UCP2 mRNA Abundance Research Articles

Differential effects of thyroid hormone manipulation and β adrenoceptor agonist administration on uncoupling protein mRNA abundance in adipose tissue and thermoregulation in neonatal pigs

We have shown that there is significant disparity in the expression of uncoupling proteins (UCP) 2 and 3 between modern-commercial and ancient-Meishan porcine genotypes, commercial pigs also have higher plasma triiodothyronine (T3) in on the first day of life. T3 and the sympathetic nervous system are both known to regulate UCPs in rodents and humans; their role in regulating these proteins in the pig is unknown. This study examined whether thyroid hormone manipulation or administration of a selective β3 adrenoceptor agonist (ZD) influenced plasma hormones, colonic temperature and UCP expression in adipose tissue of two breeds of pig. To mimic the differences observed in thyroid hormone status, piglets from Meishan and commercial litters were randomly assigned to control (1 ml/kg water), T3 (10 mg/kg) (Meishan only), methimazole (a commonly used antithyroid drug) (50 mg/kg) (commercial only) or ZD (10 mg/kg) oral administration for the first 4 days of postnatal life. Adipose tissue UCP2/3 mRNA abundance was measured on day 4 using PCR. T3 administration raised plasma T3 concentrations and increased colonic temperature on day 4. UCP3 mRNA abundance was higher in Meishan, than commercial piglets and was downregulated following T3 administration. Irrespective of genotype, ZD increased UCP2 mRNA abundance. Expression of neither UCP2 nor 3 was related to colonic temperature, regardless of treatment. In conclusion, we have demonstrated a dissociation between thyroid hormones and the sympathetic nervous system in the regulation of UCPs in porcine adipose tissue. We have also suggested that expression of adipose tissue UCP2 and 3 are not related to body temperature in piglets.

Uncoupling protein expression in skeletal muscle and adipose tissue in response to in vivo porcine somatotropin treatment

These experiments examined the potential roles of somatropin (pST) and IGF-I in the regulation of uncoupling protein (UCP)2 and UCP3 and their regulatory proteins peroxisome proliferator activated receptor (PPAR) α, γ and δ using in vivo pST treatment of swine and in vitro supplementation of pST or IGF-I to adipose slices. Six, 90 kg barrows were treated with recombinant pST (10 mg) for 2 week while another six pigs were injected with buffer. Total RNA from outer subcutaneous adipose (OSQ) and middle subcutaneous adipose (MSQ) tissues, leaf fat, liver and longissimus (LM) was amplified by reverse transcription-PCR with quantification of transcripts by capillary electrophoresis with laser-induced fluorescence detection. UCP2 mRNA abundance increased in liver ( P < 0.001) and all three adipose tissues by pST treatment ( P < 0.05). Administration of pST increased UCP3 mRNA abundance by 42% in LM ( P < 0.01). PPARα mRNA abundance increased with pST treatment by 29% in liver ( P < 0.05), while decreasing 25% in LM ( P < 0.05). PPARγ mRNA abundance decreased 32% ( P < 0.01) while PPARδ increased 48% in LM ( P < 0.01) with pST administration. In vitro, pST reduced UCP2 mRNA abundance in OSQ and MSQ tissue slices ( P < 0.05). UCP3 mRNA abundance decreased in OSQ ( P < 0.05) but increased in MSQ ( P < 0.05) with pST. In contrast, IGF-I increased UCP2 and UCP3 mRNA abundance in both MSQ and OSQ slices ( P < 0.05). These experiments suggest pST, IGF-I and metabolic adaptations to pST contribute to regulating UCP2 and UCP3.

Impact of dietary protein content on uncoupling protein mRNA abundance in swine

The present study was designed to determine if dietary protein can alter uncoupling protein (UCP) expression in swine, as has been shown in rats, and attempt to identify the mechanism. Eight pigs (~50 kg body mass) were fed an 18% crude protein (CP) diet while another eight pigs were switched to a diet containing 12% crude protein (CP) and fed these diets until 110 kg body mass. The outer (OSQ) and middle (MSQ) subcutaneous adipose tissues, liver, leaf fat, longissimus (LM), red portion of the semitendinosus (STR) and the white portion of the ST (STW) were analyzed for gene expression by real-time PCR. Feeding of 12% CP did not alter growth or carcass composition, relative to 18% CP (P>0.05). Serum growth hormone, non-esterified fatty acids, triglycerides and urea nitrogen were reduced with the feeding of 12% CP (P<0.05). The UCP2 mRNA abundance was reduced in LM, STR, MSQ and OSQ with feeding of 12% CP (P<0.05), as was UCP3 mRNA abundance in MSQ and STW (P<0.01). Peroxisome proliferation activated receptor α (PPARα) and PPARγ were reduced in MSQ and STR (P<0.05) with feeding 12% CP as was the PPARα regulated protein, acyl CoA oxidase (ACOX, P<0.05). These data suggest that feeding 12% CP relative to 18% CP reduces serum NEFA, which reduces PPARα and PPARγ expression and consequently reduces UCP2 lipoperoxidation in OSQ and STR and also reduced UCP3 associated fatty acid transport in MSQ and STW.

β-Adrenergic regulation of uncoupling protein expression in swine

This study examined the beta-adrenergic regulation of uncoupling protein (UCP) 2 and UCP3 gene expression in porcine tissues. In vitro experiments examined changes in UCP2 and UCP3 gene expression in middle (MSQ) and outer (OSQ) subcutaneous adipose tissues from crossbred neutered male pigs. Incubation of tissue slices (24 h) with 0 to 1000 nM isoproterenol increased UCP2 and UCP3 mRNA abundance in MSQ and OSQ, relative to 18S rRNA (P<0.05). For the in vivo experiment, nine randomly selected pigs (80 kg) were presented with a diet supplemented with 10.0 ppm ractopamine for 2 weeks. Another eight pigs were maintained on a control diet. Dietary ractopamine did not affect adipose UCP2 or UCP3 gene expression (P>0.05). However, UCP2 mRNA abundance was depressed in semitendinosus white (STW, P<0.05) and semitendinosus red (STR, P<0.001) by ractopamine feeding. Also, ractopamine decreased UCP3 mRNA abundance by 28% in STW (P<0.05). The in vitro data suggest that beta-adrenergic agonists directly affect adipose tissue UCP expression, although these adipose effects can be masked by the in vivo physiology. The in vivo data indicate that beta-adrenergic agonists may function in regulating UCP2 and UCP3 expression in selected muscles.

Differential effects of leptin administration on the abundance of UCP2 and glucocorticoid action during neonatal development

In the neonate, adipose tissue and the lung both undergo a rapid transition after birth, which results in dramatic changes in uncoupling protein abundance and glucocorticoid action. Leptin potentially mediates some of these adaptations and is known to promote the loss of uncoupling protein (UCP)1, but its effects on other mitochondrial proteins or glucocorticoid action are not known. We therefore determined the effects of acute and chronic administration of ovine recombinant leptin on brown adipose tissue (BAT) and/or lung in neonatal sheep. For the acute study, eight pairs of 1-day-old lambs received, sequentially, 10, 100, and 100 mug of leptin or vehicle before tissue sampling 4 h from the start of the study, whereas in the chronic study, nine pairs of 1-day-old lambs received 100 mug of leptin or vehicle daily for 6 days before tissue sampling on day 7. Acute leptin decreased the abundance of UCP2, glucocorticoid receptor, and 11beta-hydroxysteroid dehydrogenase (11beta-HSD) type 1 mRNA and increased 11beta-HSD type 2 mRNA abundance in BAT, a pattern that was reversed with chronic leptin administration, which also diminished lung UCP2 protein abundance. In BAT, UCP2 mRNA abundance was positively correlated to plasma leptin and nonesterified fatty acids and negatively correlated to mean colonic temperature in the leptin group at 7 days. In conclusion, leptin administration to the neonatal lambs causes differential effects on UCP2 abundance in BAT and lung. These effects may be important in the development of these tissues, thereby optimizing lung function and fat growth.

EFFECT OF ULTRASONIC STIMULATION ON mRNA ABUNDANCE OF UNCOUPLING PROTEIN (UCP) 2 AND UCP 3 IN GASTROCNEMIUS MUSCLE OF RATS

1. The hypothesis that ultrasonic stimulation upregulates uncoupling protein (UCP) 2 and UCP3 in gastrocnemius muscle by a different mechanism of exercise was investigated in Wister rats. 2. The ultrasnonic-stimulated group was given ultrasonic stimulation to the leg (1 MHz frequency, 1 W/cm2 intensity, 10 min continuously). 3. The exercise group was given exercise training by swimming for 10 min in plastic barrels filled with warm water. 4. After 3 h, rats were killed and the gastrocnemius muscle was removed rapidly, weighed and frozen in liquid nitrogen for real-time quantitative reverse transcription-polymerase chain reaction analysis. 5. In gastrocnenius muscles of ultrasonic-stimulated rats, UCP3 mRNA abundance was significantly increased 3.6-fold and UCP2 mRNA abundance was significantly increased 2.2-fold compared with control rats. 6. In gastrocnenius muscles of exercised rats, UCP3 mRNA abundance was significantly increased 3.5-fold compared with control rats, but no change in UCP2 mRNA abundance was observed. 7. Plasma free fatty acid (FFA) levels were also significantly increased in the ultrasonic stimulation group, as well as the exercise group, compared with the control group. 8. These findings show that ultrasonic stimulation lipolyses subcutaneous fat into FFA and glycerol and upregulates UCP2 and UCP3 mRNA by a mechanism different to that of exercise.

Influence of genotype on the differential ontogeny of uncoupling protein 2 and 3 in subcutaneous adipose tissue and muscle in neonatal pigs

The present study aimed to determine whether porcine genotype and/or postnatal age influenced mRNA abundance or protein expression of uncoupling protein (UCP)2 or 3 in subcutaneous adipose tissue (AT) and skeletal muscle (SM) and the extent to which these differences are associated with breed-specific discordance in endocrine and metabolic profiles. Piglets from commercial and Meishan litters were ranked according to birth weight. Tissue samples were obtained from the three median piglets from each litter on either day 0, 4, 7, 14 or 21 of neonatal life. UCP2 protein abundance in AT was similar between genotypes on the first day of life, but it was elevated at all subsequent postnatal ages (P<0.05) in AT of Meishan piglets. In contrast, UCP2 mRNA abundance was lower in Meishans up to 14 days of age. UCP2 mRNA expression was not correlated with protein abundance in either breed at any age. UCP3 mRNA in AT was similar between breeds up to day 7; thereafter, expression was higher (general linear model, P<0.05) in Meishan piglets. Conversely, UCP3 mRNA expression in SM was higher in commercial piglets after day 7. Colonic temperature remained lower in Meishan than commercial piglets throughout the study; this was most obvious in the immediate post-partum period when Meishan piglets had lower (P<0.05) plasma triiodothyronine. In conclusion, we have demonstrated that porcine genotype influences the expression and abundance of UCP2 and 3, an influence which may, in part, be due to the distinctive endocrine profiles associated with each genotype.

Decreased mitochondrial proton leak and reduced expression of uncoupling protein 3 in skeletal muscle of obese diet-resistant women.

Weight loss in response to caloric restriction is variable. Because skeletal muscle mitochondrial proton leak may account for a large proportion of resting metabolic rate, we compared proton leak in diet-resistant and diet-responsive overweight women and compared the expression and gene characteristics of uncoupling protein (UCP)2 and UCP3. Of 1,129 overweight women who completed the University of Ottawa Weight Management Clinic program, 353 met compliance criteria and were free of medical conditions that could affect weight loss. Subjects were ranked according to percent body weight loss during the first 6 weeks of a 900-kcal meal replacement protocol. The highest and lowest quintiles of weight loss were defined as diet responsive and diet resistant, respectively. After body weight had been stable for at least 10 weeks, 12 of 70 subjects from each group consented to muscle biopsy and blood sampling for determinations of proton leak, UCP mRNA expression, and genetic studies. Despite similar baseline weight and age, weight loss was 43% greater, mitochondrial proton leak-dependent (state 4) respiration was 51% higher (P = 0.0062), and expression of UCP3 mRNA abundance was 25% greater (P < 0.001) in diet-responsive than in diet-resistant subjects. There were no differences in UCP2 mRNA abundance. None of the known polymorphisms in UCP3 or its 5' flanking sequence were associated with weight loss or UCP3 mRNA abundance. Thus, proton leak and the expression of UCP3 correlate with weight loss success and may be candidates for pharmacological regulation of fat oxidation in obese diet-resistant subjects.

Differential regulation of expression of genes encoding uncoupling proteins 2 and 3 in brown adipose tissue during lactation in mice

Thermogenic activity in brown adipose tissue (BAT) decreases during lactation; the down-regulation of the gene encoding uncoupling protein 1 (UCP1) is involved in this process. Our studies show that UCP2 mRNA expression does not change during the breeding cycle in mice. In contrast, UCP3 mRNA is down-regulated in lactation but it recovers after weaning, in parallel with UCP1 mRNA. This leads to a decrease in the content of UCP3 in BAT mitochondria during lactation. Lowering the energy-sparing necessities of lactating dams by decreasing litter size or feeding with a high-fat diet prevented the down-regulation of UCP1 mRNA and UCP3 mRNA. In most cases this resulted in a less marked decrease in UCP1 and UCP3 protein in BAT mitochondria owing to lactation. Fasting for 24h caused a different response in UCP1 and UCP3 mRNA expression: it decreased UCP1 mRNA levels but had no effect on UCP3 mRNA abundance in virgin mice; it even increased UCP3 mRNA expression in lactating dams. These changes did not lead to modifications in UCP1 or UCP3 protein abundance. Whereas acute treatment with peroxisome-proliferator-activated receptor (PPAR)α and PPARγ agonists increased UCP1 mRNA levels only in lactating dams, UCP3 mRNA expression was induced by both kinds of PPAR activator in lactating dams and by PPARα agonists in virgin mice. It is concluded that modifications of UCP2 mRNA levels are not part of the physiological adaptations taking place in BAT during lactation. In contrast, the down-regulation of UCP3 mRNA expression and mitochondrial UCP3 content is consistent with a role for the gene encoding UCP3 in the decrease in metabolic fuel oxidation and thermogenesis in BAT during lactation.

Differential regulation of expression of genes encoding uncoupling proteins 2 and 3 in brown adipose tissue during lactation in mice.

Thermogenic activity in brown adipose tissue (BAT) decreases during lactation; the down-regulation of the gene encoding uncoupling protein 1 (UCP1) is involved in this process. Our studies show that UCP2 mRNA expression does not change during the breeding cycle in mice. In contrast, UCP3 mRNA is down-regulated in lactation but it recovers after weaning, in parallel with UCP1 mRNA. This leads to a decrease in the content of UCP3 in BAT mitochondria during lactation. Lowering the energy-sparing necessities of lactating dams by decreasing litter size or feeding with a high-fat diet prevented the down-regulation of UCP1 mRNA and UCP3 mRNA. In most cases this resulted in a less marked decrease in UCP1 and UCP3 protein in BAT mitochondria owing to lactation. Fasting for 24 h caused a different response in UCP1 and UCP3 mRNA expression: it decreased UCP1 mRNA levels but had no effect on UCP3 mRNA abundance in virgin mice; it even increased UCP3 mRNA expression in lactating dams. These changes did not lead to modifications in UCP1 or UCP3 protein abundance. Whereas acute treatment with peroxisome-proliferator-activated receptor (PPAR)alpha and PPARgamma agonists increased UCP1 mRNA levels only in lactating dams, UCP3 mRNA expression was induced by both kinds of PPAR activator in lactating dams and by PPARalpha agonists in virgin mice. It is concluded that modifications of UCP2 mRNA levels are not part of the physiological adaptations taking place in BAT during lactation. In contrast, the down-regulation of UCP3 mRNA expression and mitochondrial UCP3 content is consistent with a role for the gene encoding UCP3 in the decrease in metabolic fuel oxidation and thermogenesis in BAT during lactation.

Impaired expression of the uncoupling protein-3 gene in skeletal muscle during lactation: fibrates and troglitazone reverse lactation-induced downregulation of the uncoupling protein-3 gene.

The expression of uncoupling protein (UCP)-3 mRNA in skeletal muscle is dramatically reduced during lactation in mice. The reduction in UCP-3 mRNA levels lowers the amount of the UCP-3 protein in skeletal muscle mitochondria during lactation. Spontaneous or abrupt weaning reverses the downregulation of the UCP-3 mRNA but not the reduction in UCP-3 protein levels. In lactating and virgin mice, however, fasting increases UCP-3 mRNA levels. Changes in UCP-3 mRNA occur in parallel with modifications in the levels of free fatty acids, which are reduced in lactation and are upregulated due to weaning or fasting. Modifications in the energy nutritional stress of lactating dams achieved by manipulating litter sizes do not influence UCP-3 mRNA levels in skeletal muscle. Conversely, when mice are fed a high-fat diet after parturition, the downregulation of UCP-3 mRNA and UCP-3 protein levels due to lactation is partially reversed, as is the reduction in serum free fatty acid levels. Treatment of lactating mice with a single injection of bezafibrate, an activator of the peroxisome proliferator-activated receptor (PPAR), raises UCP-3 mRNA in skeletal muscle to levels similar to those in virgin mice. 4-chloro-6-[(2,3-xylidine)-pirimidinylthio] acetic acid (WY-14,643), a specific ligand of the PPAR-alpha subtype, causes the most dramatic increase in UCP-3 mRNA, whereas troglitazone, a specific activator of PPAR-gamma, also significantly increases UCP-3 mRNA abundance in skeletal muscle of lactating mice. However, in virgin mice, bezafibrate and WY-14,643 do not significantly affect UCP-3 mRNA expression, whereas troglitazone is at least as effective as it is in lactating dams. It is proposed that the UCP-3 gene is regulated in skeletal muscle during lactation in response to changes in circulating free fatty acids by mechanisms involving activation of PPARs. The impaired expression of the UCP-3 gene is consistent with the involvement of UCP-3 gene regulation in the reduction of the use of fatty acids as fuel by the skeletal muscle and in impaired adaptative thermogenesis, both of which are major metabolic adaptations that occur during lactation.