We have developed an in vitro muscle preparation suitable for metabolic studies with human muscle tissue and have investigated the effects of obesity and non-insulin-dependent diabetes mellitus (NIDDM) on glucose transport. Transport of 3-0-methylglucose and 2-deoxyglucose was stimulated approximately twofold by insulin in muscle from normal nonobese subjects and stimulation occurred in the normal physiological range of insulin concentrations. In contrast to insulin stimulation of 3-0-methylglucose and 2-deoxyglucose transport in muscle from normal, nonobese subjects, tissue from morbidly obese subjects, with or without NIDDM, were not responsive to insulin. Maximal 3-0-methylglucose transport was lower in muscle of obese than nonobese subjects.Morbidly obese patients, with or without NIDDM, have a severe state of insulin resistance in glucose transport. The novel in vitro human skeletal muscle preparation herein described should be useful in investigating the mechanism of this insulin resistance.

Glucose transport in skeletal muscle is mediated by two distinct transporter isoforms, designated muscle/adipose glucose transporter (Glut4) and erythrocyte/HepG2/brain glucose transporter (Glut1), which differ in both abundance and membrane distribution. The present study was designed to investigate whether differences in insulin responsiveness of red and white muscle might be due to differential expression of the glucose transporter isoforms. Glucose transport, as well as Glut1 and Glut4 protein and mRNA levels, were determined in red and white portions of the quadriceps and gastrocnemius muscles of male Sprague-Dawley rats (body wt. approx. 250 g). Maximal glucose transport (in response to 100 nM-insulin) in the perfused hindlimb was 3.6 times greater in red than in white muscle. Red muscle contained approx. 5 times more total Glut4 protein and 2 times more Glut4 mRNA than white muscle, but there were no differences in the Glut1 protein or mRNA levels between the fibre types. Our data indicate that differences in responsiveness of glucose transport in specific skeletal muscle fibre types may be dependent upon the amount of Glut4 protein. Because this protein plays such an integral part in glucose transport in skeletal muscle, any impairment in its expression may play a role in insulin resistance.

Leptin, an adipocyte-derived hormone that directly regulates both adiposity and energy homeostasis, decreases food intake and appears to partition metabolic fuels toward utilization and away from storage. Because skeletal muscle expresses the leptin receptor and plays a major role in determining energy metabolism, we studied leptin's effects on glucose and fatty acid (FA) metabolism in isolated mouse soleus and extensor digitorum longus (EDL) muscles. One muscle from each animal served as a basal control. The contralateral muscle was treated with insulin (10 mU/ml), leptin (0.01-10 ug/ml), or insulin plus leptin, and incorporation of [ C]glucose or [14 C]oleate into CO 2 and into either glycogen or triacylglycerol (TAG) was determined. Leptin increased soleus muscle FA oxidation by 42% (P < 0.001) and decreased incorporation of FA into TAG by 35% (P < 0.01) in a dose-dependent manner. In contrast, insulin decreased soleus muscle FA oxidation by 40% (P < 0.001) and increased incorporation into TAG by 70% (P < 0.001). When both hormones were present, leptin attenuated both the antioxidative and the lipogenic effects of insulin by 50%. Less pronounced hormone effects were observed in EDL muscle. Leptin did not alter insulin-stimulated muscle glucose metabolism. These data demonstrate that leptin has direct and acute effects on skeletal muscle. Diabetes 46:1360-1363, 1997 H omozygous ob/ob mice lack functional leptin, a 16-kDa peptide hormone that is expressed and secreted by adipose tissue, and exhibit an obesity syndrome characterized by hyperphagia, hypothermia, hyperlipidemia, hyperinsulinemia, and insulin resistance (1). When administered to ob/ob mice, leptin reduces food intake and increases energy expenditure (2,3). Received for publication 28 March 1997 and accepted in revised form 7 May 1997.BSA, bovine serum albumin; EDL, extensor digitorum longus; FA, fatty acid; mKRB, modified Krebs-Ringer buffer; TAG, triacylglycerol.Both lean and obese animals treated with leptin lose fat mass but retain lean body mass (2,3). In ob/ob mice, leptin also normalizes serum concentrations of glucose, insulin, and lipids (2). The latter effects are observed at low leptin doses that do not affect body weight, suggesting that leptin's metabolic effects precede its effects on food intake and weight (2). In pair-feeding studies, leptin-injected mice lose 30-50% more weight and 50-100% more fat mass than pair-fed controls (4). Leptin treatment increases energy expenditure (2), whereas energy is conserved during food restriction (4,5). These data strongly suggest that leptin modulates energy homeostasis in part through mechanisms that are independent of food intake and that it directs metabolic fuels toward oxidation and away from storage.Skeletal muscle accounts for a large proportion of insulinstimulated glucose uptake and whole-body lipid oxidation and is the major tissue contributing to resting metabolic rate. Because each of these factors is affected by leptin in ob/ob mice (2-4) and because skeletal muscle expresses both lo...

Because muscle triacylglycerol (TAG) accumulation might contribute to insulin resistance in leptin-deficient ob/obmice, we studied the acute (60- to 90-min) effects of leptin and insulin on [14C]glucose and [14C]oleate metabolism in muscles isolated from lean and obese ob/ob mice. In ob/ob soleus, leptin decreased glycogen synthesis 36–46% ( P < 0.05), increased oleate oxidation 26% ( P < 0.05), decreased oleate incorporation into TAG 32% ( P < 0.05), and decreased the oleate partitioning ratio (oleate partitioned into TAG/CO2) 44% ( P < 0.05). Insulin decreased oleate oxidation 31% ( P < 0.05), increased oleate incorporation into TAG 46% ( P< 0.05), and increased the partitioning ratio 125% ( P < 0.01). Adding leptin diminished insulin’s antioxidative, lipogenic effects. In soleus from lean mice, insulin increased the partitioning ratio 142%, whereas leptin decreased it 51%, as previously reported (Muoio, D. M., G. L. Dohm, F. T. Fiedorek, E. B. Tapscott, and R. A. Coleman. Diabetes 46: 1360–1363, 1997). The phosphatidylinositol 3-kinase inhibitor wortmannin blocked insulin’s effects on lipid metabolism but only attenuated leptin’s effects. Increasing glucose concentration from 5 to 10 mM did not affect TAG synthesis, suggesting that insulin-induced lipogenesis is independent of increased glucose uptake. These data indicate that leptin opposes insulin’s promotion of TAG accumulation in lean and ob/ob muscles. Because acute leptin exposure does not correct insulin resistance in ob/ob muscles, in vivo improvements in glucose homeostasis appear to require other long-term factors, possibly TAG depletion.

A major defect contributing to impaired insulin action in human obesity is reduced glucose transport activity in skeletal muscle.This study was designed to determine whether the improvement in whole body glucose disposal associated with weight reduction is related to a change in skeletal muscle glucose transport activity and levels of the glucose transporter protein GLUT4. Seven morbidly obese (body mass index = 45.8±2.5, mean ± SE) patients, including four with non-insulin-dependent diabetes mellitus (NIDDM), underwent gastric bypass surgery for treatment of their obesity. In vivo glucose disposal during a euglycemic clamp at an insulin infusion rate of 40 mU/mi per min was reduced to 27% of nonobese controls (P < 0.01) and improved to 78% of normal after weight loss of 43.1±3.1 kg (P < 0.01). Maximal insulin-stimulated glucose transport activity in incubated muscle fibers was reduced by _ 50% in obese patients at the time of gastric bypass surgery but increased twofold (P < 0.01) to 88% of normal in five separate patients after similar weight reduction. Muscle biopsies obtained from vastus lateralis before and after weight loss revealed no significant change in levels of GLUT4 glucose transporter protein. These data demonstrate conclusively that insulin resistance in skeletal muscle of mobidly obese patients with and without NIDDM cannot be causally related to the cellular content of GLUT4 protein. The results further suggest that morbid obesity contributes to whole body insulin resistance through a reversible defect in skeletal muscle glucose transport activity. The mechanism for this improvement may involve enhanced transporter translocation and/or activation. (J. Clin.