Insulin-like effect of low-dose leptin on glucose transport in Langendorff rat hearts.

BACKGROUND: In a murine myotube cell line (C 2 C 12 myotubes), leptin at low physiological concentrations (1 ng/ml) has been shown to stimulate glucose transport and glycogen synthesis. The aim of the present study was to test whether an analogous leptin effect on glucose transport is detectable in the heart. METHODS AND RESULTS: We used the isolated Langendorff rat heart preparation with hemodynamic function control. Using polymerase chain reaction (RT-PCR), a 346- and 375-base fragment indicative for the short and long leptin receptor isoform was detected in the rat heart. Glucose transport rates were calculated using equimolar double tracer perfusion with the non-metabolizable glucose analog 3-O-methylglucose (3-O-MG) and the non-transportable tracer mannitol and two-compartimental modeling. 3-O-MG uptake at a perfusate glucose concentration of 11 mM was measured over 15 minutes in control hearts, hearts perfused with insulin (10 mU/ml), leptin (1 ng/ml) or insulin (10 mU/ml) plus leptin (1 ng/ml; n = 8 in each group). The basal 3-O-MG transport rate of 0,7351 +/- 0,051 micro mol/min/g wet weight was increased 4.18 fold with insulin, 2.69 fold with leptin, and 4.2 fold with leptin plus insulin. Simultaneous monitoring of hemodynamic function revealed a minor and transient effect of leptin on left ventricular pressure, which was strongly augmented in coperfusion with insulin. CONCLUSIONS: The data suggest that leptin at low physiological concentrations is able to exert a partial insulin like effect on glucose uptake. We speculate that the effect might be mediated by both leptin receptor isoforms. This leptin effect is additive to the effect of insulin and might therefore contribute to the insulin independent basal glucose supply of the heart. It can not be completely excluded that the observed leptin effect on glucose transport is secondary to altered myocardial function.

Induction of adiponectin gene expression in human myotubes by an adiponectin-containing HEK293 cell culture supernatant.

AIMS/HYPOTHESIS: Adiponectin, an adipocytokine known to be down-regulated in obesity-linked disorders, is considered to be a potential key mediator of insulin sensitivity. In this study, we asked whether adiponectin is able to regulate ten selected genes possibly associated with insulin sensitivity in human skeletal muscle cells. METHODS: To this end, we treated in vitro differentiated human myotubes with the culture supernatant of HEK293 cells stably transfected with human recombinant adiponectin and assessed gene expression by RT-PCR. Intracellular adiponectin protein was quantified by radioimmunoassay and visualized by Western blotting. RESULTS: In contrast to the control supernatant, the adiponectin-containing supernatant consistently induced expression of adiponectin mRNA in human myotubes from eight different donors (mean increase: 90-fold over control; n=8, p<0.001). This increase in mRNA was paralleled by a rise in intracellular adiponectin protein (mean increase: 8.3-fold over control; n=4, p<0.05). Expression of the other nine candidate genes was not altered. In human skin fibroblasts and HepG2 cells, the adiponectin-enriched supernatant did not induce relevant amounts of adiponectin mRNA. CONCLUSIONS/INTERPRETATION: In conclusion, we show here that adiponectin gene expression is specifically inducible in skeletal muscle cells.

Evidence for greater oxidative substrate flexibility in male carriers of the Pro 12 Ala polymorphism in PPARgamma2.

The Pro12Ala polymorphism of the peroxisome proliferator-activated receptor gamma2 (PPARgamma2) gene is associated with reduced type 2 diabetes risk and increased insulin sensitivity. It is possible that the oxidative shift from lipid to glucose as a fuel is more efficient in Ala allele carriers. To test this hypothesis, we examined carbohydrate and lipid oxidation by indirect calorimetry in lean, glucose tolerant subjects with (X/Ala, n = 25) and without the Pro12Ala polymorphism (Pro/Pro, n = 73) basally and after insulin stimulation during a 2-hour eugylcaemic hyperinsulinaemic clamp. Insulin sensitivity was non-significantly greater in X/Ala (0.13 +/- 0.01 micromol/kg/min/pM) than in Pro/Pro (0.12 +/- 0.01 micromol/kg/min/pM, p = 0.27). Basally, there were no lipid nor carbohydrate oxidation differences between the groups. Interestingly, the decrease in lipid oxidation during insulin stimulation was significantly greater in male X/Ala (- 0.51 +/- 0.06 mg/kg/min) than in male Pro/Pro (- 0.35 +/- 0.04 mg/kg/min, p = 0.03). No difference was observed in females. Analogously, the change in carbohydrate oxidation in male X/Ala (1.34 +/- 0.2 mg/kg/min) was significantly greater than in male Pro/Pro (1.03 +/- 0.12 mg/kg/min, p = 0.05). The respiratory quotient increased more, but not significantly more, in male X/Ala (0.11 +/- 0.01) than in male Pro/Pro subjects (0.08 +/- 0.01, p = 0.08) but similarly in females. These results indicate that the mechanism by which the Ala allele improves insulin sensitivity might involve enhanced suppression of lipid oxidation permitting more efficient (predominantly non-oxidative) glucose disposal. It is unclear why this could be demonstrated only in males, although gender differences in substrate oxidation are well documented.

The prevalent Gly1057Asp polymorphism in the insulin receptor substrate-2 gene is not associated with impaired insulin secretion.

Disruption of the insulin receptor substrate-2 was shown to cause type 2 diabetes in mice. This could be largely attributed to abnormal beta-cell development. In humans, a prevalent polymorphism in insulin receptor substrate-2 (Gly1057Asp) was not found be associated with type 2 diabetes in linkage and association studies. We tested the hypothesis that an extreme challenge of the beta cell might reveal subtle abnormalities in carriers of this polymorphism undetected by conventional insulin secretion tests. Therefore, in addition to assessing beta-cell function by oral glucose tolerance testing (n = 318, normal glucose tolerance), we measured the secretory response to maximal stimulation by hyperglycemia (10 mM), glucagon-like peptide-1, and arginine administered in an additive fashion (n = 77, nondiabetic). The allelic frequency of the Asp allele was approximately 37%. Neither the beta-cell function indices from the oral glucose tolerance test nor the secretory response during the hyperglycemic clamp differed measurably between carriers and controls. Moreover, maximal plasma C-peptide concentrations in response to the combined glucose, glucagon-like peptide-1, and arginine stimulus was not different between Gly/Gly (10,745 +/- 1,186 pmol/liter) and X/Asp (10,800 +/- 490 pmol/liter, P = 0.99). In conclusion, our findings strongly suggest that the Gly1057Asp polymorphism in insulin receptor substrate-2 is not associated with beta-cell dysfunction. The normal maximal insulin secretory response makes it unlikely that this common polymorphism results in abnormal beta-cell development.

Impaired non-esterified fatty acid suppression is associated with endothelial dysfunction in insulin resistant subjects.

In a recent study, we found a significant association between insulin resistance (IR) and disturbed flow-associated (endothelial-dependent) vasodilation in first-degree relatives of subjects with type 2 diabetes. However, the mechanisms linking insulin resistance and endothelial dysfunction (ED) have not been fully elucidated. Experimental data have pointed out that non-esterified fatty acids (NEFA) have a modulating effect on NO-synthase activity, and therefore on endothelial function. The aim of our study was to evaluate whether insulin resistance associated impaired NEFA suppression is present in subjects with ED. We examined 53 first-degree relatives (FDR) of patients with type 2 diabetes (32f, 21 m, mean age 35 years). Endothelial function was measured as flow-associated vasodilation (FAD%) of the brachial artery. Insulin sensitivity was evaluated with a standard hyperinsulinemic glucose clamp (insulin infusion rate of 1 mU/kg/min). While under fasting conditions, NEFA did not differ between groups with high or low FAD (0.415+/-0.033 vs. 0.394 +/- 0.040 mmol/l; p = n. s.), reduced FAD% was significantly associated with higher non-esterified fatty acids concentrations during steady state of the glucose clamp (0.072+/-0.022 vs. 0.039+/-0.016mmol/l; p=0.04). This association was independent of insulin levels under fasting conditions and during the glucose clamp. In conclusion, our results reveal a significant association between endothelial dysfunction and impaired non-esterified fatty acid suppression in insulin resistant subjects. As insulin resistance of lipolysis is a feature of the insulin resistance syndrome, these results suggest that elevated NEFA concentrations could play a role linking endothelial dysfunction and insulin resistance in vivo.

The Gly972Arg polymorphism in the insulin receptor substrate-1 gene contributes to the variation in insulin secretion in normal glucose-tolerant humans.

The Gly972Arg polymorphism in the insulin receptor substrate (IRS)-1 was found in some studies to have a higher prevalence in type 2 diabetic subjects than in control subjects. Previously, transfection of IRS-1 with this polymorphism into insulin-secreting cells resulted in a marked reduction of glucose-stimulated insulin secretion compared with the wild-type transfected cells. In the present study, we compared insulin secretion in well-matched normal glucose-tolerant subjects with and without this polymorphism. Several validated indexes of beta-cell function from the oral glucose tolerance test were significantly lower in X/Arg (n = 31) compared with Gly/Gly (n = 181) (P between 0.002 and 0.05), whereas insulin sensitivity (measured with a euglycemic clamp) was not different. During a modified hyperglycemic clamp, insulin secretion rates were significantly lower in Gly/Arg (n = 8) compared with Gly/Gly (n = 36) during the first phase (1,711+/-142 vs. 3,014+/-328 pmol/min, P = 0.05) and after maximal stimulation with arginine (5,340+/-639 vs. 9,075+/-722 pmol/min, P = 0.03). In summary, our results suggest that the Gly972Arg polymorphism in IRS-1 is associated with decreased insulin secretion in response to glucose but not with insulin sensitivity. It is possible that this polymorphism causes insulin resistance at the level of the beta-cell and contributes to the polygenic etiology of type 2 diabetes.

The PPARgamma2 polymorphism pro12Ala is associated with better insulin sensitivity in the offspring of type 2 diabetic patients.

Recently, a highly prevalent polymorphism of the PPARgamma2-receptor (Pro12Ala) was described and found to be associated with reduced transcriptional activity. Both human and animal studies suggested that this polymorphism may be associated with increased insulin sensitivity. However, an effect independent of other factors known to influence insulin sensitivity has yet to be demonstrated. Therefore, we compared insulin sensitivity using the hyperinsulinemic-euglycemic clamp technique in 37 subjects heterozygous for the PPARgamma2-Pro12Ala mutation and 37 control subjects negative for the PPARgamma2-Pro12Ala. The control group was selected from 190 subjects by pair-matching for sex, BMI, fat distribution and body composition. In the group heterozygous for the polymorphism steady-state plasma insulin during the clamp was significantly lower (63.3 microU/ml +/- 2.8) than in the control group (74.9 microU/ml +/- 4.0, p = 0.02). While MCR of glucose was similar in the PPARgamma2-Pro12Ala group (8.1 ml/kg x min x 100 +/- 0.5) and the control group (7.6 ml/kg x min x 100 +/- 3.0, p = 0.7), the insulin sensitivity index was significantly higher in the PPARgamma2-Pro12Ala group (12.5 mg/kg x min x microU/ml +/- 0.9 vs. 9.7 mg/kg x min x microU/ml +/- 0.8, p = 0.039). In addition, an arbitrary lipolysis index (decrease in FFA divided by increase in insulin) was also found to be marginally higher in the PPARgamma2-Pro12Ala group (8.0 +/- 0.9) compared to the control group (6.1 +/- 0.7, p = 0.097). In conclusion, these data suggest that the PPARgamma2-Pro12Ala mutation is associated with better insulin sensitivity of glucose disposal and possibly, also of antilipolysis.

The silent PPARgamma exon 6 CAC(His) --> CAT(His) polymorphism does not affect the plasma leptin levels in a collective of first degree relatives of type 2 diabetes patients from South West Germany.

The peroxisome proliferator activated receptors-gamma (PPARgamma) belong to the superfamily of nuclear transcription factors acting as master genes regulating events in adipocyte differentiation. Thus, PPARgamma is a candidate gene for affecting insulin sensitivity and the pathogenesis of insulin resistance. PPARs trigger endocrine response of two important adipose tissue-derived signalling factors, leptin and tumor necrosis factor-alpha. Leptin is the afferent signal in a negative feedback loop regulating adipose tissue mass and energy balance. It generates insulin-like signals for glucose transport and glycogen synthesis via leptin receptors and the PI3-kinase and could, therefore, play a role as a mediator of obesity-related insulin resistance. Recently, a silent substitution in the coding sequence of the PPARgamma2 gene, leading to the substitution of a C by a T in exon 6 (nt 161), was described. In a recent study, it was proposed that mutations in PPARgamma could play a role in individuals who are at increased risk for developing obesity and type 2 diabetes mellitus by influencing leptin levels. We therefore examined the prevalence of the CAC(His) --> CAT(His) mutation in non-diabetic first degree relatives of subjects with type 2 diabetes to determine a possible association of this mutation to leptin levels and insulin sensitivity. 138 probands were characterised by oral glucose tolerance tests, euglycemic-hyperinsulinemic glucose-clamp and by measuring leptin levels. We found 93 (67.4%) probands without the CAC(His) --> CAT(His) substitution and 45 heterozygotes (36.6%). When the whole group was analysed for an association of the mutation with plasma leptin concentration and insulin sensitivity, no statistical significance could be demonstrated. Independently of the mutation, leptin levels were significantly (p<0.001) higher in female subjects.

Endothelial dysfunction is detectable in young normotensive first-degree relatives of subjects with type 2 diabetes in association with insulin resistance.

BACKGROUND: Endothelial dysfunction (ED) is regarded as an early step in the development of atherosclerosis. Among the pathogenetic factors leading to atherosclerosis, the role of insulin resistance and hyperinsulinemia as independent risk factors is still under debate. In this study, we examined the association between ED and insulin resistance in normotensive and normoglycemic first-degree relatives (FDRs) of patients with type 2 diabetes mellitus (DM). METHODS AND RESULTS: Endothelium-dependent and -independent vasodilation of the brachial artery was measured with high-resolution ultrasound (13 MHz) in 53 normotensive FDRs (21 men, 32 women; mean age, 35 years) with normal oral glucose tolerance, 10 age- and sex-matched normal control subjects, and 25 DM patients (mean age, 57 years). According to the tertiles of the clamp-derived glucose metabolic clearance rate (MCR), the FDRs were further classified as insulin resistant with an MCR or =7.8 mL. kg(-1). min(-1), and borderline with an MCR of 5.9 to 7.7 mL. kg(-1). min(-1). Flow-associated dilation was 4.1+/-0.9% in insulin-resistant FDRs, 6.7+/-1.1% in borderline FDRs, 9.0+/-1.2% in insulin-sensitive FDRs (P=0.002), 7.7+/-2.9% in control subjects (P=NS versus FDRs), and 3.8+/-1.0% in DM patients (P=0.03). In multiple regression analysis, low MCR was significantly correlated with ED independent of age, sex, smoking, body mass index, percent body fat, serum insulin, and lipids. CONCLUSIONS: There is a significant association between ED and insulin resistance in young FDRs of DM subjects independent of the classic cardiovascular risk factors.

The tumour necrosis factor alpha -238 G --> A and -308 G --> A promoter polymorphisms are not associated with insulin sensitivity and insulin secretion in young healthy relatives of Type II diabetic patients.

AIMS/HYPOTHESIS: Tumour necrosis factor-alpha (TNF-alpha) is believed to influence skeletal muscle insulin resistance. Two G --> A transitions in the promoter region of TNF-alpha at position -238 and -308 have been identified that could play a part in transcriptional regulation of the gene. Insulin resistance is an independent familial trait that predicts the development of Type II (non-insulin-dependent) diabetes mellitus. We investigated the influence on insulin sensitivity and insulin secretion of both polymorphisms in a cohort of young healthy relatives of patients with Type II diabetes. METHODS: We examined 109 first-degree relatives of Caucasian patients with a history of Type II diabetes, who underwent extensive metabolical and anthropometrical phenotyping, and determined the TNF-alpha -238 and -308 G --> A promoter polymorphisms. RESULTS: For the -238 polymorphism, 3 probands (76.1%) were homozygous for the G-allele, 25 probands (22.9%) were heterozygous and 1 proband (0.9%) was homozygous for the A-allele. For the -308 polymorphism, 83 probands (76.1%) were homozygous for the G-allele, 24 probands (22.0%) were heterozygous and 2 probands (1.18%) were homozygous for the A-allele. Probands with and without the polymorphism did not differ in insulin sensitivity (p = 0.78), insulin-concentrations and C-peptide concentrations in oral glucose tolerance tests (p > 0.05). CONCLUSIONS/INTERPRETATION: We could not detect an association between insulin sensitivity or insulin secretion and TNF-alpha promoter polymorphisms in our cohort. The polymorphisms occur at the same frequencies in probands with either low or high insulin sensitivity.

Amino acid polymorphism Gly 972 Arg in IRS-1 is not associated to lower clamp-derived insulin sensitivity in young healthy first degree relatives of patients with type 2 diabetes.

The Gly 972 Arg variant in the insulin receptor substrate-1 (IRS-1) gene may interact with the pathogenesis of common insulin-resistance disorders raising the hypothesis that the mutation may predispose to type 2 diabetes. We examined the codon 972 variant in 144 non-diabetic first degree relatives of patients with type 2 diabetes (FDR), who underwent extensive phenotyping: Glucose tolerance was determined by an oral glucose load, insulin sensitivity by euglycaemic-hyperinsulinaemic glucose clamp (glucose metabolic clearance rate, MCR) and body composition by bioelectrical impedance. 20 (14%) of the FDR showed the Gly 972 Arg variant in heterozygous form, 2 (1.3%) probands were homozygous. Carriers of the polymorphism did not differ in MCR independent of body weight and total body fat. The polymorphism does not seem to determine clamp-derived insulin sensitivity. Despite identical fasting plasma glucose, carriers of the polymorphism showed a slightly lower fasting serum insulin and lower insulin response to an oral glucose load but higher glucose concentrations. In an obese subgroup (BMI > 25) the polymorphism did not show a higher frequency and was not associated with lower insulin sensitivity. In the investigated group of young, healthy relatives of type 2 diabetes patients, the frequency of the mutation corresponded to that of a diabetic population. In summary our data show that the polymorphism is not suitable to predict insulin resistance.

Oral administration of RAC-alpha-lipoic acid modulates insulin sensitivity in patients with type-2 diabetes mellitus: a placebo-controlled pilot trial.

Alpha-lipoic acid (ALA), a naturally occuring compound and a radical scavenger was shown to enhance glucose transport and utilization in different experimental and animal models. Clinical studies described an increase of insulin sensitivity after acute and short-term (10 d) parenteral administration of ALA. The effects of a 4-week oral treatment with alpha-lipoic acid were evaluated in a placebo-controlled, multicenter pilot study to determine see whether oral treatment also improves insulin sensitivity. Seventy-four patients with type-2 diabetes were randomized to either placebo (n = 19); or active treatment in various doses of 600 mg once daily (n = 19), twice daily (1200 mg; n = 18), or thrice daily (1800 mg; n = 18) alpha-lipoic acid. An isoglycemic glucose-clamp was done on days 0 (pre) and 29 (post). In this explorative study, analysis was done according to the number of subjects showing an improvement of insulin sensitivity after treatment. Furthermore, the effects of active vs. placebo treatment on insulin sensitivity was compared. All four groups were comparable and had a similar degree of hyperglycemia and insulin sensitivity at baseline. When compared to placebo, significantly more subjects had an increase in insulin-stimulated glucose disposal (MCR) after ALA treatment in each group. As there was no dose effect seen in the three different alpha-lipoic acid groups, all subjects receiving ALA were combined in the "active" group and then compared to placebo. This revealed significantly different changes in MCR after treatment (+27% vs. placebo; p < .01). This placebo-controlled explorative study confirms previous observations of an increase of insulin sensitivity in type-2 diabetes after acute and chronic intravenous administration of ALA. The results suggest that oral administration of alpha-lipoic acid can improve insulin sensitivity in patients with type-2 diabetes. The encouraging findings of this pilot trial need to be substantiated by further investigations.

The PPARgamma2 amino acid polymorphism Pro 12 Ala is prevalent in offspring of Type II diabetic patients and is associated to increased insulin sensitivity in a subgroup of obese subjects.

AIMS/HYPOTHESIS: Recently a mutation in the coding sequence of the adipocyte specific isoform peroxisome proliferator-activated receptor gamma2 (PPARgamma2) was described, leading to the substitution of Proline to Alanine at codon 12. Mutations in PPARgamma2 could have a role in people who are at increased risk for the development of obesity and Type II (non-insulin-dependent) diabetes mellitus. METHODS: Non-diabetic first-degree relatives (n = 108) of subjects with Type II diabetes were characterized by oral glucose tolerance tests and euglycaemic hyperinsulinaemic glucose clamp to determine insulin sensitivity. RESULTS: We found 75 (69%) probands without the PPARgamma ProAla12 substitution, 28 heterozygotes (26%) and 5 (4%) homozygotes. When the whole group was analysed for an association between the mutation and insulin sensitivity, no statistical significance could be shown. Only in the group with severe obesity more than 30 kg/m2, an association (p = 0.016) of the polymorphism with an increase in insulin sensitivity was found. CONCLUSION/INTERPRETATION: These observations suggest that the mutation in the PPARgamma2 molecule may have a role in subgroups prone to the development of obesity and Type II diabetes.

Association of increased intramyocellular lipid content with insulin resistance in lean nondiabetic offspring of type 2 diabetic subjects.

Insulin resistance plays an important role in the pathogenesis of type 2 diabetes; however, the multiple mechanisms causing insulin resistance are not yet fully understood. The aim of this study was to explore the possible contribution of intramyocellular lipid content in the pathogenesis of skeletal muscle insulin resistance. We compared insulin-resistant and insulin-sensitive subjects. To meet stringent matching criteria for other known confounders of insulin resistance, these individuals were selected from an extensively metabolically characterized group of 280 first-degree relatives of type 2 diabetic subjects. Some 13 lean insulin-resistant and 13 lean insulin-sensitive subjects were matched for sex, age, BMI, percent body fat, physical fitness, and waist-to-hip ratio. Insulin sensitivity was determined by the hyperinsulinemic-euglycemic clamp method (for insulin-resistant subjects, glucose metabolic clearance rate [MCR] was 5.77+/-0.28 ml x kg(-1) x min(-1) [mean +/- SE]; for insulin-sensitive subjects, MCR was 10.15+/-0.7 ml x kg(-1) x min(-1); P<0.002). Proton magnetic resonance spectroscopy (MRS) was used to measure intramyocellular lipid content (IMCL) in both groups. MRS studies demonstrated that in soleus muscle, IMCL was increased by 84% (11.8+/-1.6 vs. 6.4+/-0.59 arbitrary units; P = 0.008 ), and in tibialis anterior muscle, IMCL was increased by 57% (3.26+/-0.36 vs. 2.08+/-0.3 arbitrary units; P = 0.017) in the insulin-resistant offspring, whereas the extramyocellular lipid content and total muscle lipid content were not statistically different between the two groups. These data demonstrate that in these well-matched groups of lean subjects, IMCL is increased in insulin-resistant offspring of type 2 diabetic subjects when compared with an insulin-sensitive group matched for age, BMI, body fat distribution, percent body fat, and degree of physical fitness. These results indicate that increased IMCL represents an early abnormality in the pathogenesis of insulin resistance and suggest that increased IMCL may contribute to the defective glucose uptake in skeletal muscle in insulin-resistant subjects.

Insulin action and secretion in healthy, glucose tolerant first degree relatives of patients with type 2 diabetes mellitus. Influence of body weight.

It is a matter of controversy, whether insulin action or secretion - or both - are disturbed in first degree relatives of patients with type 2 diabetes. We intended to assess both the compensatory and the obesity-related part of insulin secretion. In order to dissect out the latter, matching for insulin sensitivity was mandatory to normalize for the compensatory part of hyperinsulinemia. In 154 healthy, glucose tolerant first degree relatives of patients with type 2 diabetes we directly quantified both insulin sensitivity (by euglycemic-glucose-clamp technique) and insulin secretion (oral glucose load; stimulated serum c-peptide). Insulin sensitivity was scattered over a wide range with a considerable overlap of both first degree relatives of patients with type 2 diabetes and 97 controls without a family history of diabetes. Average insulin sensitivity was higher in controls (8.0+/-0.3 vs. 7.1 + 0.2 ml x kg-l x min-1, p < 0.05). Prevalence of insulin resistance (defined as controls, lowest tertile for insulin sensitivity) was 40% in first degree relatives of patients with type 2 diabetes. Insulin secretion after oral glucose was significantly increased in insulin resistant first degree relatives of patients with type 2 diabetes compared to insulin sensitive first degree relatives of patients with type 2 diabetes. Early phase relative insulin secretion (30 min) expressed as x-fold increase above basal was smaller in insulin resistant first degree relatives of patients with type 2 diabetes than in insulin sensitive counterparts (5.3+/-0.4 vs. 7.3+/-0.5; p < 0.01). Body mass index was distributed over the whole range in insulin resistant first degree relatives of patients with type 2 diabetes. In the insulin sensitive subgroup absolute and relative secretion did not differ in obese (Body mass index >25 kg/m2) and insulin sensitivity-matched lean. In obese insulin resistant first degree relatives of patients with type 2 diabetes absolute hyperinsulinemia was combined with reduced and delayed relative early insulin release. In summary, degree and prevalence of insulin resistance is higher in first degree relatives of patients with type 2 diabetes than in controls. However, both groups are of heterogenous metabolic composition and family history as major discriminator should not be overestimated. Our data suggest, that hyperinsulinemia cannot simply be explained as a compensatory event to balance insulin resistance. Hypersecretion is associated with insulin resistance predominantly in combination with obesity. It might be speculated that adipose tissue derived signals to the beta-cell might lead to hypersecretion only in the genetic background that also leads to insulin resistance.

Perfusion-independent effect of bradykinin and fosinoprilate on glucose transport in Langendorff rat hearts.

Angiotensin-converting enzyme (ACE) inhibitor-stimulated glucose metabolism and perfusion in muscle tissue seem to be, at least in part, mediated by kinins. However, the relative contribution of direct metabolic or secondary hemodynamically induced effects is unclear. It was the aim of this study to characterize the effects of ACE inhibition and bradykinin on glucose transport while changes in cardiocoronary function that might influence glucose transport were minimized. Hearts from Wistar rats were perfused by a Langendorff preparation and a set of functional parameters were simultaneously measured. Bradykinin (10[-11] M) and fosinoprilate (10[-7] M) were administered at concentrations that did not affect coronary flow. Insulin was employed as reference at half-maximal concentration. The nonmetabolizable glucose analog 3-O-[14C]methyl-D-glucose and the nontransportable tracer L-[3H]glucose were coperfused for the calculation of glucose transport. Using a 2-compartment mathematical model we found that the glucose transport rate, which was doubled with insulin, was increased almost 3-fold by either bradykinin or fosinoprilate. In the presence of the B2 bradykinin receptor antagonist HOE 140 (D-Arg[Hyp3,Thi5,D-Tic7,Oic8]-bradykinin; icatibant), the effect of both agents was completely abolished. Both agents also induced minor changes in contractility/relaxation parameters that again were completely neutralized with icatibant. A perfusion-independent but B2-kinin receptor-dependent stimulating effect on glucose transport by either bradykinin or fosinoprilate is concluded. This effect could, in analogy to insulin be due to increased glucose transporter translocation, increased endothelium-derived nitric oxide formation, or--despite constant coronary flow conditions--secondary to altered cardiac function.

Insulin-induced translocation of GLUT 4 in skeletal muscle of insulin-resistant Zucker rats.

The genetically obese Zucker rat (fa/fa) is an animal model with severe insulin resistance of the skeletal muscle. We investigated whether a defect of insulin-dependent glucose transporter (GLUT 4) translocation might contribute to the pathogenesis of the insulin-resistant state. fa/fa rats, lean controls (Fa/Fa) as well as normal Wistar rats were injected intraperitoneally with insulin and were killed after 2 or 20 min, respectively. Subcellular fractions were prepared from hind-limb skeletal muscle and were characterized by determination of marker-enzyme activities and immunoblotting applying antibodies against alpha 1 Na+/K+ ATPase. The relative amounts of GLUT 1 and GLUT 4 were determined in the fractions by immunoblotting with the respective antibodies. Insulin induced an approximately two-fold increase of GLUT 4 in a plasma membrane and transverse tubule enriched fraction and a decrease in the low density enriched membrane fraction in all three groups of rats. There was a high individual variation in GLUT 4 translocation efficiency within the groups. However, no statistically significant difference was noted between the groups. No effect of insulin was detectable on the distribution of GLUT 1 or alpha 1 Na+K+ ATPase. The data suggest that skeletal muscle insulin resistance of obese Zucker rats is not associated with a lack of GLUT 4 translocation.

Local generation of kinins in working skeletal muscle tissue in man.

The effect of standardized isometric forearm work on circulating and local kinin concentrations was investigated in 12 healthy volunteers using the forearm catheter technique. Radioimmunological kinin determination in arterial blood and in the venous effluent of forearm muscle tissue was performed using a modification of Shimamoto's technique of blood sampling and kinin extraction. Under basal conditions, there was no arterio-venous difference of kinins. Throughout the whole experiment, arterial--reflecting systemically circulating--kinins did not change. In muscle venous blood, immunoreactive kinins were not significantly elevated during work, whereas a marked increase was detected in the recovery period (5.0 +/- 0.6 vs. 10.2 +/- 2.0 pmol/l; p less than 0.01). The data demonstrate, that kinins are locally generated in calculated amounts (32.7 +/- 8.4 fmol/(100 g x min) that are known to be sufficient to induce local vasodilatory and metabolic effects at the site of muscle contraction, but below the threshold for systemic cardiovascular actions.

[The method of the isolated perfused rat heart using the Langendorff model in nutrition research]. / Die Methodik des isoliert-perfundierten Rattenherzens nach Langendorff in der Ernährungsforschung.

The preparation technique and principle of the Langendorff model for the perfusion of isolated rat heart, as well as the assessment of circulatory and metabolic parameters in this model are described, and the role of the model in metabolic and nutrition research is discussed.

Effects of kallikrein (K), bradykinin (B) and insulin (I), on substrate metabolism in the isolated perfused rat heart.

In the isolated rat heart preparation, perfused at physiological glucose levels, effects of kallikrein (K), bradykinin (B) and insulin (I) on carbohydrate metabolism have been compared. A flow- and I-independent K-effect, obviously working via proteolytic kinin formation, was shown. While I elevated glucose-uptake into the heart preferably via accelerated transport across the membrane and increased rates of glycogen synthesis, B and K significantly activated PFK and reduced myocardial glucose uptake. The latter effect is probably due to an accelerated glycolytic flux leading to decreased G6P-levels and thereby stimulating glycogenolysis. Since aprotinin was able to abolish the K-effect, the latter may possibly be mediated via kinin release.