Walter Renn

The PPARγ2 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 γ2 (PPARγ2) was described, leading to the substitution of Proline to Alanine at codon 12. Mutations in PPARγ2 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 PPARγ 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 PPARγ2 molecule may have a role in subgroups prone to the development of obesity and Type II diabetes. [Diabetologia (1999) 42: 758–762]

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-α (TNF-α) is believed to influence skeletal muscle insulin resistance. Two G → A transitions in the promoter region of TNF-α 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-α–238 and –308 G→ A promoter polymorphisms.¶Results. For the –238 polymorphism, 83 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-α promoter polymorphisms in our cohort. The polymorphisms occur at the same frequencies in probands with either low or high insulin sensitivity. [Diabetologia (2000) 43: 181–184]

Effect of Hypoglycemia on β-Adrenergic Sensitivity in Normal and Type 1 Diabetic Subjects

OBJECTIVE The purpose of this study was to assess the potential role of reduced tissue sensitivity to catecholamines in the pathogenesis of hypoglycemia unawareness in patients with type 1 diabetes. RESEARCH DESIGN AND METHODS The effect of a single episode of hypoglycemia on β-adrenergic sensitivity was studied in 10 type 1 diabetic patients with apparently normal awareness of hypoglycemia (age 29 ± 5 years, diabetes duration 13 ± 8 years, HbA1c 7.3 ± 0.9%) and 10 age-matched healthy control subjects. β-adrenergic sensitivity was measured with the isoproterenol test after a hyperinsulinemic euglycemic clamp and after a hyperinsulinemic hypoglycemic clamp. β-adrenergic sensitivity was expressed as the dose of intravenous isoproterenol that increased the heart rate by 25 beats/min (IC25). RESULTS During hypoglycemia, diabetic subjects had an impaired plasma epinephrine response compared with that of the control subjects (16.7 ± 5.0 vs. 40.1 ± 6.8 ng/ml, P = 0.02). In control subjects, the IC25 was lower after hypoglycemia than after euglycemia (0.83 ± 0.22 vs. 1.13 ± 0.21 μg, P = 0.02) indicating an increase in β-adrenergic sensitivity. In diabetic subjects, on the other hand, the IC25 was greater after hypoglycemia than after euglycemia (1.00 ± 0.26 vs. 0.65 ± 0.14 μ, P = 0.04), indicating a decrease in β-adrenergic sensitivity. CONCLUSIONS In normal subjects, a single episode of hypoglycemia increases β-adrenergic sensitivity. In diabetic subjects, in contrast, hypoglycemia reduces β-adrenergic sensitivity. These results provide evidence that in type 1 diabetic patients, some maladaptation of tissue sensitivity to catecholamines contributes to the development of hypoglycemia unawareness. A unifying hypothesis is presented for the pathogenesis of hypoglycemia unawareness in type 1 diabetic patients incorporating the concepts of both a reduced catecholamine response and reduced adrenergic sensitivity.

Desensitization of platelets to iloprost. Loss of specific binding sites and heterologous desensitization of adenylate cyclase.

The binding characteristics of [3H]prostacyclin and [3H]iloprost ([3H]5-[(E)-(1S,5S,6R,7R)-7-hydroxy-6-[(E)-(3S,4RS) -3-hydroxy-4-methyl-1-octen-6-inyl]-bicyclo[3.3.0]octan-3-yl idene] -pentanoic acid) and platelet adenylate cyclase activities were investigated in platelet-rich plasma preincubated with iloprost. The exposure of platelets to 0.1 microM iloprost (12 h, 20 degrees C) caused a significant loss of iloprost binding sites (P less than 0.01) without causing changes in binding affinity. This loss of specific [3H]iloprost binding was time- and dose-dependent. The reduction of iloprost receptor density was accompanied by an impaired responsiveness of platelet adenylate cyclase to iloprost, prostaglandin D2 and forskolin. In contrast, basal adenylate cyclase activity was not affected by iloprost pretreatment. The diminished response of the enzyme to GTP and NaF pointed to an involvement of the stimulatory guanyl nucleotide-binding protein (Gs) in iloprost-induced heterologous desensitization. Consequently, [32P]NAD+ and cholera toxin were used for the direct labelling of Gs. Platelet membranes desensitized to iloprost incorporated less label into the 45 kD subunit of Gs. These data suggest that the site of action of iloprost for heterologous desensitization of human platelet adenylate cyclase is located on Gs.

Lipolysis in skeletal muscle is rapidly regulated by low physiological doses of insulin

Aims/hypothesis. Both patients with Type II (non-insulin-dependent) diabetes mellitus and normoglycaemic, insulin resistant subjects were shown to have an increased lipid content in skeletal muscle, which correlates negatively with insulin sensitivity. Recently, it was shown that during a hyperinsulinaemic euglycaemic clamp interstitial glycerol was reduced not only in adipose tissue but also in skeletal muscle. To assess whether lipolysis of muscular lipids is also regulated by low physiological concentrations of insulin, we used the microdialysis technique in combination with a 3-step hyperinsulinaemic glucose clamp. Methods. Nineteen lean, healthy subjects (12 m/7 f) underwent a glucose clamp with various doses of insulin (GC I = 0.1, GC II = 0.25 and GC III = 1.0mU · kg–1· min–1). Two double lumen microdialysis catheters each were inserted in the paraumbilical subcutaneous adipose tissue and in skeletal muscle (tibialis anterior) to measure interstitial glycerol concentration (index of lipolysis) and ethanol outflow (index of tissue flow). Results. During the different steps of the glucose clamp, glycerol in adipose tissue was reduced to 81 ± 7 % (GC I), 55 ± 8 % (GC II) and 25 ± 5 % (GC III), respectively, of basal. In contrast, glycerol in skeletal muscle declined to 73 ± 5 % (GC I) and to 57 ± 6 % (GC II) but was not further reduced at GC III. Tissue flow was higher in the skeletal muscle and remained unchanged in both compartments throughout the experiment. Conclusion/interpretation. This study confirms the presence of glycerol release in skeletal muscle. Lipolysis in skeletal muscle and adipose tissue are suppressed similarly by minute and physiological increases in insulin but differently by supraphysiological increases. Inadequate suppression of intramuscular lipolysis resulting in increased availability of non-esterified fatty acids, could represent a potential mechanism involved in the pathogenesis of impaired glucose disposal, i. e. insulin resistance, in muscle. [Diabetologia (1999) 42: 1171–1174]

Platelet prostacyclin binding in coronary artery disease

Reduced responsiveness of platelets to prostacyclin, reported in vitro in patients with coronary artery disease, has been thought to be a factor predisposing toward coronary thrombosis and vasospasm as a result of enhanced in vivo release of cyclic endoperoxides and thromboxane A2 by the platelets. In this study, specific binding of prostacyclin to intact platelets was determined in patients with coronary artery disease by direct binding studies using 9-3H-prostacyclin sodium salt. In addition, the inhibitory effect of prostacyclin on primary aggregation induced by adenosine diphosphate and cyclic adenosine monophosphate (cyclic AMP) accumulation stimulated by prostacyclin was examined. Twenty patients with angiographically documented coronary artery disease and stable angina, 8 patients with acute myocardial infarction, 14 healthy volunteers and 10 patients with normal angiograms were studied. In patients with stable angina, binding capacity and affinity of platelet prostacyclin binding sites and prostacyclin-induced cyclic AMP accumulation were not different from those of control subjects. In patients with acute myocardial infarction, however, binding capacity of platelet prostacyclin receptors was significantly reduced (0.69 +/- 0.45 versus 1.35 +/- 0.37 pmol/10(9) platelets, p = 0.001) and the postreceptor response, represented by platelet responsiveness to prostacyclin and prostacyclin-induced cyclic AMP synthesis, was impaired. Because all patients with myocardial infarction were receiving intravenous heparin and nitroglycerin, which might interfere with platelet prostacyclin binding, competition experiments were performed in vitro. Neither heparin (3 to 250 IU/ml) nor nitroglycerin (0.8 to 22 microM) displaced specifically bound 9-3H-prostacyclin. L-Epinephrine in concentrations up to 10 microM also exhibited no competition with specific platelet prostacyclin binding.(ABSTRACT TRUNCATED AT 250 WORDS)

Leptin levels in humans are acutely suppressed by isoproterenol despite acipimox-induced inhibition of lipolysis, but not by free fatty acids

Leptin secretion is complexly regulated in humans. Insulin has been shown to stimulate leptin secretion, whereas in vitro data suggest that catecholamines and free fatty acids (FFAs) inhibit leptin secretion. To dissect differential effects on leptin secretion, we performed two experimental protocols in 11 lean healthy subjects in addition to a saline infusion plus oral acipimox to suppress lipolysis (SAL + ACX) as a control experiment: (1) isoproterenol (approximately 30 ng/kg x min, to increase the heart rate by approximately 50 bpm) plus oral acipimox (ISO + ACX, 240 minutes) and (2) Intralipid (Pharmacia & Upjohn, Erlangen, Germany) plus heparin (LIP, 420 minutes). During SAL + ACX, FFAs decreased from 0.44 +/- 0.04 to 0.06 +/- 0.02 mmol/L (P = .001), while serum insulin and leptin remained unchanged. During ISO + ACX, FFAs decreased similarly from 0.41 +/- 0.13 to 0.09 +/- 0.02 mmol/L (P= .001), while insulin increased from 47 +/- 8 to a maximum of 116 +/- 15 pmol/L (P= .001) and serum leptin decreased acutely from 6.4 +/- 2.1 to a minimum of 5.4 +/- 1.8 ng/mL after 90 minutes (P = .003 vSAL + ACX). After 150 minutes, leptin returned to control levels. During LIP, the elevation of FFAs from 0.34 +/- 0.04 to 1.71 +/- 0.19 mmol/L (P = .001) had no effect on serum insulin or leptin concentrations (both P = nonsignificant). In conclusion, our results show that in humans, isoproterenol acutely suppresses leptin levels independently of increased FFAs, and elevated FFAs have no acute effect on leptin levels. The fact that an inhibition of leptin secretion occurred despite conditions that are known to suppress intracellular cyclic adenosine monophosphate (cAMP) levels, as demonstrated by suppressed lipolysis, suggests that signaling mechanisms other than those mediated by cAMP must be involved in modulating leptin secretion.

Diabetes teaching program improves glycemic control and preserves perception of hypoglycemia

Improvement of HbA1c is frequently accompanied by deteriorating awareness of hypoglycemia. We studied the effect of improved metabolic control on hypoglycemia perception in 33 type 1 diabetic patients during 3 months after an inpatient diabetes education program of 5 days. Patients were grouped according to the presence (H, n = 11) or the absence (N, n = 22) of a history of repeated severe hypoglycemia. To measure awareness of blood glucose (BG) and hypoglycemia, we calculated their accuracy of BG perception (error grid analysis) and sensitivity for BG levels < 3.9 mmol/l, respectively, during the first (I) and second (II) period of the 3 months using the method of BG estimation. HbA1c decreased from 8.0 +/- 0.3% before to 7.1 +/- 0.2% 3 months after the program (P < 0.001) with no difference between H and N. Neither accuracy of BG perception (40.6 +/- 3.8 (I) versus 43.6 +/- 4.1% (II), P = 0.25) nor sensitivity for low BG levels (49.1 +/- 4.2 (I) versus 54.9 +/- 4.9% (II), P = 0.12) changed significantly. Group H had a lower overall accuracy of BG estimation (P = 0.048) and a lower overall sensitivity for detecting BG levels < 3.9 mmol/l (P = 0.03) than group N. Group H was able to improve accuracy of BG estimation (H: 24.8 +/- 6.2 (I) versus 36.9 +/- 8.3% (II), P = 0.04) while group N was not (48.5 +/- 3.9 (I) versus 46.9 +/- 4.6% (II), P = 0.5). In conclusion, improvement of metabolic control after intensive diabetes education had no adverse effect on the perception of low BG levels. On the contrary, patients with a history of severe hypoglycemia improved their awareness of BG.

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.

Pain Intensity and Blood Pressure Reactions During a Cold Pressor Test in IDDM Patients

OBJECTIVE To determine the contribution of altered pain perception to the impaired blood pressure reactions during a cold pressor test in diabetic patients. Reduced blood pressure increases have been observed in diabetic patients during a cold pressor test and have been attributed to an impaired efferent sympathetic function. RESEARCH DESIGN AND METHODS We investigated pain intensities and blood pressure reactions simultaneously during a cold pressor test in 30 IDDM patients (diabetes duration 12 ± 6 years, HbA1c 7.5 ± 1.4%) and in 30 normal control subjects with comparable sex distribution, age, height, BMI, physical fitness, and smoking habits. RESULTS Initial pain intensities and respective time courses did not differ between the two groups. The initial blood pressure response was significantly smaller in diabetic patients (P < 0.002). Correlations of diastolic blood pressure increases in diabetic patients with initial pain intensity, standard cardiovascular reflex tests, age, clock time, smoking habits, disease duration, and actual blood glucose concentrations did not reach statistical significance. Pain intensity and diastolic blood pressure increases, however, were correlated to HbA1c concentrations in diabetic patients. CONCLUSIONS Impaired pain perception is not the cause of the impaired reactions of blood pressure in diabetic patients during the cold pressor test, leaving very early deterioration of either cerebral processing of pain stimuli, cardiac function, efferent sympathetic nerves, or decreased vascular reactivity as possible explanations.