HbA(1c) in adults without known diabetes from southern Europe. Impact of the new diagnostic criteria in clinical practice.

AIMS: To analyse the differences in the prevalence of diabetes and dysglycaemia using fasting plasma glucose and HbA(1c) criteria. METHODS: Analytical cross-sectional study undertaken in a random sample of 2144 individuals (age 18-80 years) without known diabetes from the primary care setting in Malaga (Spain). Dysglycaemia was defined as fasting plasma glucose 5.6-6.9 mmol/l or HbA(1c) 39-46 mmol/mol (5.7-6.4%) and diabetes as fasting plasma glucose ≥ 7.0 mmol/l or HbA(1c)≥ 48 mmol/mol (≥ 6.5%). RESULTS: The proportion of subjects who were normoglycaemic was significantly higher using fasting plasma glucose than HbA(1c) (83.5 vs. 65%) (P < 0.0001). Compared with fasting plasma glucose, HbA(1c) detects more cases of dysglycaemia (32 vs. 14.8%) (P < 0.0001) and diabetes (3 vs. 1.7%) (P < 0.0001). CONCLUSIONS: In our environment, using HbA(1c) for the diagnosis of pre-diabetes and diabetes could increase the target population for preventive and therapeutic measures. Further cost-effectiveness studies are needed before the widespread diagnostic use of HbA(1c) can be recommended.

Diagnostic difficulties in glucokinase hyperinsulinism.

Glucokinase hyperinsulinism is a rare variant of congenital hyperinsulinism caused by activating mutations in the glucokinase gene and has been reported so far to be a result of overactivity of glucokinase within the pancreatic beta-cell. Here we report on a new patient with difficulties to diagnose persistent hyperinsulinism and discuss diagnostic procedures of this as well as the other reported individuals. After neonatal hypoglycemia, the patient was reevaluated at the age of 3 years for developmental delay. Morning glucose after overnight fast was 2.5-3.6 mmol/l. Fasting tests revealed supressed insulin secretion at the end of fasting (1.4-14.5 pmol/l). In addition, diagnostic data of the patients reported so far were reviewed. A novel heterozygous missense mutation in exon 10 c.1354G>C (p.Val452Leu) was found and functional studies confirmed the activating mutation. There was no single consistent diagnostic criterion found for our patient and glucokinase hyperinsulinism individuals in general. Often at the time of hypoglycemia low insulin levels were found. Therefore insulin concentrations at hypoglycemia, or during fasting test as well as reactive hypoglycemia after an oral glucose tolerance test were not conclusive for all patients. A glucose lowering effect in extra-pancreatic tissues independent from hyperinsulinism that results in diagnostic difficulties may contribute to underestimation of glucokinase hyperinsulinism. Mutational analysis of the GCK-gene should be performed in all individuals with unclear episodes of hypoglycemia even without documented hyperinsulinism during hypoglycemia. Delay of diagnosis might result in mental handicap of the affected individuals.

Presence of functional cannabinoid receptors in human endocrine pancreas.

AIMS/HYPOTHESIS: We examined the presence of functional cannabinoid receptors 1 and 2 (CB1, CB2) in isolated human islets, phenotyped the cells producing cannabinoid receptors and analysed the actions of selective cannabinoid receptor agonists on insulin, glucagon and somatostatin secretion in vitro. We also described the localisation on islet cells of: (1) the endocannabinoid-producing enzymes N-acyl-phosphatidyl ethanolamine-hydrolysing phospholipase D and diacylglycerol lipase; and (2) the endocannabinoid-degrading enzymes fatty acid amidohydrolase and monoacyl glycerol lipase. METHODS: Real-time PCR, western blotting and immunocytochemistry were used to analyse the presence of endocannabinoid-related proteins and genes. Static secretion experiments were used to examine the effects of activating CB1 or CB2 on insulin, glucagon and somatostatin secretion and to measure changes in 2-arachidonoylglycerol (2-AG) levels within islets. Analyses were performed in isolated human islets and in paraffin-embedded sections of human pancreas. RESULTS: Human islets of Langerhans expressed CB1 and CB2 (also known as CNR1 and CNR2) mRNA and CB1 and CB2 proteins, and also the machinery involved in synthesis and degradation of 2-AG (the most abundant endocannabinoid, levels of which were modulated by glucose). Immunofluorescence revealed that CB1 was densely located in glucagon-secreting alpha cells and less so in insulin-secreting beta cells. CB2 was densely present in somatostatin-secreting delta cells, but absent in alpha and beta cells. In vitro experiments revealed that CB1 stimulation enhanced insulin and glucagon secretion, while CB2 agonism lowered glucose-dependent insulin secretion, showing these cannabinoid receptors to be functional. CONCLUSIONS/INTERPRETATION: Together, these results suggest a role for endogenous endocannabinoid signalling in regulation of endocrine secretion in the human pancreas.

Hiperinsulinismo monogénico / Monogenic hyperinsulinism

El término hiperinsulinismo monogénico se refiere a casos de hiperinsulinemia causados por mutaciones en un solo gen. Los pacientes presentan hipoglucemias de ayuno recurrentes, niveles inadecuados de insulina e incremento de la glucemia tras la administración de glucagón endovenoso. Además, no existe cetonemia, cetonuria ni acidosis. La principal causa de este cuadro clínico son las canelopatías, en las que el hiperinsulinismo está producido por alteraciones estructurales de los canales de potasio dependientes del ATP como consecuencia de mutaciones en el receptor de la sulfonilurea 1 (SUR1) o en el rectifi cador interno de los canales de potasio (Kir6.2). La segunda causa más común es el síndrome de hiperinsulinismo-hiperamonemia, originado por mutaciones activadoras de la enzima glutamato deshidrogenasa (GDH). Este síndrome se caracteriza por cuadros de hipoglucemia hiperinsulinémica con niveles elevados de amonio, que pueden ser provocados por la ingestión de una comida rica en proteínas. Otra causa de hiperinsulinismo monogénico es el hiperinsulinismo inducido por mutaciones activadoras en el gen de la glucocinasa (GGK). Finalmente, debe incluirse también la mutación en la enzima mitocondrial 3-hidroxiacil-CoA deshidrogenasa de cadena corta (SCHAD), que cataliza el tercero de los cuatro pasos de la oxidación de los ácidos grasos en la mitocondria

The term monogenic hyperinsulinism refers to cases of hyperinsulinism caused by mutations in a single gene. The affected patients show recurrent fasting hypoglycemia, inadequate serum insulin levels, and an increase in plasma glucose levels following the administration of intravenous glucagon. In addition, there is an absence of ketonemia, ketonuria and acidosis. The main causes of these syndromes are channelopathies, in which hyperinsulinism is caused by structural changes in the ATP-sensitive potassium channels due to mutations in sulfonylurea receptor 1 (SUR1) or in Kir6.2, the pre-forming subunit of this channel. The second most frequent cause is the hyperinsulinism/hyperammonemia syndrome, caused by activating mutations of the glutamate dehydrogenase (GDH) enzyme. This syndrome is characterized by episodes of hypoglycemia with hyperinsulinism and elevated levels of ammonium, which can be triggered by the ingestion of a protein- rich meal. Monogenic hyperinsulinism can also be induced by activating mutations of the glucokinase gene. Finally, mutations of mitochondrial short-chain 3-hydroxyacyl-coenzyme A dehydrogenase (SCHAD), which catalyses the third of the four steps in mitochondrial fatty acid oxidation, should also be included

Glutaminase activity is confined to the mantle of the islets of Langerhans.

Glutamatergic signalling plays an important role in the coordination of hormone secretion from the endocrine pancreas. Thus, glutamate production must be a process exquisitely regulated to ensure a proper transmitter function. Recently we have reported that the endocrine pancreas co-expresses two isoforms of the protein glutaminase (GA), denoted as kidney-type (KGA) and liver-type (LGA). However, how GA activity, and therefore glutamate production, is regulated in the islets represents a critical issue that remains unresolved. Since the purification of these enzymes from rat islets is a daunting task, in order to characterize each isoform we have taken advantage of the spatial segregation of these isoenzymes in pancreas. To assist us with this goal, we have developed a new procedure that enables us to assay GA activity in situ. The assay is highly specific for GA as indicated by its dependence on glutamine and orthophosphate. Surprisingly, LGA, which is abundantly expressed by beta-cells, did not show detectable activity under the assay conditions. All the GA activity detected in pancreatic islets was attributed to KGA and was confined to the mantle of the islets. Double labelling analyses strongly suggested that alpha-cells should be regarded as the site of glutamate production in the endocrine pancreas.

Differential analysis of donor characteristics for pancreas and islet transplantation.

Suitable selection of donors is key to the success of human islet isolation and transplantation. Although several important donor-related factors have been identified previously, they needed to be confirmed in our setting. The aims of this study were: (1) to compare the characteristics of islet donors with those of pancreas donors (national transplant registry). (2) to compare the characteristics of islet donors resulting in a successful isolation in our facility with the characteristics of pancreas donors, and (3) to compare the characteristics of islet donors at this facility, whether or not isolation was successful, with donors elsewhere whose islets were transplanted and included in the Collaborative Islet Transplant Registry. The 35 islet isolations completed at our facility were analyzed for various characteristics. Significant differences were seen in donor age body mass index (BMI), and body weight between our islet donors and our pancreas donors (P < .001). These differences were maintained in the subgroup analysis corresponding to donors of successful isolations compared to pancreas donors (P < .01). Most successful isolations in our islet isolation facility were associated with donors of BMI >25. The percentage of successful isolation (>300,000 IEq) was higher among donors with a body weight >90 kg. We concluded that there was little overlap between the donor profiles for pancreas transplantation and for islet transplantation. More specific selection criteria relative to both BMI and body weight for islet donors may result in greater success of pancreas islet isolation and transplantation.

Internal assessment of a novel islet isolation facility in Spain.

UNLABELLED: Islet transplantation is a promising therapy in the treatment of diabetes mellitus. Herein we present the result from the first series of islet isolations carried out in our new islet isolation facility. The aims of study were to analyze the influence of various donor characteristics on the success of islet isolation and compare these outcomes with other European and American groups. Data from 22 completed islet isolation were used to compare donor and isolation variables among successful (>300,000 IEQs) versus unsuccessful isolations. The successful isolation rate from our laboratory was 31.8%. We did not see any significant differences between successful and unsuccessful groups according to donor characteristics, although age was close to significance (38.57 +/- 10.29 versus 48.33 +/- 12.39; P = .08). Donor age (1.12 [1.23; 0.99]) and body mass index (0.065 [1.32; 3.08]) were associated with isolation success in a logistic regression model. We did not find differences among intraprocedure variables with the exception of IEQ prepurification (409,073 +/- 115,041 versus 263,776 +/- 128,988; P < .05). IEQpre and IEQpost were positively correlated (P < .05). In comparison with other groups, we observed differences in some cases related to islet yield prepurification (P < .05) but not postpurification. Purity from our islet preparations was the highest from all considered groups (P < .05). Recovery was similar in all groups. CONCLUSIONS: In our experience, donor characteristics have no influence on the success rate. The digestion step is a critical factor for success. Our results with respect to IE yield were close to that of experienced groups.

Monounsaturated n-9 fatty acids and adipocyte lipolysis in rats.

To investigate the role of the monounsaturated n-9 fatty acids (MUFA) in the lipolytic activity of adipocytes, a study was carried out in which an increase in MUFA was produced in the tissues by two different methods; by the dietary enrichment of oleic acid or by producing an essential fatty acid deficiency syndrome. For this, forty-five male Sprague-Dawley rats were fed with a normal-energy diet and were subdivided into three groups. The diets varied in the type of dietary fat; palmitic acid, olive oil, or soyabean oil+palmitic acid. At the end of the study measurements were taken of weight, plasma leptin, tissue concentration of fatty acids, fat-cell size in the epididymal and the omental adipose tissues, adipocyte lipolytic activity of both tissues after stimulation with adrenaline, and the capacity of insulin to inhibit lipolysis. The baseline and adrenaline-stimulated lipolytic activity were greater and the anti-lipolytic capacity of insulin lower in the animals undergoing an increase in MUFA in the tissues (palmitic-acid and olive-oil diets). The area under the curve of glycerol, used as an indicator of lipolytic activity, was positively correlated with the concentration of MUFA and negatively with polyunsaturated fatty acids in the adipose tissues. It is concluded that an increase in tissue MUFA, however obtained, induces an increase in lipolytic activity.