Elisabeth Hjøllund

Insulin receptor binding and insulin action in human fat cells: Effects of obesity and fasting

We have studied (125I)-insulin binding and insulin dose response relationships of (14C)-methylglucose transport conversion of (14C)-glucose to CO2 and total lipids, and lipolysis at 37 degrees C and pH 7.4 in adipocytes from obese patients before (n = 15) and after fasting for 10 days (n = 6). Studies of adipocytes from obese before fasting showed a significant reduction of insulin binding when expressed to cell surface area and rightward shifts of the insulin dose response curves (decreased insulin sensitivity) for glucose transport, glucose oxidation, lipogenesis and antilipolysis. The decreased insulin sensitivity of adipocytes from obese was most likely the functional consequence of the impaired insulin binding. Moreover, decreased maximal glucose transport capacities were present in rat cells from obese both in the basal and maximally insulin stimulated states. Similarly, the percentage response above basal level to maximal insulin stimulation of glucose oxidation and lipogenesis was impaired to these cells. The latter findings suggest post receptor defects localized both to the transport system per se and to intracellular mechanisms involved in the metabolism of glucose. Conversely, the post receptor pathways for the insulin induced antilipolysis was intact in fat cells from obese man. Studies after fasting showed an increase of adipocyte insulin binding accompanied by an increased sensitivity to the antilipolytic effect of insulin with unchanged maximal responsiveness. However, due to marked post receptor alterations, the insulin stimulated glucose utilization was severely blunted. Thus, the glucose transport system of adipocytes from all fasted subjects was totally unresponsive to insulin, while some of the fasted patients had a slight response of glucose oxidation and lipogenesis in the presence of insulin in maximally effective concentrations.

Increased insulin binding to adipocytes and monocytes and increased insulin sensitivity of glucose transport and metabolism in adipocytes from non-insulin-dependent diabetics after a low-fat/high-starch/high-fiber diet

Nine non-insulin-dependent diabetics were studied before and after 3 weeks on an isoenergetic high-fiber/high-starch/low-fat diet (alternative diet), and nine non-insulin-dependent diabetics were studied on their usual diet. In the group that ate the alternative diet, the intake of fiber and starch increased 120% and 53%, whereas fat intake decreased 31%. Diabetes control improved as demonstrated by decreased fasting plasma glucose (P less than 0.05) and 24-hour urinary glucose excretion (P less than 0.05). The in vivo insulin action increased (KIVITT increased, P less than 0.05) with no change in fasting serum insulin levels. In fat cells obtained from patients in the alternative-diet group, insulin receptor binding increased (P less than 0.05) after the change of diet. Insulin binding to purified monocytes (more than 95% monocytes) also increased (P less than 0.05), whereas no change was found in insulin binding to erythrocytes. When lipogenesis was studied at a tracer glucose concentration at which glucose transport seems to be rate limiting, insulin sensitivity increased (P less than 0.02). This is the predicted consequence of increased receptor binding. Moreover, when CO2 production and lipogenesis were studied at a higher glucose concentration, where steps beyond transport seem to be rate limiting for glucose metabolism, increased insulin sensitivity was also observed. In contrast, no change was found in maximal insulin responsiveness. Fat and blood cells from the patients who continued on their usual diet showed no changes of the mentioned quantities.(ABSTRACT TRUNCATED AT 250 WORDS)

Insulin Receptor Binding to Fat and Blood Cells and Insulin Action in Fat Cells from Insulin-dependent Diabetics

Insulin binding to fat and blood cells and the effect of insulin on glucose transport, glucose oxidation, lipogenesis, and lipolysis in fat cells were measured in 23 conventionally treated insulin-dependent diabetics (IDDs) with a mean duration of diabetes for 10 yr. In the fasting state diabetics had increased plasma levels of free insulin (P < 0.001) accompanied by a 38% reduction of insulin binding to fat cells (P < 0.001), whereas insulin binding to monocytes and erythrocytes was normal. The concentrations of insulin causing halfmaximal effect on glucose transport and antilipolysis in adipocytes were both higher in diabetics than in normals (P < 0.001), suggesting impaired sensitivity to insulin that may be the functional consequence of the adipocyte insulin receptor defect. The basal rates and the maximal responses to insulin of glucose transport and lipolysis were both within the limits of normals, indicating the absence of postreceptor abnormalities in these effector systems. In contrast, both glucose oxidation and lipogenesis in fat cells from diabetics had normal sensitivity to insulin. This finding was very surprising and suggests possible postreceptor modulation o the insulin sensitivity of glucose metabolism in insulin-treated diabetics counterbalancing the effects of decreased insulin receptor binding. Moreover, both the basal rates for glucose oxidation and lipogenesis and the absolute response of glucose oxidation and lipogenesis rates to maximal insulin stimulation were significantly reduced in fat cells from diabetics indicating postreceptor impairments of glucose metabolism. In conclusion: In long-term-treated IDDs in usual metabolic control and with fasting, peripheral hyperinsulinemia insulin binding to circulating blood cells was normal. In contrast, a peripheral, fixed tissue, as fat cells, exhibited decreased insulin bindingwith impaired insulin sensitivity of some pathways (glucose transport and antilipolysis) and postreceptor impairments of other pathways (lipogenesis and glucose oxidation).

Impaired insulin receptor binding and postbinding defects of adipocytes from normal and diabetic pregnant women

To evaluate the relative contribution of insulin binding and postbinding defects of glucose utilization in peripheral tissue during normal and diabetic pregnancy, we have studied the in vitro insulin action of isolated adipocytes from eight nondiabetic pregnant women and nine pregnant women with insulin-dependent diabetes mellitus who were undergoing cesarian section. The pregnant women were compared with a matched group of normal nonpregnant women undergoing gynecologic surgery. Insulin binding to adipocytes measured at tracer insulin concentration was reduced by 45% (P < 0.01) in normal pregnant women and by 30% (P < 0.02) in pregnant women with diabetes. In contrast, no changes were found between the three groups in insulin binding to pure monocytes and erythrocytes. The glucose transport system in fat cells from both groups of pregnant women was characterized by impaired maximal (P < 0.05) and half-maximal (P < 0.05) response to insulin. When fat cell glucose metabolism was studied, pregnant diabetic women exhibited decreased basal lipogenesis (P < 0.05) and decreased maximal responses of lipogenesis and glucose oxidation to insulin stimulation (P < 0.05). Similar but less pronounced abnormalities were seen in glucose metabolism of adipocytes from nondiabetic pregnant women. In conclusion, both in late normal and diabetic pregnancy, insulin binding to adipocytes is significantly reduced and accompanied by decreased insulin sensitivity and reduced maximal insulin responsiveness of glucose transport and by impaired basal and maximally insulin-stimulated glucose metabolism.

Effects of sulfonylureas on adipocyte and skeletal muscle insulin action in patients with non-insulin-dependent diabetes mellitus

The effect of glibenclamide treatment on insulin action in isolated fat cells was studied in eight moderately obese patients with non-insulin-dependent diabetes mellitus (NIDDM). Insulin receptor binding and the effect of insulin on glucose transport and lipogenesis were determined before and after 3 months of glibenclamide therapy. At the end of the treatment period, mean daytime plasma glucose concentrations were reduced (10.8 +/- 0.4 versus 7.0 +/- 0.3 mmol/L, p less than 0.001) whereas mean daytime plasma insulin level was increased (40 +/- 12 versus 71 +/- 9 mU/L, p less than 0.001). Adipocyte insulin receptor binding as well as basal glucose transport and metabolism were unaffected by drug treatment. In contrast, insulin-stimulated glucose transport and lipogenesis were both significantly enhanced (p less than 0.05). These findings are comparable to those of another study involving seven moderately obese subjects with NIDDM who had biopsies of the lateral vastus muscle taken for measurement of insulin receptor function and glycogen synthase activity before and during 2 months of gliclazide treatment. In that study insulin receptors purified with wheatgerm agglutinin showed unchanged insulin binding and receptor kinase activity. Moreover, gliclazide had no impact on maximal glycogen synthase activity. However, under physiologic hyperinsulinemic conditions gliclazide therapy was associated with an increased sensitivity of glycogen synthase for its allosteric activation by glucose-6-phosphatase (p less than 0.04). In conclusion, sulfonylurea treatment of NIDDM enhances insulin-stimulated peripheral glucose utilization in part through a potentiation of insulin action on adipose tissue glucose transport and lipogenesis and skeletal muscle glycogen synthase.

Transport and metabolism of d-glucose in human adipocytes. Studies of the dependence on medium glucose and insulin concentrations

Uptake and metabolism of the physiologically labelled D-glucose (D-[U-14C]glucose) has been characterized in human adipocytes at several unlabelled D-glucose concentrations in the absence and presence of insulin. Following a 90 min incubation, about 80% of the intracellular radioactivity was incorporated into total lipids at tracer glucose concentration, as well as at higher glucose concentrations in basal and insulin-stimulated cells, whereas 20% was recovered as hydrophilic metabolites. The only 14C-labelled metabolite escaping the cells in detectable amounts was CO2, which accounted about 4%. At trace glucose concentrations (5 mumol/l), the rate of glucose uptake was linear with time. Comparative studies of initial glucose uptake after 10 s and tracer D-glucose conversion to total lipids after 90 min showed high coefficients of correlation between basal rates (r = 0.87), maximal response above basal level to insulin (r = 0.92) and insulin sensitivity (r = 0.78). Thus, under these conditions glucose transport is rate-limiting for net glucose uptake, and measurements over long time intervals of rates for total cell-associated radioactivity or lipogenesis may serve as reliable estimates of initial glucose influx rates. However, the conversion rate of tracer glucose to metabolites decreased progressively with the glucose concentration and with an apparent Km of about 0.2 mmol/l. The three metabolic pathways exhibited similar percentage decreases in their activities, suggesting that a common enzymatic step is rate-limiting. In comparison, the Km for initial D-glucose uptake rate was about 7 mmol/l. Hence, the capacity for total glucose metabolism comprised only a small fraction of the glucose transport capacity at medium glucose concentrations above tracer concentrations. Both basal, half-maximal and maximal insulin-stimulated rates of adipocyte glucose utilization were dependent on the glucose concentration. Thus, comparing lipogenesis at tracer and at 0.5 mmol/l medium glucose concentration, it was shown that the higher medium glucose concentration was associated with a 60% lowering of the basal rate, a 35% reduction in the percentage response above baseline to maximal insulin stimulation and a 4-fold increase in the insulin sensitivity. Obviously, these findings reflect some intracellular step(s) being rate-limiting at medium glucose levels above tracer values.

Adipocyte insulin binding and action in moderately obese NIDDM patients after dietary control of plasma glucose: reversal of postbinding abnormalities.

Studies of fat cells from patients with newly diagnosed, untreated non-insulin-dependent diabetes mellitus (NIDDM) have revealed severe abnormalities in insulin action on glucose transport and metabolism. To determine whether these defects can be reversed if good glycemic control is reached by dietary treatment, eight moderately obese NIDDM subjects were studied at diagnosis and again when the patients had been in good glycemic control induced by low-energy dieting for at least 2 mo (absence of glycosuria and fasting plasma glucose <7 mM). Average body weight decreased by 8 kg (P < .05). Fasting plasma glucose decreased from 11.5 ± 1.2 to 6.9 ± 0.9 mM, whereas fasting serum insulin concentrations were unchanged. Adipocyte insulin binding at tracer concentration (15 pM, 37°C) was not changed significantly (1.94 ± 0.52 to 2.05 ± 0.62% per 30 cm2 surface area/ml). The basal (noninsulin-stimulated) glucose transport (tracer glucose concentration 5 μM) increased from 25 ± 12 to 44 ± 14 pmol · 90 min−1 · 10 cm−2 surface area (P < .02). The maximally insulin-stimulated glucose transport rate increased from 35 ± 20 to 78 ± 26 pmol/90 min (P < .01). The percentage insulin response above basal levels increased from 31 ± 40 to 89 ± 58% (P < .01). The insulin sensitivity (half-maximally stimulating insulin concentrations) was also improved (P < .05). Glucose conversion rates to total lipids increased 34 ± 62 and 65 ± 80% in basal cells and maximally insulin-stimulated cells, respectively (.2 > P > .1 , .1 > P > .05). The percentage insulin response above basal level increased from 30 ± 24 to 67 ± 52% (P < .05). The percentage insulin response above basal level of glucose conversion to CO2 also increased (P < .05). The results after diet treatment were not different from those of normal controls. We conclude that dietary treatment of moderately obese NIDDM patients that leads to good glycemic control results in reversal of the abnormalities observed in adipocytes of untreated NIDDM subjects.

Defective Non-insulin-mediated and Insulin-mediated Glucose Transport and Metabolism in Adipocytes from Obese and Lean Patients with Untreated Type 2 Diabetes Mellitus

Insulin binding, glucose transport, and glucose metabolism were investigated in isolated adipocytes from 11 lean and 13 obese patients with non‐insulin‐dependent diabetes mellitus. Insulin binding at 15°C was reduced by 35% (p<0.01) in both lean and obese diabetic patients, whereas insulin binding (or uptake) at 37°C was similar in diabetic patients and healthy controls. In lean diabetic patients both non‐insulin‐mediated (basal) and maximally insulin‐stimulated glucose transport and metabolism were significantly reduced (all p<0.01). The percentage responses to insulin were also markedly reduced (p<0.05, p<0.02). In obese diabetic patients basal glucose transport was reduced (p<0.01) but basal glucose metabolism was not. Insulin‐stimulated glucose transport and metabolism were significantly reduced (p<0.01, p<0.05). The percentage responses were reduced compared to healthy controls (p<0.05, p<0.05) but higher than in lean diabetic patients (p<0.05).