Human Skeletal Muscle Protein Research Articles

Exercise does not alter subcellular localization, but increases phosphorylation of insulin-signaling proteins in human skeletal muscle

The subcellular localization of insulin signaling proteins is altered by various stimuli such as insulin, insulin-like growth factor I, and oxidative stress and is thought to be an important mechanism that can influence intracellular signal transduction and cellular function. This study examined the possibility that exercise may also alter the subcellular localization of insulin signaling proteins in human skeletal muscle. Nine untrained males performed 60 min of cycling exercise (approximately 67% peak pulmonary O2 uptake). Muscle biopsies were sampled at rest, immediately after exercise, and 3 h postexercise. Muscle was fractionated by centrifugation into the following crude fractions: cytosolic, nuclear, and a high-speed pellet containing membrane and cytoskeletal components. Fractions were analyzed for protein content of insulin receptor, insulin receptor substrate (IRS)-1 and -2, p85 subunit of phosphatidylinositol 3-kinase, Akt, and glycogen synthase kinase-3 (GSK-3). There was no significant change in the protein content of the insulin signaling proteins in any of the crude fractions after exercise or 3 h postexercise. Exercise had no significant effect on the phosphorylation of IRS-1 Tyr612 in any of the fractions. In contrast, exercise increased (P < 0.05) the phosphorylation of Akt Ser473 and GSK-3alpha/beta Ser9/21 in the cytosolic fraction only. In conclusion, exercise can increase phosphorylation of downstream insulin signaling proteins specifically in the cytosolic fraction but does not result in changes in the subcellular localization of insulin signaling proteins in human skeletal muscle. Change in the subcellular protein localization is therefore an unlikely mechanism to influence signal transduction pathways and cellular function in skeletal muscle after exercise.

Adipocyte Differentiation‐related Protein in Human Skeletal Muscle: Relationship to Insulin Sensitivity

To determine whether adipocyte differentiation-related protein (ADRP), a lipid droplet-associated protein that binds to and sequesters intracellular fatty acids, is 1) expressed in human skeletal muscle and 2) differentially regulated in human skeletal muscle obtained from obese non-diabetic (OND) and obese diabetic (OD) subjects. Ten OND subjects and 15 OD subjects underwent a weight loss or pharmacological intervention program to improve insulin sensitivity. Anthropometric data, hemoglobin A(1C), fasting glucose, lipids, and glucose disposal rate were determined at baseline and at completion of studies. Biopsies of the vastus lateralis muscle (SkM) were obtained in the fasting state from OND and OD subjects. Protein expression was determined by Western blotting. ADRP was highly expressed in SkM from OND (4.4 +/- 1.54 AU/10 microg, protein, n = 10) and OD (5.02 +/- 1.33 AU/10 microg, n = 12) subjects. OND subjects undergoing weight loss had decreased triglyceride levels and improved insulin action. SkM ADRP content increased with weight loss from 5.14 +/- 2.15 AU/10 microg to 9.92 +/- 1.57 AU/10 microg (p < 0.025). OD subjects were treated with either troglitazone or metformin, together with glyburide, for 3 to 4 months. Both treatments attained similar levels of glycemic control. OD subjects with lower baseline ADRP content (2.85 +/- 1.07 AU/10 microg, n = 6) displayed up-regulation of ADRP expression (to 9.27 +/- 2.76 AU/10 microg, p < 0.025). ADRP is the predominant lipid droplet-associated protein in SkM, and low ADRP expression is up-regulated in circumstances of improved glucose tolerance. Up-regulation of ADRP may act to sequester fatty acids as triglycerides in discrete lipid droplets that could protect muscle from the detrimental effects of fatty acids on insulin action and glucose tolerance.

Dietary protein intake impacts human skeletal muscle protein fractional synthetic rates after endurance exercise

This investigation evaluated the physiological impact of different dietary protein intakes on skeletal muscle protein synthesis postexercise in endurance runners. Five endurance-trained, male runners participated in a randomized, crossover design diet intervention, where they consumed either a low (0.8 g/kg; LP)-, moderate (1.8 g/kg; MP)-, or high (3.6 g/kg; HP)-protein diet for 4 wk. Diets were designed to be eucaloric with carbohydrate, fat, and protein approximating 60, 30, and 10%; 55, 30, and 15%; and 40, 30, and 30% for LP, MP, and HP, respectively. Substrate oxidation was assessed via indirect calorimetry at 3 wk of the dietary interventions. Mixed-muscle protein fractional synthetic rate (FSR) was measured after an endurance run (75 min at 70% V(O2 peak)) using a primed, continuous infusion of [(2)H(5)]phenylalanine. Protein oxidation increased with increasing protein intake, with each trial being significantly different from the other (P < 0.01). FSR after exercise was significantly greater for LP (0.083%/h) and MP (0.078%/h) than for HP (0.052%/h; P < 0.05). There was no difference in FSR between LP and MP. This is the first investigation to establish that habitual dietary protein intake in humans modulates skeletal muscle protein synthesis after an endurance exercise bout. Future studies directed at mechanisms by which level of protein intake influences skeletal muscle turnover are needed.

Impact of surgical trauma on human skeletal muscle protein synthesis.

Muscle protein catabolism is a considerable clinical problem following surgery. However, the impact of surgical trauma on muscle protein synthesis is not well characterized. In this pilot study, we therefore investigated whether the severity of surgical trauma is related to a decrease in muscle protein synthesis rate in humans. Metabolically healthy patients (n=28) were included in the study. Eight of the patients were day-care patients undergoing minor breast surgery (defined as minor surgery). The other 20 patients were subjected to major abdominal surgery and were therefore scheduled to stay overnight in the recovery room during the first postoperative night (defined as major surgery). Protein FSRs (fractional synthesis rates) in skeletal muscle were determined during a measurement period of 90 min before surgery and immediately after termination of surgery. FSR in skeletal muscle of the minor surgery patients was 1.72+/-0.25%/24 h before surgery and 1.67+/-0.29%/24 h after surgery (P=0.68). In the major surgery group, FSR was 1.62+/-0.30%/24 h before surgery and 1.57+/-0.40%/24 h (P=0.59) immediately following surgery. The observations made in this pilot study could not confirm a size-related decrease in muscle protein synthesis immediately following minor and major surgery. This finding is discussed in relation to confounders, postoperative course and to muscle protein degradation. The shortage of knowledge in this field is emphasized.

Expression of Na+/HCO3- co-transporter proteins (NBCs) in rat and human skeletal muscle.

Sodium/bicarbonate co-transport (NBC) has been suggested to have a role in muscle pH regulation. We investigated the presence of NBC proteins in rat and human muscle samples and the fibre type distribution of the identified NBCs. Western blotting of muscle homogenates and sarcolemmal membranes (sarcolemmal giant vesicles) were used to screen for the presence of NBCs. Immunohistochemistry was used for the subcellular localization. The functional test revealed that approximately half of the pH recovery in sarcolemmal vesicles produced from rat muscle is mediated by bicarbonate-dependent transport. This indicates that the NBCs are preserved in the vesicles. The western blotting experiments demonstrated the existence of at least two NBC proteins in skeletal muscle. One NBC protein (approximately 150 kDa) seems to be related to the kidney/pancreas/heart isoform NBC1, whereas the other protein (approximately 200 kDa) is related to the NBC4 isoform. The two NBC proteins represent the electrogenic isoforms named NBCe1 and NBCe2. Membrane fractionation and immunofluorescence techniques confirmed that the two NBCs are located in the sarcolemmal membrane as well as in some internal membranes, probably the T-tubules. The two NBCs localized in muscle have distinct fibre type distributions. Skeletal muscle possesses two variants of the sodium/bicarbonate co-transporter (NBC) isoforms, which have been called NBCe1 and NBCe2.

Studies of plasma membrane fatty acid-binding protein and other lipid-binding proteins in human skeletal muscle.

The first putative fatty acid transporter identified was plasma membrane fatty acid-binding protein (FABPpm). Later it was demonstrated that this protein is identical to the mitochondrial isoform of the enzyme aspartate aminotransferase. In recent years data from several cell types have emerged, indicating that FABPpm plays a role in the transport of long-chain saturated and unsaturated fatty acids. In the limited number of studies in human skeletal muscle it has been demonstrated that dietary composition and exercise training can influence the content of FABPpm. Ingestion of a fat-rich diet induces an increase in FABPpm protein content in human skeletal muscle in contrast to the decrease seen during consumption of a carbohydrate-rich diet. A similar effect of a fat-rich diet is also observed for cytosolic fatty acid-binding protein and fatty acid translocase/CD36 protein expression. Exercise training up regulates FABPpm protein content in skeletal muscle, but only in male subjects; no significant differences were observed in muscle FABPpm content in a cross-sectional study of female volunteers of varying training status, even though muscle FABPpm content did not depend on gender in the untrained state. A higher utilization of plasma long-chain fatty acids during exercise in males compared with females could explain the gender-dependent influence of exercise training on FABPpm. The mechanisms involved in the regulation of the function and expression of FABPpm protein remain to be clarified.

Muscle- and fibre type-specific expression of glucose transporter 4, glycogen synthase and glycogen phosphorylase proteins in human skeletal muscle.

The muscle- and fibre type-specific expression of skeletal muscle glucose transporter 4 (GLUT4), glycogen synthase (GS) and glycogen phosphorylase (GP) was investigated in six young male subjects. Single muscle fibres were dissected from vastus lateralis (VL), soleus (SO) and triceps brachii (TB) muscle biopsy samples. On the basis of myosin heavy chain (MHC) expression, fibres were pooled into three groups (MHC I, MHC IIA and MHC IIX) and the GLUT4, GS and GP content of 15-40 pooled fibres determined using SDS-PAGE and immunological detection. In VL, the GLUT4 content in the pooled muscle fibres expressing MHC I was approximately 33% higher ( P<0.05) than in fibres expressing MHC IIA or IIX. There was no difference in GLUT4 content between fibres expressing MHC IIA or IIX, nor were there any differences in GS and GP content between any of the fibre types. In SO, there was no difference in GLUT4, GS and GP between fibres expressing MHC I or IIA. No fibres expressing type IIX were detected. In TB, fibres expressing MHC IIA and IIX had significantly ( P<0.05) more GP (66% and 55 % in MHC IIA and MHCIIX, respectively) than those expressing MHC I, whilst there was no difference in GP between MHC IIA and MHC IIX fibres. The GLUT4 and the GS content was similar in fibres expressing MHC I, IIA and IIX in the TB. Our data directly demonstrate that some proteins, like GLUT4 and GP, are expressed in a fibre type-specific manner in some, but not all, muscles, whilst other proteins, like GS, are not. In human skeletal muscle the GLUT4, GS and GP content thus seems to be related primarily to factors other than the fibre type as defined by the expression of contractile protein. These findings imply that it is not possible to generalize fibre type-dependent protein expression on the basis of biopsies from only one muscle.

Effect of insulin on human skeletal muscle mitochondrial ATP production, protein synthesis, and mRNA transcripts

Mitochondria are the primary site of skeletal muscle fuel metabolism and ATP production. Although insulin is a major regulator of fuel metabolism, its effect on mitochondrial ATP production is not known. Here we report increases in vastus lateralis muscle mitochondrial ATP production capacity (32-42%) in healthy humans (P < 0.01) i.v. infused with insulin (1.5 milliunits/kg of fat-free mass per min) while clamping glucose, amino acids, glucagon, and growth hormone. Increased ATP production occurred in association with increased mRNA levels from both mitochondrial (NADH dehydrogenase subunit IV) and nuclear [cytochrome c oxidase (COX) subunit IV] genes (164-180%) encoding mitochondrial proteins (P < 0.05). In addition, muscle mitochondrial protein synthesis, and COX and citrate synthase enzyme activities were increased by insulin (P < 0.05). Further studies demonstrated no effect of low to high insulin levels on muscle mitochondrial ATP production for people with type 2 diabetes mellitus, whereas matched nondiabetic controls increased 16-26% (P < 0.02) when four different substrate combinations were used. In conclusion, insulin stimulates mitochondrial oxidative phosphorylation in skeletal muscle along with synthesis of gene transcripts and mitochondrial protein in human subjects. Skeletal muscle of type 2 diabetic patients has a reduced capacity to increase ATP production with high insulin levels.

Human skeletal muscle and erythrocyte proteins involved in acid-base homeostasis: adaptations to chronic hypoxia

Human skeletal muscle and erythrocyte proteins involved in acid-base homeostasis: adaptations to chronic hypoxia

Human skeletal muscle and erythrocyte proteins involved in acid-base homeostasis: adaptations to chronic hypoxia

Chronic hypoxia is accompanied by changes in blood and skeletal muscle acid-base control. We hypothesized that the underlying mechanisms include altered protein expression of transport systems and the enzymes involved in lactate, HCO3- and H+ fluxes in skeletal muscle and erythrocytes. Immunoblotting was used to quantify densities of the transport systems and enzymes. Muscle and erythrocyte samples were obtained from eight Danish lowlanders at sea level and after 2 and 8 weeks at 4100 m (Bolivia). For comparison, samples were obtained from eight Bolivian natives. In muscle membranes there were no changes in fibre-type distribution, lactate dehydrogenase isoforms, Na+,K+-pump subunits or in the lactate-H+ co-transporters MCT1 and MCT4. The Na+-H+ exchanger protein NHE1 was elevated by 39 % in natives compared to lowlanders. The Na+-HCO3- co-transporter density in muscle was elevated by 47-69 % after 2 and 8 weeks at altitude. The membrane-bound carbonic anhydrase (CA) IV in muscle increased in the lowlanders by 39 %, whereas CA XIV decreased by 23-47 %. Levels of cytosolic CA II and III in muscle and CA I and II in erythrocytes were unchanged. The erythrocyte lactate-H+ co-transporter MCT1 increased by 230-405 % in lowlanders and was 324 % higher in natives. The erythrocyte inorganic anion exchanger (Cl--HCO3- exchanger AE1) was increased by 149-228 %. In conclusion, chronic hypoxia induces dramatic changes in erythrocyte proteins, but only moderate changes in muscle proteins involved in acid-base control. Together, these changes suggest a hypoxia-induced increase in the capacity for lactate, HCO3- and H+ fluxes from muscle to blood and from blood to erythrocytes.

Human skeletal muscle creatine transporter mRNA and protein expression in healthy, young males and females.

The present study investigated whether there were any differences between males and females in respect to creatine transporter (CreaT) gene expression and/or total creatine (TCr) content in human vastus lateralis muscle. Skeletal muscle obtained from young healthy male (n = 13, age: 23.2 +/- 5.0 years) and female subjects (n = 12, age: 21.7 +/- 4.3 years) was analyzed for CreaT mRNA, CreaT protein and TCr content. Total CreaT protein content in the muscle was similar (p > 0.05) between the sexes. Two bands (approximately 55 and 73 kDa) of the CreaT protein were detected in all muscle samples. Both the 55 and the 73 kDa bands were present in similar (p > 0.05) amounts in males compared with females. The 73 kDa band was in greater abundance (p < 0.05) than the 55 kDa band, irrespective of gender. In addition, CreaT mRNA expression relative to beta-actin mRNA and the TCr content (males: 117.8 +/- 2.2, females: 125.3 +/- 4.3 mmol.kg(-1) dry mass) were also unaffected (p > 0.05) by gender. These data demonstrate that gender does not influence skeletal muscle TCr content and CreaT gene expression in young human subjects.

Branched Chain Amino Acids Activate Messenger Ribonucleic Acid Translation Regulatory Proteins in Human Skeletal Muscle, and Glucocorticoids Blunt This Action

Branched Chain Amino Acids Activate Messenger Ribonucleic Acid Translation Regulatory Proteins in Human Skeletal Muscle, and Glucocorticoids Blunt This Action

Branched chain amino acids activate messenger ribonucleic acid translation regulatory proteins in human skeletal muscle, and glucocorticoids blunt this action.

Branched chain amino acids (BCAA) are particularly effective anabolic agents. Recent in vitro studies suggest that amino acids, particularly leucine, activate a signaling pathway that enhances messenger ribonucleic acid translation and protein synthesis. The physiological relevance of these findings to normal human physiology is uncertain. We examined the effects of BCAA on the phosphorylation of eukaryotic initiation factor 4E-binding protein 1 (eIF4E-BP1) and ribosomal protein S6 kinase (p70(S6K)) in skeletal muscle of seven healthy volunteers. We simultaneously examined whether BCAA affect urinary nitrogen excretion and forearm skeletal muscle protein turnover and whether the catabolic action of glucocorticoids could be mediated in part by inhibition of the action of BCAA on the protein synthetic apparatus. BCAA infusion decreased urinary nitrogen excretion (P < 0.02), whole body phenylalanine flux (P < 0.02), plasma phenylalanine concentration (P < 0.001), and improved forearm phenylalanine balance (P = 0.03). BCAA also increased the phosphorylation of both eIF4E-BP1 (P < 0.02) and p70(S6K) (P < 0.03), consistent with an action to activate the protein synthetic apparatus. Dexamethasone increased plasma phenylalanine concentration (P < 0.001), prevented the BCAA-induced anabolic shift in forearm protein balance, and inhibited their action on the phosphorylation of p70(S6K). We conclude that in human skeletal muscle BCAA act directly as nutrient signals to activate messenger ribonucleic acid translation and potentiate protein synthesis. Glucocorticoids interfere with this action, and that may be part of the mechanism by which they promote net protein catabolism in muscle.

Metabolic effects of vanadyl sulfate in humans with non—insulin-dependent diabetes mellitus: In vivo and in vitro studies

To investigate the efficacy and mechanism of action of vanadium salts as oral hypoglycemic agents, 16 type 2 diabetic patients were studied before and after 6 weeks of vanadyl sulfate (VOSO4) treatment at three doses. Glucose metabolism during a euglycemic insulin clamp did not increase at 75 mg/d, but improved in 3 of 5 subjects receiving 150 mg VOSO4 and 4 of 8 subjects receiving 300 mg VOSO4. Basal hepatic glucose production (HGP) and suppression of HGP by insulin were unchanged at all doses. Fasting glucose and hemoglobin A1c (HbA1c) decreased significantly in the 150- and 300-mg VOSO4 groups. At the highest dose, total cholesterol decreased, associated with a decrease in high-density lipoprotein (HDL). There was no change in systolic, diastolic, or mean arterial blood pressure on 24-hour ambulatory monitors at any dose. There was no apparent correlation between the clinical response and peak serum level of vanadium. The 150- and 300-mg vanadyl doses caused some gastrointestinal intolerance but did not increase tissue oxidative stress as assessed by thiobarbituric acid-reactive substances (TBARS). In muscle obtained during clamp studies prior to vanadium therapy, insulin stimulated the tyrosine phosphorylation of the insulin receptor, insulin receptor substrate-1 (IRS-1), and Shc proteins by 2- to 3-fold, while phosphatidylinositol 3-kinase (PI 3-kinase) activity associated with IRS-1 increased 4.7-fold during insulin stimulation (P = .02). Following vanadium, there was a consistent trend for increased basal levels of insulin receptor, Shc, and IRS-1 protein tyrosine phosphorylation and IRS-1-associated PI 3-kinase, but no further increase with insulin. There was no discernible correlation between tyrosine phosphorylation patterns and glucose disposal responses to vanadyl. While glycogen synthase fractional activity increased 1.5-fold following insulin infusion, there was no change in basal or insulin-stimulated activity after vanadyl. There was no increase in the protein phosphatase activity of muscle homogenates to exogenous substrate after vanadyl. Vanadyl sulfate appears safe at these doses for 6 weeks, but at the tolerated doses, it does not dramatically improve insulin sensitivity or glycemic control. Vanadyl modifies proteins in human skeletal muscle involved in early insulin signaling, including basal insulin receptor and substrate tyrosine phosphorylation and activation of PI 3-kinase, and is not additive or synergistic with insulin at these steps. Vanadyl sulfate does not modify the action of insulin to stimulate glycogen synthesis. Since glucose utilization is improved in some patients, vanadyl must also act at other steps of insulin action.

Effects of exercise and insulin on insulin signaling proteins in human skeletal muscle.

Insulin and exercise independently increase glucose metabolism in muscle. Moreover, exercise training or a prior bout of exercise increases insulin-stimulated glucose uptake in resting skeletal muscle. The present study was undertaken to compare how physiological hyperinsulinemia and moderate intensity aerobic exercise affect the tyrosine phosphorylation state and activity of insulin signaling molecules in healthy, physically inactive volunteers. Subjects had biopsies of the vastus lateralis muscle before and immediately after 30 min of either hyperinsulinemia (euglycemic insulin clamp) or moderate-intensity exercise on a cycle ergometer (approximately 60% of VO2max). Insulin receptor and IRS-1 tyrosine phosphorylation, association of the p85 regulatory subunit of PI 3-kinase with IRS-1, IRS-1 associated PI 3-kinase activity, and glycogen synthase activity were determined in muscle biopsy specimens taken from healthy subjects before and after insulin or exercise. Physiological hyperinsulinemia increased the rate of glucose disposal from 11.4 +/- 1.5 to 25.6 +/- 6.7 micromol x kg(-1) x min(-1) (P < 0.01), insulin receptor and IRS-1 tyrosine phosphorylation (173 +/- 19% and 159 +/- 35% of basal values, respectively, P < 0.05), association of the p85 regulatory subunit of PI 3-kinase with IRS-1 (159 +/- 10%, P < 0.05), and glycogen synthase fractional velocity (136 +/- 11%, P < 0.01). Exercise also increased glucose disposal, from 10.4 +/- 0.5 to 15.6 +/- 1.7 micromol x kg(-1) x min(-1) (P < 0.01) and glycogen synthase fractional velocity (253 +/- 35% of basal, P < 0.01). The exercise-induced increase in glycogen synthase was greater than that due to insulin (P < 0.05). In contrast to insulin, exercise decreased tyrosine phosphorylation of the insulin receptor to 72 +/- 10% of basal values (P < 0.05 vs basal and P < 0.05 vs insulin) and had no effect on IRS-1 tyrosine phosphorylation, or association of p85 with IRS-1. The exercise-induced decreased insulin receptor tyrosine phosphorylation could explain the well-known effect of exercise to enhance the sensitivity of muscle to insulin.

Presence in human skeletal muscle of an AMP deaminase-associated protein that reacts with an antibody to human plasma histidine-proline-rich glycoprotein.

Histidine-proline-rich glycoprotein (HPRG) is a protein that is synthesized by parenchimal liver cells. The protein has been implicated in a number of plasma-specific processes, including blood coagulation and fibrinolysis. We have recently reported the association of an HPRG-like protein with rabbit skeletal muscle AMP deaminase (AMPD). The results of the immunological analysis reported here demonstrate that an antibody against human plasma HPRG reacts with an AMPD preparation from human skeletal muscle. To probe the localization of the putative HPRG-like protein in human skeletal muscle, serial sections from frozen biopsy specimens were processed for immunohistochemical and histoenzymatic stains. A selective binding of the anti-HPRG antibody to Type IIB muscle fibers was detected, suggesting a preferential association of the novel protein to the AMPD isoenzyme contained in the fast-twitch glycolytic fibers.

Age-dependent increases in oxidative damage to DNA, lipids, and proteins in human skeletal muscle

A role for oxidative damage in normal aging is supported by studies in experimental animals, but there is limited evidence in man. We examined markers of oxidative damage to DNA, lipids, and proteins in 66 muscle biopsy specimens from humans aged 25 to 93 years. There were age-dependent increases in 8-hydroxy-2-deoxyguanosine (OH 8dG), a marker of oxidative damage to DNA, in malondialdehyde (MDA), a marker of lipid peroxidation, and to a lesser extent in protein carbonyl groups, a marker of protein oxidation. The increases in OH 8dG were significantly correlated with increases in MDA. These results provide evidence for a role of oxidative damage in human aging which may contribute to age-dependent losses of muscle strength and stamina.

Does Growth Hormone Therapy in Conjunction With Resistance Exercise Increase Muscle Force Production and Muscle Mass in Men and Women Aged 60 Years or Older?

Improved muscle protein mass and increments in maximum voluntary muscle force have rarely been observed in men and women aged 60 years and older who were treated with rhGH. Although rhGH administration has been reported to increase lean body mass in older men and women, it is doubtful that this increase is localized to skeletal muscle contractile proteins. When rhGH administration was combined with 16 weeks of resistance exercises, increases in muscle mass, muscle protein synthesis, and muscle force were not greater in the rhGH-treated group than in a weight training group that received placebo injections. Side effects of rhGH treatment in elderly people are prevalent, not trivial, and further limit its usefulness as an effective anabolic agent for promoting muscle protein accretion in men and women. In particular, the induction of insulin resistance and carpal tunnel compression reduces the efficacy of rhGH replacement therapy in elderly individuals. The evidence for a GH-induced increase in human skeletal muscle protein and maximum voluntary muscle force is weak. The optimum dose and GH-replacement paradigm (GHRH, GH-secretagogues) have not been identified. Whether rhGH therapy improves muscle protein mass and force in individuals with severe cachexia associated with major trauma, burns, surgery, or muscular dystrophy is controversial and under investigation.

Expression of mRNAs encoding uncoupling proteins in human skeletal muscle: effects of obesity and diabetes.

To explore the potential role of the uncoupling protein (UCP) family in human obesity and diabetes, we have used the reverse transcription-polymerase chain reaction to quantify UCP mRNA expression in human skeletal muscle. Levels of mRNA for UCP2, and for both short (UCP3S) and long (UCP3L) forms of UCP3, were highly correlated in individuals, indicating that gene transcription of these UCPs may be coordinately regulated by common mechanisms. In normal glucose-tolerant individuals, muscle UCP2 mRNA levels were positively correlated with percentage of body fat and with BMI (r = 0.6 and P < 0.05 for both). UCP3S mRNA levels were also positively correlated with percentage of body fat (r = 0.52, P < 0.05), and UCP3L mRNA tended to increase as a function of obesity (0.05 < P < 0.1). UCP mRNA levels, however, were not correlated with resting metabolic rate. UCP3S and UCP3L mRNA levels (P < 0.05) and the UCP2 mRNA level (P = 0.09) were increased by 1.8- to 2.7-fold in type 2 diabetes, an effect that could not be explained by obesity. No significant difference was found for UCP2, UCP3S, or UCP3L mRNA levels between insulin-sensitive and insulin-resistant nondiabetic subgroups. We conclude that 1) skeletal muscle mRNA levels encoding UCP2 and UCP3 are correlated among individuals and may be coordinately regulated; 2) UCP3 expression is not regulated by differential effects on UCP3L and UCP3S forms of the mRNA; and 3) UCP mRNA expression tends to increase in muscle as a function of obesity but not of resting metabolic rate or insulin resistance, and is increased in patients with type 2 diabetes.

Emerin, deficiency of which causes Emery-Dreifuss muscular dystrophy, is localized at the inner nuclear membrane.

X-linked recessive Emery-Dreifuss muscular dystrophy (EDMD) is an inherited muscle disorder characterized by the clinical triad of progressive wasting of humero-peroneal muscles, early contractures of the elbows, Achilles tendons and postcervical muscles, and cardiac conduction block with a high risk of sudden death. The gene for EDMD on Xq28 encodes a novel protein named emerin that localizes at the nuclear membrane of skeletal, cardiac and smooth muscles and some other non-muscle tissues. To investigate a possible physiological role for emerin, we examined the ultrastructural localization of the protein in human skeletal muscle and HeLa cells, using ultrathin cryosections. We found that the immune-labeled colloidal gold particles were localized on the nucleoplasmic surface of the inner nuclear membrane, but not on the nuclear pore. Emerin stayed on the cytoplasmic surface of the nuclear lamina, even after detergent treatment that solubilizes membrane lipids and washes out membrane proteins. These results suggest that emerin anchors at the inner nuclear membrane through the hydrophobic stretch, and protrudes from the hydrophilic region to the nucleoplasm where it interacts with the nuclear lamina. We speculate that emerin contributes to maintain the nuclear structure and stability, as well as nuclear functions, particularly in muscle tissues that have severe stress with rigorous contraction-relaxation movements and calcium flux.