Specific Insulin Receptor Binding Research Articles

Adipocyte insulin action in hypogonadotrophic hypogonadism.

In-vitro studies of adipose tissue have shown that patients with polycystic ovarian syndrome (PCOS) have marked insulin resistance, the abnormalities being more pronounced during amenorrhoea compared to following an ovulatory cycle. If the insulin resistance in PCOS is a reflection of anovulation then patients with hypogonadotrophic hypogonadism (HH) should also have a reduction in insulin sensitivity. This study was designed to investigate insulin sensitivity in patients with HH. Seven patients with HH were studied and compared with eight age and body mass index matched female controls. Adipocyte insulin receptor binding was measured and adipocyte insulin action was assessed by measuring initial rates of 3-O-methylglucose uptake and inhibition of lipolysis. The specific insulin receptor binding per 10 cm(2) cell surface was 0.95 +/- 0. 25% in HH and 1.85 +/- 0.14% in control patients (P < 0.01). Maximum rates of glucose uptake were also impaired in HH compared with controls (3-O-methylglucose transport 0.81 +/- 0.22 versus 1.83 +/- 0.2 pmol/10 cm(2)/5 s)(P < 0.01). Hence, patients with HH have impaired insulin sensitivity to a degree similar to that seen in PCOS, suggesting a direct effect of anovulation on insulin sensitivity.

Adipocyte insulin action during the normal menstrual cycle

The relationship between the menstrual cycle and insulin sensitivity is unclear. The aim of this study was to investigate insulin sensitivity during the normal menstrual cycle using the physiological insulin target organ adipose tissue. A total of 23 normal healthy volunteers were studied, nine of whom were in the follicular phase, and 14 of whom were age and body mass index-matched and in the luteal phase of the menstrual cycle. Adipocyte insulin receptor binding was measured and adipocyte insulin action was assessed by measuring initial rates of 3-O-methylglucose uptake and by inhibition of lipolysis. The maximum specific insulin receptor binding was significantly higher in subjects studied during the follicular phase of the menstrual cycle compared to subjects studied during the luteal phase (1.81 +/- 0.13 versus 1.36 +/- 0.15% per 10 cm2 cell surface, P < 0.05). Maximum rates of 3-O-methylglucose transport were 1.70 +/- 0.22 versus 1.75 +/- 0.22 pmol/10 cm2/5 s in the follicular and luteal phase respectively and were not significantly different between the two groups. The maximum percentage lipolysis inhibition observed was 42. 5 +/- 7.5% in the follicular phase and 39.9 +/- 7.4% in the luteal phase (not significant). This study demonstrated that there is a reduction in insulin receptor binding in the luteal phase of the normal ovulatory menstrual cycle. The post-receptor action of insulin is not affected between the two phases of the menstrual cycle.

Antiobesity effect of diazoxide in obese zucker rats

Hyperinsulinism and insulin resistance are characteristic findings in obese subjects. Obesity in both humans and experimental animals is associated with a reduced number of insulin receptors and a decreased insulin-mediated glucose disposal, whereas sensitivity to insulin's antilipolytic action is unaltered. To evaluate the antiobesity effect of diazoxide (DZ), an inhibitor of glucose-stimulated insulin release, 7-week-old Zucker obese and lean rats were studied. Obese and lean rats were grouped into DZ-treated (150 mg/kg/d) and control (C) groups. DZ-treated obese rats consumed similar amounts of calories per kilogram body weight (BW) compared with C obese animals, but gained less weight ( P < .01). Postabsorptive plasma free fatty acids (FFA), cholesterol, and triglycerides were significantly higher in obese versus lean animals ( P < .01). DZ treatment reduced plasma triglyceride levels in obese animals ( P < .001), but had no significant effect on FFA or cholesterol concentrations. Plasma glucose concentrations in the postabsorptive state and during glucose tolerance tests (GTTs) were significantly lower in DZ obese versus C obese rats ( P < .01) despite a decrease in plasma insulin concentrations in DZ-treated animals ( P < .01). In contrast, DZ lean rats developed glucose intolerance ( P < .05). Sensitivity and responsiveness to the antilipolytic effect of insulin in isolated adipocytes were significantly decreased in DZ obese as compared with C obese rats ( P < .01). Moreover, adipocyte specific insulin receptor binding was increased in both DZ lean and DZ obese animals ( P < .01). This was accompanied by increased basal and insulin-stimulated glucose transport in both genotypes ( P < .01). In conclusion, DZ increased insulin receptor binding and glucose transport while decreasing hyperinsulinemia and insulin sensitivity to the antilipolytic action of insulin. This combined effect resulted in improved glucose tolerance and a decrease in weight gain in obese rats, implying that pharmacologic modification of the disturbed insulin metabolism of obesity may be therapeutically beneficial.

Severe impairment of insulin action in adipocytes from amenorrheic subjects with polycystic ovary syndrome

Adipose tissue was used to characterize the metabolic abnormality of insulin resistance in polycystic ovary syndrome (PCOS). Nine patients with PCOS were studied during a period of amenorrhea and confirmed to be chronically anovulatory by vaginal ultrasound and plasma progesterone measurements. These were compared with six age- and body mass index (BMI)-matched controls (BMI, 27.2 ± 2.2 in PCOS and 24.7 ± 2.3 in control subjects). Insulin receptor binding was measured and insulin action was assessed by measuring initial rates of 3- O-methylglucose uptake and by inhibition of lipolysis. The maximum specific insulin receptor binding was 0.62% ± 0.12% and 1.78% ± 0.18% per 10-cm 2 cell surface (mean ± SEM) in PCOS and control subjects, respectively ( P < .001). Maximum rates of glucose transport were also impaired as compared with controls, with 3- O-methylglucose transport being 0.90 ± 0.15 versus 1.57 ± 0.28 pmol/10 cm 2/5s, respectively ( P < .05). The concentration of insulin required for half-maximal stimulation of glucose uptake was 165 ± 36 versus 32 ± 10 pmol in PCOS and control subjects, respectively ( P < .05). The maximum percentage lipolysis inhibition (mean ± SEM) was 9.5% ± 1.6% in PCOS and 28.3% ± 7.2% in control patients, respectively ( P < .01). These data demonstrate that there are both insulin binding and postreceptor defects in adipocytes from amenorrheic PCOS subjects. The degree of defect in adipocyte insulin action is greater than would have been anticipated from in vivo data.

Features of insulin receptor interaction in placenta from normal, overt and poorly controlled gestational diabetic patients.

Features of insulin binding to trophoblast plasma membranes were studied in six normal pregnant women (NP), six overt diabetes (ODP) and six poorly controlled glycemic gestational patients (PCDP) i.e. women who did not strictly follow the management of diabetes mellitus during pregnancy. A decreased maximum specific insulin receptor binding per 0.1 mg membrane protein in placenta from PCDP (12%) was found comparing with that from ODP or NP (17.5% and 36.2%, respectively, P < 0.01), The insulin binding in PCDP declined at a faster rate until it reached minimum when studied at a higher temperature (25-37 degrees C). The binding equilibrium was likewise attained faster at this temperature than that at lower temperature of 4 degrees C for all studied groups. The insulin receptor binding in all studied groups was pH dependent. The maximum binding in ODP and PCDP groups was attained at pH 7.8 while for NP maximum binding was at pH 7.4. The competitive binding assay was carried out with 14 concentrations of unlabelled insulin and the half maximal displacement of 125I-insulin was at 8 x 10(-9) M, 6 x 10(-9) M and 4 x 10(-9) M for NP, ODP and PCDP, respectively (P < 0.05) suggesting the differences in the effect of glycemic control on the insulin binding. Furthermore the binding yielded curvilinear Scatchard plots with the apparent affinity of the receptors being affected in the ODP and PCDP groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of in vivo estrogen treatment on adipose tissue metabolism and nuclear estrogen receptor binding in isolated rat adipocytes

We have previously demonstrated the existence of nuclear estrogen receptors in isolated adipocytes (Pedersen et al. (1991) Biochim. Biophys. Acta 1093, 80–86). In the present study we have investigated the regulatory properties of these nuclear estrogen receptors, in addition to the metabolic effects of estrogen on adipose tissue metabolism. Estrogen treatment (20 μg 17β-estradiol in NaCl for 7 days) decreased lipoprotein lipase activity (LPL) in the adipose tissue by 62% ( p < 0.05), decreased adipocyte size by 27% ( p < 0.01) and diminished the normal postovariectomy weight gain. Furthermore, estrogen treatment increased the nuclear estrogen receptor binding in adipocytes; in addition, there was a tendency for increased cytosolic estrogen receptor content as well. Time course studies revealed that already 6 h after a single estrogen injection the B max increased from 3.82 ± 0.3 fmol/10 6 cells to 9.8 ± 3.6 fmol/10 6 cells ( p < 0.1) and 24 h after a single injection the B max was maximally increased to 12.7 ± 5.5 fmol/10 6 cells ( p < 0.05). The K d was similar at all time points (about 3–5 nM). Furthermore, the specific insulin receptor binding was increased in adipocytes from estrogen treated rats. The specific insulin binding was maximally increased by 149 ± 6% ( p < 0.001) after 4 days of daily estrogen injections. The increased binding seemed to be due to an increased number of insulin receptors on adipocytes from estrogen treated rats with no alteration of the ED 50 value. In conclusion it was found that estrogen treatment has a positive feedback effect on its own nuclear receptor. In addition, estrogen treatment was found to have several metabolic effects in the adipose tissue. Estrogen treatment increased the insulin receptor number, decreased LPL activity, increased the lipolytic response in adipocytes and finally decreased body weight and adipocyte cell size.

Effects of insulin on adrenoceptor binding and the rate of catecholamine-induced lipolysis in isolated human fat cells.

The mechanisms by which insulin inhibits catecholamine-induced lipolysis in fat cells are unknown. In this study the possible role of an interaction between insulin and the adrenoceptors on human fat cells was investigated. Insulin inhibited, in a dose-dependent fashion, the specific binding of hydrophobic as well as hydrophilic nonselective beta-receptor radioligands but had no effect on the binding of alpha 2-selective radioligands. The results of saturation experiments and competition-inhibition experiments under both equilibrium conditions and nonequilibrium conditions revealed that insulin reduced the total number of beta-adrenergic binding sites (maximum effect 25%) without changing the beta-adrenoceptor affinity. This insulin effect was rapid and reversible; one-third of the effect occurred within 1 min of incubation and it was completely reversed within 30 min after withdrawal of insulin. It could be mimicked by a polyclonal rabbit insulin receptor antibody but not by insulin mimickers acting distal to the initial interaction between the hormone and its specific insulin-receptor binding site. The beta-adrenoceptor binding to a plasma membrane-enriched fraction decreased at the same time as it increased to a microsomal enriched fraction after insulin treatment, indicating a redistribution of beta-adrenoceptors in the cell. In lipolysis experiments performed under conditions like those in the binding experiments, insulin inhibited the rate of lipolysis with a lag period of 3 min. Furthermore, the hormone caused a dose-dependent maximum 10-fold shift to the right of the dose-response curve for isoprenaline-induced lipolysis without changing the amplitude of the curve. This effect of insulin was specific for the beta-adrenergic receptors system, since insulin markedly decreased the amplitude of the dose-response curve for parathyroid hormone-induced lipolysis. In addition, the effect of insulin on isoprenaline-induced lipolysis could be mimicked by long-lasting fractional inactivation of the beta-adrenoceptors. The dose-response relationships for the inhibitory effects of insulin on beta-adrenoceptor binding and the lipolytic sensitivity to isoprenaline were almost identical. Half-maximum and maximum effects occurred at about 5 and 100 microunits/ml of insulin, respectively. In conclusion, the exposure of human fat cells to physiological insulin doses is followed by a rapid and dose-dependent translocation of beta-adrenoceptors from the exterior to the interior of the cell and a subsequent dose-dependent decrease in the lipolytic sensitivity to beta-adrenergic agonists, without a change in maximum lipolysis.(ABSTRACT TRUNCATED AT 400 WORDS)

Monocyte-T lymphocyte interaction for regulation of insulin receptors of the activated T lymphocyte.

During activation the lymphocyte attains functional insulin receptors with precise regulation, a consequence of insulin concentration manipulations. These studies test the hypothesis that insulin receptor (+) monocytes monitor insulin concentrations, so instructing the T lymphocytes. Monocyte-enriched populations were incubated with insulin (0-10(-6) M) followed by co-culture with T lymphocytes and an activating stimulus. A dose-related fall in T lymphocyte insulin receptor binding was observed that was specific for the monocyte as the signalling cell and for insulin as the signal received. Monocytes from normal volunteers during a euglycemic, hyperinsulinemic clamp were cultured with T lymphocytes and an activating stimulus. A decline in specific insulin receptor binding on T lymphocytes was observed, which Scatchard analysis demonstrated to be a consequence of reduction in receptor numbers. These studies demonstrate that the receptor (+) monocyte perceives the concentration of insulin and passes this information to T lymphocytes regulating the number of activation-induced insulin receptors. The interplay between the monocyte and T lymphocyte parallels the interaction of these cell types for recognition of antigen.

Insulin receptor binding and metabolic effects of insulin in human subcutaneous adipose tissue in untreated non-insulin dependent diabetes mellitus.

Insulin action at the target tissue level in non-insulin dependent diabetes mellitus was investigated using human adipose tissue. Specific adipocyte receptor binding of insulin and the effects of the hormone on glucose oxidation and lipolysis were determined in subcutaneous adipose tissue. The study included 25 patients with untreated non-insulin dependent diabetes mellitus and 38 healthy control subjects matched for age, sex and body weight. Insulin stimulated adipose tissue glucose oxidation in a dose-dependent way in the control subjects. On the other hand, a marked inhibition of this insulin effect was observed in the diabetics. A weak stimulation was observed only at high unphysiological hormone concentrations [greater than or equal to 0.7 nmol/l] and the maximal insulin response was 6 times lower than that in the control subjects. However, neither specific insulin receptor binding nor the antilipolytic effect of insulin were inhibited in diabetes. Similar results with insulin binding and the metabolic effects of insulin were obtained in non-obese normoinsulinemic diabetics as compared to moderately obese hyperinsulinemic diabetics. It is concluded that adipose tissue insulin resistance in non-insulin dependent diabetes mellitus only involves glucose metabolism and not antilipolysis. Furthermore, it may solely be due to postreceptor defects in insulin action and seems not to be influenced by obesity or oversecretion of insulin.

Induction of insulin receptor expression of human leukemic cells by 1α,25 dihydroxyvitamin D 3

The HL-60 and U-937 leukemic cell line and explants of fresh human leukemia cells were differentiated in vitro by incubation with 1α,25 dihydroxyvitamin D 3 (Vit D). Morphologic change to monocytes was demonstrated in the promyelocytic (HL-60) line but not in the U-937 line and four of five leukemic cells. One patient with chronic granulocytic leukemia in the myeloid blast phase had incomplete morphologic change with Vit D treatment. In all cells studied, an increase in specific insulin receptor binding was independent of morphologic maturation. An increase in insulin receptor expression by Vit D was an early monocytic membrane marker dissociated from morphologic and functional alterations.

Insulin action does not change during the menstrual cycle in normal women.

Normal women have alterations in carbohydrate metabolism during pregnancy and when taking oral contraceptives, and clinical observations suggest that diabetic women need more insulin during menstruation. We, therefore, studied insulin action in normal women during the menstrual cycle in the follicular, luteal, and menstrual phases. Glucose tolerance was similar at all three times. Specific insulin receptor binding to monocytes did not change during the menstrual cycle. Euglycemic insulin clamp studies at four different insulin infusion rates (15, 40, 120, and 240 mU/M2 X min) showed no differences in insulin sensitivity or responsiveness throughout the menstrual cycle, and hepatic glucose output did not change. These studies suggest that if insulin action is impaired during menstruation in diabetic women it is because of factors that are not detected in normal women.

Influence of fasting and refeeding on the antilipolytic effect of insulin in human fat cells obtained from obese subjects.

The antilipolytic effect of insulin was investigated in obese subjects before and after 7 days of total fasting, and 1 h after oral refeeding with 100 g glucose. Isolated fat cells were prepared from subcutaneous gluteal adipose tissue and incubated in vitro. Specific insulin receptor binding and insulin inhibition of basal and isoprenaline-stimulated lipolysis were determined. During the fasting period, a 15% increase (P less than 0.05) in high-affinity insulin binding and a concomitant 3-4-fold increase in insulin sensitivity were noted, and there was a marked enhancement of the maximum insulin-induced inhibition of basal lipolysis, from 4 to 10 mumol of glycerol/10(7) cells/2 h. The maximum insulin-induced inhibition of isoprenaline-induced lipolysis was similar before and after fasting, about 10 mumol/10(7) cells/2 h. Glucose refeeding induced a 30% decrease (P less than 0.02) in high-affinity insulin binding and a 20-60-fold decrease (P less than 0.01) in the sensitivity of the antilipolytic effect of insulin under basal conditions and in the presence of isoprenaline. The maximum antilipolytic effect of insulin, however, was not altered by glucose refeeding. Thus, in the basal state, maximum antilipolytic effect was larger after refeeding as compared with that before fasting. The high-affinity insulin binding and insulin sensitivity were significantly lower after refeeding than before fasting. Before the fasting period, neither the insulin binding nor the antilipolytic effect of the hormone was altered by oral glucose. It is concluded that fasting and glucose refeeding are associated with marked alterations in the antilipolytic effect of insulin on human fat cells of obese subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of fasting on insulin receptor binding and insulin action in different human subcutaneous fat depots.

The possible existence of regional variations in fasting-mediated changes on insulin action an human adipose tissue was investigated in vitro. Subcutaneous adipose tissue was obtained from the femoral, abdominal, and gluteal areas of obese but otherwise normal subjects (16 women and 7 men) before and after 7 days of total fasting. Specific insulin receptor binding to isolated fat cells was similar in femoral and abdominal adipose tissues before and after fasting. However, the latter condition was associated with a significant increase in insulin receptor binding at low hormone concentrations (less than 2 nmol/liter) in gluteal adipocytes. Insulin stimulated glucose oxidation in a dose-dependent way in all 3 adipose regions before fasting. In the femoral and gluteal sites after fasting, the maximum insulin effect was significantly decreased, but a dose-dependent insulin effect was still present, and there was no change in insulin sensitivity. However, abdominal adipose tissue after fasting was completely unresponsive to insulin stimulation when the hormone was added in increasing concentrations up to 80 nmol/liter. The results in fasting women were similar to those in the whole study group. In conclusion, there appear to be marked regional variations in fasting-mediated changes in insulin action on glucose metabolism in human adipose tissue. Alterations at the postreceptor level which lead to insulin resistance appear to be of greater importance than the counteracting receptor changes.

Influence of obesity on the antilipolytic effect of insulin in isolated human fat cells obtained before and after glucose ingestion.

The antilipolytic effect of insulin was studied in 9 obese and 10 age- and sex-matched subjects of normal weight. Isolated fat cells were taken before and 1 h after an 100 g oral glucose load. Insulin inhibition of basal and isoprenaline-induced rates of lipolysis were determined by using a sensitive bioluminescent glycerol assay. When compared with the controls, the obese group showed a lower glucose tolerance, a higher insulin secretion, and a lower specific insulin receptor binding per adipocyte surface area, which would suggest an insulin-resistant state. Before oral glucose, however, the sensitivity of the antilipolytic effect of insulin was enhanced 10-fold in obesity (P less than 0.01), but the maximum antilipolytic effect was not altered. Glucose ingestion induced a 10-25-fold increase in insulin sensitivity (P less than 0.01) and a 10% but not significant increase in specific adipocyte insulin receptor binding in the nonobese group. In the obese group, however, neither the insulin binding nor the antilipolytic effect of the hormone was increased by oral glucose. After oral glucose, insulin sensitivity was similar in the two groups. The concentration of the hormone which produced a half maximum effect was about 1 microU/ml. Similar results were obtained with insulin inhibition of basal and isoprenaline-stimulated glycerol release. It is concluded that, after an overnight fast, the sensitivity of the antilipolytic effect of insulin is markedly enhanced in adipocytes of "insulin-glucose resistant" obese subjects, presumably because of alterations at postreceptor levels of insulin action. In obesity, the antilipolytic effect of insulin seems normal after glucose ingestion. Furthermore, in adipocytes of subjects of normal weight, oral glucose rapidly stimulates the sensitivity of the antilipolytic effect of insulin, apparently because of changes at postreceptor sites. This short-term regulation of insulin action following the ingestion of glucose does not seem to be present in obesity.

Marked increase in insulin sensitivity of human fat cells 1 hour after glucose ingestion.

The effect of glucose ingestion on insulin action was investigated in isolated human fat cells. Subcutaneous adipose tissue was obtained from eight normal adult volunteers before and 1 h after oral intake of 100 g of glucose. Lipolysis (glycerol release) and specific insulin receptor binding were determined. Insulin binding increased significantly by 20-30% after glucose ingestion. This was due to an increase in insulin binding affinity, without any change in the receptor number. The concentration of insulin producing half-maximum inhibition (ED(50)) of basal lipolysis was 50 muU/ml before and 0.25 muU/ml after glucose ingestion (P < 0.01), which represented a 200-fold difference. As regards isoprenaline-induced lipolysis, the ED(50) for insulin inhibition was 30 muU/ml before and 2.5 muU/ml after oral glucose (P < 0.01), which was a 12-fold difference. The maximum insulin-induced inhibition of basal and isoprenaline-induced lipolysis were not altered after oral glucose. It is concluded that glucose ingestion is accompanied by a marked increase in insulin sensitivity of human fat cells and this may be an important modulating factor in the overall scheme of insulin action.