The effect of weight reduction on skeletal muscle UCP2 and UCP3 mRNA expression and UCP3 protein content in Type II diabetic subjects

Aims/hypothesis. The aim of this study was to examine the effect of weight loss on UCP2/UCP3 mRNA expression and UCP3 protein content in subjects with Type II (non-insulin-dependent) diabetes mellitus.¶Methods. We studied seven Type II diabetic subjects who followed a 10-week very low calorie diet. Expression of skeletal muscle UCP2 and UCP3 mRNA was measured using RT-competitive PCR and UCP3 protein content by western blotting, before and after the diet. Total and plasma fatty acid oxidation was measured using infusion of ¹³C labelled palmitate.¶Results. Body weight decreased from 105.5 ± 8.2 kg to 91.6 ± 7.2 kg (p < 0.001), after 10 weeks of diet intervention. Expression of UCP2 and UCP3 mRNA were significantly reduced after 10 weeks of diet (p < 0.05) but UCP3 protein contents were not significantly altered. Notably, the change in UCP3L mRNA expression and UCP3 protein content after the very low calorie diet were negatively associated with changes in body weight (r = – 0.97, p = 0.006 and r = – 0.83, p = 0.043, respectively) and BMI (r = – 0.99, p = 0.0007 and r = – 0.9, p = 0.016, respectively). Furthermore, changes in UCP3L mRNA expression and UCP3 protein content induced by the diet were positively correlated with changes in cytosolic fatty acid-binding protein content (r = 0.93, p = 0.023 and r = 0.84, p = 0.039, respectively). No correlation between diet-induced changes in UCP3 protein and resting energy expenditure or plasma non-esterified fatty acid concentrations were found.¶Conclusion/interpretation. The negative correlation between the change in UCP3 protein content after weight loss and the change in BMI, suggests that the decrease in UCP3 during weight loss could prevent further weight loss. The finding that the change in UCP3 protein content correlates with the change in skeletal muscle fatty acid-binding protein content, suggests a role for UCPs in the handling of lipids as a fuel. [Diabetologia (2000) 43: 1408–1416]     

Uncoupling protein-3 content is decreased in peripheral skeletal muscle of patients with COPD.

Mechanical efficiency is reduced in patients with chronic obstructive pulmonary disease (COPD). Furthermore, altered fibre-type distribution and metabolic profile has been observed in peripheral skeletal muscle of COPD patients. Since skeletal muscular uncoupling protein-3 (UCP3) has been implicated in the regulation of energy metabolism, the aim of this study was to assess UCP3 in peripheral skeletal muscle of COPD patients and healthy controls. A total of 16 COPD patients and 11 healthy age-matched control subjects were studied. Mechanical efficiency was measured by means of cycle ergometry. Biopsies were taken from the vastus lateralis, and UCP3 and cytochrome c (as a marker for mitochondrial content) levels were assessed by Western blotting. Muscle fibre types and metabolic profile were examined histochemically. UCP3 levels were markedly decreased in COPD compared to controls. In COPD patients, there was a positive correlation between UCP3 content and the forced expiratory volume in one second. UCP3 content was not related to mechanical efficiency, or other muscular data such as fibre types, markers of oxidative/glycolytic energy metabolism or cytochrome c. The authors of this study conclude that uncoupling protein-3 content is decreased in peripheral skeletal muscle of patients with chronic obstructive pulmonary disease and is related to disease severity, but not to mechanical efficiency. The low uncoupling protein-3 content is independent of the loss of oxidative capacity observed in these patients.     

Fiber type dependent upregulation of human skeletal muscle UCP2 and UCP3 mRNA expression by high-fat diet

OBJECTIVE: To test the hypothesis that consumption of a high-fat diet leads to an increase in UCP mRNA expression in human skeletal muscle. In a group of endurance athletes, with a range in fiber type distribution, we hypothesized that the effect of the high-fat diet on UCP2 and UCP3 mRNA expression is more pronounced in muscle fibers which are known to have a high capacity to shift from carbohydrate to fat oxidation (type IIA fibers). DESIGN: Ten healthy trained athletes (five males, five females) consumed a low-fat diet (17+/-0.9 en% of fat) and high-fat diet (41.4+/-1.4 en% fat) for 4 weeks, separated by a 4 week wash-out period. Muscle biopsies were collected at the end of both dietary periods. MEASUREMENTS: Using RT-PCR, levels of UCP2 and UCP3 mRNA expression were measured and the percentage of type I, IIA and IIB fibers were determined using the myofibrillar ATPase method in all subjects. RESULTS: UCP3L mRNA expression tended to be higher on the high-fat diet, an effect which reached significance when only males were considered (P=0.037). Furthermore, diet-induced change in mRNA expression of UCP3T (r: 0.66, P=0.037), UCP3L (r: 0.61, P=0.06) and UCP2 (r: 0.70, P=0.025), but not UCP3S, correlated significantly with percentage dietary fat on the high-fat diet. Plasma FFA levels were not different during the two diets. Finally, the percentage of type IIA fibers was positively correlated with the diet-induced change in mRNA expression for UCP2 (r: 0.7, P=0.03), UCP3L (r: 0.73, P=0.016) and UCP3T (r: 0.68, P=0.03) but not with UCP3S (r: 0.06, NS). CONCLUSION: UCP2 and UCP3 mRNAs are upregulated by a high-fat diet. This upregulation is more pronounced in humans with high proportions of type IIA fibers, suggesting a role for UCPs in lipid utilization.     

The effect of mild cold exposure on UCP3 mRNA expression and UCP3 protein content in humans

OBJECTIVE: In rodents, adaptive thermogenesis in response to cold exposure and high-fat feeding is accomplished by the activation of the brown adipose tissue specific mitochondrial uncoupling protein, UCP1. The recently discovered human uncoupling protein 3 is a possible candidate for adaptive thermogenesis in humans. In the present study we examined the effect of mild cold exposure on the mRNA and protein expression of UCP3. SUBJECTS: Ten healthy male volunteers (age 24.4 +/- 1.6 y; height 1.83 +/- 0.02 m; weight 77.3 +/- 3.0 kg; percentage body fat 19 +/- 2). DESIGN: Subjects stayed twice in the respiration chamber for 60 h (20.00-8.00 h); once at 22 degrees C (72 degrees F), and once at 16 degrees C (61 degrees F). After leaving the respiration chamber, muscle biopsies were taken and RT-competitive-PCR and Western blotting was used to measure UCP3 mRNA and protein expression respectively. RESULTS: Twenty-four-hour energy expenditure was significantly increased at 16 degrees C compared to 22 degrees C (P<0.05). At 16 degrees C, UCP3T (4.6 +/- 1.0 vs 7.7 +/- 1.5 amol/microg RNA, P=0.07), UCP3L (2.0 +/- 0.5 vs 3.5 +/- 0.9 amol/microg RNA, P=0.1) and UCP3S (2.6 +/- 0.6 vs 4.2 +/- 0.7 amol/microg RNA, P=0.07) mRNA expression tended to be lower compared with at 22 degrees C, whereas UCP3 protein content was, on average, not different. However, the individual differences in UCP3 protein content (16-22 degrees C) correlated positively with the differences in 24 h energy expenditure (r=0.86, P<0.05). CONCLUSION: The present study suggests that UCP3 protein content is related to energy metabolism in humans and might help in the metabolic adaptation to cold exposure. However, the down-regulation of UCP3 mRNA with mild cold exposure suggests that prolonged cold exposure will lead to lower UCP3 protein content. What the function of such down-regulation of UCP3 could be is presently unknown.     

Changes in FAT/CD36, UCP2, UCP3 and GLUT4 gene expression during lipid infusion in rat skeletal and heart muscle

OBJECTIVE: It has been reported that an increased availability of free fatty acids (NEFA) not only interferes with glucose utilization in insulin-dependent tissues, but may also result in an uncoupling effect of heart metabolism. We aimed therefore to investigate the effect of an increased availability of NEFA on gene expression of proteins involved in transmembrane fatty acid (FAT/CD36) and glucose (GLUT4) transport and of the uncoupling proteins UCP2 and 3 at the heart and skeletal muscle level. STUDY DESIGN: Euglycemic hyperinsulinemic clamp was performed after 24 h Intralipid(R) plus heparin or saline infusion in lean Zucker rats. Skeletal and heart muscle glucose utilization was calculated by 2-deoxy-[1-(3)H]-D-glucose technique. Quantification of FAT/CD36, GLUT4, UCP2 and UCP3 mRNAs was obtained by Northern blot analysis or RT-PCR. RESULTS: In Intralipid(R) plus heparin infused animals a significant decrease in insulin-mediated glucose uptake was observed both in the heart (22.62+/-2.04 vs 10.37+/-2.33 ng/mg/min; P<0.01) and in soleus muscle (13.46+/-1.53 vs 6.84+/-2.58 ng/mg/min; P<0.05). FAT/CD36 mRNA was significantly increased in skeletal muscle tissue (+117.4+/-16.3%, P<0.05), while no differences were found at the heart level in respect to saline infused rats. A clear decrease of GLUT4 mRNA was observed in both tissues. The 24 h infusion of fat emulsion resulted in a clear enhancement of UCP2 and UCP3 mRNA levels in the heart (99.5+/-15.3 and 80+/-4%) and in the skeletal muscle (291.5+/-24.7 and 146.9+/-12.7%). CONCLUSIONS: As a result of the increased availability of NEFA, FAT/CD36 gene expression increases in skeletal muscle, but not at the heart level. The augmented lipid fuel supply is responsible for the depression of insulin-mediated glucose transport and for the increase of UCP2 and 3 gene expression in both skeletal and heart muscle.     

Low-intensity exercise increases skeletal muscle protein expression of PPARdelta and UCP3 in type 2 diabetic patients

BACKGROUND: Physical exercise provides health benefits for people with type 2 diabetes mellitus, partly by enhancing skeletal muscle insulin action. We tested the hypothesis that changes in expression of key genes in skeletal muscles relate to exercise-induced improvements in type 2 diabetic patients. METHODS: We determined mRNA expression of 20 selected genes following a self-supervised program of walking (> 150 min per week) over a 4-month period. RESULTS: This level of physical activity improved clinical parameters in approximately half the participants, as determined by reduced hypertension and enhanced insulin sensitivity (defined by reduced plasma-insulin levels and improved homeostasis model assessment (HOMA)). Skeletal muscle mRNA expression of Cbl-associated protein (CAP), diacylglycerol kinase (DGK)delta, uncoupling protein (UCP) 3, nuclear respiratory factor (NRF)-1, and peroxisome proliferator-activated receptor (PPAR)delta tended to increase in type 2 diabetic patients with an improved clinical profile. Skeletal muscle protein expression of PPARdelta and UCP3 was increased significantly after physical exercise in patients with an improved clinical profile, but were unchanged in patients who did not show exercise-mediated improvements in clinical parameters. CONCLUSIONS: This study provides clinical evidence that improvements in insulin sensitivity can be achieved in type 2 diabetic patients after individually executed low-intensity exercise training. Moreover, the positive clinical response to exercise is correlated with changes in skeletal muscle proteins involved in the regulation of mitochondrial biogenesis and metabolism. These changes in skeletal muscle gene expression offer a possible molecular explanation for the improvements in clinical outcomes.     

Reduced gene expression of UCP2 but not UCP3 in skeletal muscle of human obese subjects

Summary Massive overweight is an increasing health problem and underlies several complications which in turn result in premature death. The mechanisms underlying the imbalance between energy intake and energy expenditure, that lead to obesity in humans, are still only partly understood. In rodents, heat generation and the burning of calories by the mitochondrial uncoupling protein 1 (UCP1) are important for metabolic control. However, UCP1 is exclusively expressed in brown fat which is only present in limited amounts in human adults. The recent characterization of two new uncoupling proteins, UCP2 and UCP3, may elucidate potentially important pathways for energy expenditure regulation in man. The aim of this study was to investigate whether obesity is accompanied by aberrations in UCP2 and UCP3 regulation. Expression of these two genes was examined using in situ hybridization in six lean and six obese, but otherwise healthy, men. The UCP2 expression was decreased by 28 % (p = 0.001) in the abdominal muscle of the obese subjects. No differences in UCP3 expression were observed between obese and control subjects, although there was great variation in the expression between subjects. In conclusion, these data suggest an impaired activity of the mitochondrial uncoupling protein UCP2, but probably not UCP3, in obese subjects. This may result in decreased energy expenditure and contribute to the development and maintenance of obesity. [Diabetologia (1998) 41: 935–939] Received: 5 December 1997 and in revised form: 18 February 1998     

Uncoupling proteins: role in insulin resistance and insulin insufficiency

Uncoupling proteins (UCPs) are modulators of mitochondrial metabolism that have been implicated in the development of both insulin resistance and insulin insufficiency, the two major pathophysiological events associated with type 2 diabetes. UCP2 mRNA is expressed in a wide range of tissues; however UCP2 protein expression is restricted to fewer tissues, including the endocrine pancreas, spleen, stomach, brain and the lung. To date, its role in the pathophysiology of diabetes has been most strongly associated with impaired glucose-stimulated insulin secretion from the beta-cell, particularly after its induction by free fatty acids. The physiological role of UCP2 remains controversial, but it may act as a downstream signal transducer of superoxide. UCP3 mRNA and protein are expressed in relatively few tissues, predominantly skeletal muscle, brown adipose tissue and heart. Increased expression of UCP3 in skeletal muscle is associated with protection from diet-induced insulin resistance in mice. In patients with type 2 diabetes UCP3 protein in muscle is reduced by 50% compared to healthy controls. The primary physiological role of the novel UCPs does not appear to be protection against positive energy balance and obesity; this is based largely on findings from studies of UCP2 and UCP3 knockout mice and from observed increases in UCP3 expression with fasting. The mechanism(s) of action of UCP2 and UCP3 are poorly understood. However, findings support roles for UCP2 and UCP3 as modifiers of fatty acid metabolism and in mitigating damage from reactive oxygen species.     

NO-1886 (ibrolipim), a lipoprotein lipase-promoting agent, accelerates the expression of UCP3 messenger RNA and ameliorates obesity in ovariectomized rats

The synthetic compound NO-1886 (ibrolipim, [4-(4-bromo-2-cyano-phenylcarbamoyl)-benzyl]-phosphonic acid diethyl ester, CAS 133208-93-2) is a lipoprotein lipase (LPL)-promoting agent that decreases plasma triglycerides, increases high-density lipoprotein cholesterol levels, and prevents fat accumulation in high fat-fed rats. However, the effect of NO-1886 on body weight, fat accumulation, and energy expenditure in ovariectomized (OVX) rats is not clear. The primary aim of this study was to ascertain whether NO-1886 ameliorated obesity in OVX rats and to examine the effects on fatty acid oxidation-related enzymes. NO-1886 decreased accumulation of visceral fat and suppressed the increase in body weight resulting from the ovariectomy. NO-1886 decreased the respiratory quotient and increased expression of the fatty acid translocase messenger RNA (mRNA) in the liver, soleus muscle, and mesenteric fat. NO-1886 also increased the expression of fatty acid-binding protein mRNA in the liver and soleus muscle and the expression of the uncoupling protein 3 (UCP3) mRNA in the heart, soleus muscle, and mesenteric fat, but not in the brown adipose tissue. Furthermore, NO-1886 did not affect UCP1 and UCP2 in brown adipose tissue. Therefore, amelioration of obesity by NO-1886 in OVX rats is possibly because of an the increased expression of fatty acid oxidation-related enzymes and UCP3, both of which are related to fatty acid transfer and fat use. Our study indicates that the LPL-promoting agent NO-1886 may be potentially beneficial in the treatment of obesity and obesity-linked health problems in postmenopausal women.     

Reduced skeletal muscle uncoupling protein-3 content in prediabetic subjects and type 2 diabetic patients: restoration by rosiglitazone treatment

CONTEXT: The mitochondrial uncoupling protein-3 (UCP3) has been implicated in the protection of the mitochondrial matrix against lipid-induced mitochondrial damage. Recent evidence points toward mitochondrial aberrations as a major contributor to the development of insulin resistance and diabetes, and UCP3 is reduced in diabetes. OBJECTIVE: We compared skeletal muscle UCP3 protein levels in prediabetic subjects [i.e. impaired glucose tolerance (IGT)], diabetic patients, and healthy controls and examined whether rosiglitazone treatment was able to restore UCP3. PATIENTS, DESIGN, INTERVENTION: Ten middle-aged obese men with type 2 diabetes mellitus [age, 61.4 +/- 3.1 yr; body mass index (BMI), 29.8 +/- 2.9 kg/m(2)], nine IGT subjects (age, 59.0 +/- 6.6 yr; BMI, 29.7 +/- 3.0 kg/m(2)), and 10 age- and BMI-matched healthy controls (age, 57.3 +/- 7.4 yr; BMI, 30.1 +/- 3.9 kg/m(2)) participated in this study. After baseline comparisons, diabetic patients received rosiglitazone (2 x 4 mg/d) for 8 wk. MAIN OUTCOME MEASURES: Muscle biopsies were sampled to determine UCP3 and mitochondrial protein (complex I-V) content. RESULTS: UCP3 protein content was significantly lower in prediabetic IGT subjects and in diabetic patients compared with healthy controls (39.0 +/- 28.5, 47.2 +/- 24.7, and 72.0 +/- 23.7 arbitrary units, respectively; P < 0.05), whereas the levels of the mitochondrial protein complex I-V were similar between groups. Rosiglitazone treatment for 8 wk significantly increased insulin sensitivity and muscle UCP3 content (from 53.2 +/- 29.9 to 66.3 +/- 30.9 arbitrary units; P < 0.05). CONCLUSION: We show that UCP3 protein content is reduced in prediabetic subjects and type 2 diabetic patients. Eight weeks of rosiglitazone treatment restores skeletal muscle UCP3 protein in diabetic patients.     

Skeletal muscle uncoupling protein-3 restores upon intervention in the prediabetic and diabetic state: implications for diabetes pathogenesis?

AIM: Skeletal muscle uncoupling protein-3 (UCP3) is reduced in type 2 diabetes, and in the pre-diabetic condition of impaired glucose tolerance (IGT). Here we examined whether intervention programs known to improve insulin sensitivity are paralleled by an increase in skeletal muscle UCP3 protein levels. METHODS: Skeletal muscle UCP3 protein content was measured before and after one year of an exercise intervention in muscle biopsies of eight diabetic subjects. In addition, UCP3 was measured in IGT subjects before and after 1 year of following a lifestyle-intervention program or serving as control. RESULTS: In the diabetic patients a significant increase of approximately 75% in UCP3 protein was found after 1 year of exercise training (P < 0.05). In IGT subjects UCP3 protein increased in the intervention group (P = 0.02), while UCP3 remained unaltered in the control group (P = 0.64). CONCLUSION: Both, exercise training and a lifestyle-intervention program increase UCP3 protein content in skeletal muscle of subjects with reduced glycaemic control, indicating a restoration towards normal UCP3 levels. These data support the idea that UCP3 has a role in the aetiology of type 2 diabetes mellitus.