty acid flux in HG mitochondria. This is supported by our finding that in female (not male) HGs, the peroxisome-rich fraction contained alpha-methylacyl-CoA racemase (AMACR), an enzyme important in the beta-oxidation of branched-chain fatty acids.

Sequential changes in the signal transduction responses of skeletal muscle following food deprivation.

Coping with reduced energy sources entails drastic morphological and functional changes in skeletal muscle, but the sequence of events required classification. We found that gastrocnemius muscle from food-deprived rats shows acute rises in peroxisome proliferator activated receptor (PPAR) gamma coactivator (PGC) -1alpha/PPAR delta nuclear protein and myosin heavy chain (MHC) Ib protein, while type I fibers accumulate and the muscle tissue appears redder. AMP levels, phosphorylation of both AMP-activated protein kinase (AMPK) and its downstream target acetyl coenzyme A carboxylase (ACC) are induced within 6 h. Rapidly increased MyoD mRNA levels are followed by an increase in uncoupling protein (UCP) 3 (UCP3) transcription. Increased serum fatty acid levels coincide with increases in mitochondrial UCP3 protein levels and fatty acid oxidation. Accompanying this is a decrease in AMPK phosphorylation, reversible upon nicotinic acid treatment, indicating that fatty acids may modulate this kinase's activity after the metabolic challenges posed by food deprivation.

3,5-diiodo-L-thyronine powerfully reduces adiposity in rats by increasing the burning of fats.

The effect of thyroid hormones on metabolism has long supported their potential as drugs to stimulate fat reduction, but the concomitant induction of a thyrotoxic state has greatly limited their use. Recent evidence suggests that 3,5-diiodo-L-thyronine (T2), a naturally occurring iodothyronine, stimulates metabolic rate via mechanisms involving the mitochondrial apparatus. We examined whether this effect would result in reduced energy storage. Here, we show that T2 administration to rats receiving a high-fat diet (HFD) reduces both adiposity and body weight gain without inducing thyrotoxicity. Rats receiving HFD + T2 showed (when compared with rats receiving HFD alone) a 13% lower body weight, a 42% higher liver fatty acid oxidation rate, appoximately 50% less fat mass, a complete disappearance of fat from the liver, and significant reductions in the serum triglyceride and cholesterol levels (-52% and -18%, respectively). Thyroid hormones and thyroid-stimulating hormone (TSH) serum levels were not influenced by T2 administration. The biochemical mechanism underlying the effects of T2 on liver metabolism involves the carnitine palmitoyl-transferase system and mitochondrial uncoupling. If the results hold true for humans, pharmacological administration of T2 might serve to counteract the problems associated with overweight, such as accumulation of lipids in liver and serum, without inducing thyrotoxicity. However, the results reported here do not exclude deleterious effects of T2 on a longer time scale as well as do not show that T2 acts in the same way in humans.

Combined cDNA array/RT-PCR analysis of gene expression profile in rat gastrocnemius muscle: relation to its adaptive function in energy metabolism during fasting.

We evaluated the effects of fasting on the gene expression profile in rat gastrocnemius muscle using a combined cDNA array and RT-PCR approach. Of the 1176 distinct rat genes analyzed on the cDNA array, 114 were up-regulated more than twofold in response to fasting, including all 17 genes related to lipid metabolism present on the membranes and all 10 analyzed components of the proteasome machinery. Only 7 genes were down-regulated more than twofold. On the basis of our analysis of genes on the cDNA array plus the data from our RT-PCR assays, the metabolic adaptations shown by rat gastrocnemius muscle during fasting are reflected by i) increased transcription both of myosin heavy chain (MHC) Ib (associated with type I fibers) and of at least three factors involved in the shift toward type I fibers [p27kip1, muscle LIM protein (MLP), cystein rich protein-2], of which one (MLP) has been shown to enhance the activity of MyoD, which would explain the known increase in the expression of skeletal muscle uncoupling protein-3 (UCP3); ii) increased lipoprotein lipase (LPL) expression, known to trigger UCP3 transcription, which tends, together with the first point, to underline the suggested role of UCP3 in mitochondrial lipid handling (the variations under the first point and this one have not been observed in mice, indicating a species-specific regulation of these mechanisms); iii) reduced expression of the muscle-specific coenzyme Q (CoQ)7 gene, which is necessary for mitochondrial CoQ synthesis, together with an increased expression of mitochondrial adenylate kinase 3, which inactivates the resident key enzyme for CoQ synthesis, 3-hydroxy-3-methylglutaryl CoA reductase (HMGR), the mRNA level for which fell during fasting; and iv) increased transcription of components of the proteasomal pathways involved in protein degradation/turnover.

Thyroid hormone affects secretory activity and uncoupling protein-3 expression in rat harderian gland.

The effects of T(3) administration on the rat Harderian gland were examined at morphological, biochemical, and molecular levels. T(3) induced hypertrophy of the two cell types (A and B) present in the glandular epithelium. In type A cells, the hypertrophy was mainly due to an increase in the size of the lipid compartment. The acinar lumina were filled with lipoproteic substances, and the cells often showed an olocrine secretory pattern. In type B cells, the hypertrophy largely consisted of a marked proliferation of mitochondria endowed with tightly packed cristae, the mitochondrial number being nearly doubled (from 62 to 101/100 microm(2)). Although the average area of individual mitochondria decreased by about 50%, the total area of the mitochondrial compartment increased by about 80% (from 11 to 19/100 microm(2)). This could be ascribed to T(3)-induced mitochondrial proliferation. The morphological and morphometric data correlated well with our biochemical results, which indicated that mitochondrial respiratory activity is increased in hyperthyroid rats. T(3), by influencing the metabolic function of the mitochondrial compartment, induces lipogenesis and the release of secretory product by type A cells. Mitochondrial uncoupling proteins 2 and 3 were expressed at both mRNA and protein levels in the euthyroid rat Harderian gland. T(3) treatment increased the mRNA levels of both uncoupling protein 2 (UCP2) and UCP3, but the protein level only of UCP3. A possible role for these proteins in the Harderian gland is discussed.

Thyroid-state influence on protein-expression profile of rat skeletal muscle.

We analyzed the whole-cell protein content of gastrocnemius muscles from rats in different thyroid states. Twenty differentially expressed proteins were unambiguously identified. They were involved in substrates and energy metabolism, stress response, cell structure, and gene expression. This study represents the first systematic identification of thyroid state-induced changes in the skeletal muscle protein-expression profile and reveals new cellular pathways as targets for thyroid hormone action.