Discovery of a functionally selective ghrelin receptor (GHSR1a) ligand for modulating brain dopamine.

SignificanceThe modulation of growth hormone secretagogue receptor-1a (GHSR1a) signaling is a promising strategy for treating brain conditions of metabolism, aging, and addiction. GHSR1a activation results in pleiotropic physiological outcomes through distinct and pharmacologically separable G protein- and ß-arrestin (ßarr)-dependent signaling pathways. Thus, pathway-selective modulation can enable improved pharmacotherapeutics that can promote therapeutic efficacy while mitigating side effects. Here, we describe the discovery of a brain-penetrant small molecule, N8279 (NCATS-SM8864), that biases GHSR1a conformations toward Gαq activation and reduces aberrant dopaminergic behavior in mice. N8279 represents a promising chemical scaffold to advance the development of better treatments for GHSR1a-related brain disorders involving the pathological dysregulation of dopamine.

Increasing incidence of type 1 diabetes mellitus in young children in Slovakia.

OBJECTIVES: We aimed to study the prevalence of the early onset Type 1 diabetes in Slovakia during the years 1996-2017. BACKGROUND: Prevalence of Type 1 diabetes in young children is increasing worldwide. However, recent data from Slovakia are missing. METHODS: All children with newly diagnosed Type 1 diabetes included in the study were diagnosed in the Children Diabetes Centre in Bratislava during the years 1996-2017. The incidence of T1D in children aged below 3 and below 5 years was calculated and compared to the T1D incidence in older children. Incidence trends were calculated with the Poissed regression. RESULTS: Gender-adjusted incidence of T1D annually increased by 5.4 % (CI: 3.9-6.8; p < 0.001) in children <3 years, and by 3.4 % (95 % CI 2.2-4.6; p<0.001) in children <5 years during the last two decades. Moreover, the proportion of young children <3 years of age among all newly diagnosed children and adolescents increased over time (4.2±2.8 % in years 1996-1998; 12.2±5.8 % in years 2004-2008, and 13.7±7.4 % during the years 2013-2017). CONCLUSION: We found a significant increase in the incidence and proportion of T1D in young children during the last two decades. Similar data were also found in other European countries. This could be explained by changing environmental conditions (Fig. 1, Ref. 32).

Neonatal hypoglycemia, early-onset diabetes and hypopituitarism due to the mutation in EIF2S3 gene causing MEHMO syndrome.

Recently, the genetic cause of several syndromic forms of glycemia dysregulation has been described. One of them, MEHMO syndrome, is a rare X-linked syndrome recently linked to the EIF2S3 gene mutations. MEHMO is characterized by Mental retardation, Epilepsy, Hypogonadism/hypogenitalism, Microcephaly, and Obesity. Moreover, patients with MEHMO had also diabetes and endocrine phenotype, but detailed information is missing. We aimed to provide more details on the endocrine phenotype in two previously reported male probands with MEHMO carrying a frame-shift mutation (I465fs) in the EIF2S3 gene. Both probands had a neonatal hypoglycemia, early onset insulin-dependent diabetes, and hypopituitarism due to dysregulation and gradual decline of peptide hormone secretion. Based on the clinical course in our two probands and also in previously published patients, neonatal hypoglycemia followed by early-onset diabetes and hypopituitarism may be a consistent part of the MEHMO phenotype.

Sulfonylurea vs insulin therapy in individuals with sulfonylurea-sensitive permanent neonatal diabetes mellitus, attributable to a KCNJ11 mutation, and poor glycaemic control.

BACKGROUND: Therapy with sulfonylurea is preferable to insulin in the majority of individuals with KCNJ11 mutations, but not all of these people achieve target levels of HbA1c in long-term follow-up. We aimed to compare sulfonylurea therapy with insulin treatment in two sulfonylurea-sensitive individuals with a KCNJ11 mutation who had poorly controlled permanent neonatal diabetes mellitus. CASE REPORT: We report on two individuals with a KCNJ11 mutation (p.R201H) who are currently aged 35 (SVK1) and 21 years (SVK2). These individuals were switched from insulin to sulfonylurea in 2005. Data from the first 4 (SVK2) and 8 years (SVK1) of the follow-up showed improved diabetes control and increased quality of life for both individuals. During the following years, however, both individuals failed to retain good diabetes control (HbA1c ≤ 53 mmol/mol; 7.0%). We therefore changed the therapy to a combination of insulin and sulfonylurea in both individuals, or to insulin monotherapy in SVK1, and compared the effects on HbA1c with those of sulfonylurea monotherapy. HbA1c levels in both individuals worsened after adding insulin to sulfonylurea [67 mmol/mol (8.3%) vs 77 mmol/mol (9.2%) in SVK1 and 106 mmol/mol (11.8%) vs 110±19 mmol/mol (12.2±1.7%) in SVK2], and after switching to only insulin therapy in SVK1 [57 mmol/mol (7.4%) vs 62 mmol/mol (7.8%)] when compared with sulfonylurea monotherapy. DISCUSSION: Our data show that sulfonylurea monotherapy might be preferable to insulin in people with permanent neonatal diabetes mellitus sensitive to sulfonylurea even when HbA1c is above target.

Prevalence of blood type A and risk of vascular complications following transcatheter aortic valve implantation.

OBJECTIVES: To assess the prevalence of blood type A among patients referred for transcatheter aortic valve implantation (TAVI) and whether it is related to vascular complications. BACKGROUNDS: Vascular complications following TAVI are associated with adverse outcomes. Various blood types, particularly type A, have been shown to be more prevalent in cardiovascular diseases and to be related to prognosis. METHODS: The prevalence of various blood types in a cohort of 491 consecutive patients who underwent TAVI was compared with a control group of 6500 consecutive hospitalised patients. The prevalence and predictors of vascular complications and bleeding events were evaluated in the blood type A group and were compared with non-type A patients. RESULTS: The mean age of TAVI patients was 83 ± 6 years, and 40 % were males. Patients were divided into two groups: blood type A (n = 220) and non-type A (n = 271). Type A was significantly more prevalent in the TAVI group than in the control group (45 vs. 38 %, p = 0.023). Compared with the non-type A group, patients with blood type A had more major and fatal bleeding (14.5 vs. 8.1 %, p = 0.027) and more vascular complications (any vascular complication: 24.5 vs. 15.9 % p = 0.016; major vascular complications: 12.3 vs. 7 % p = 0.047). In a multivariable analysis, blood type A emerged as a significant and independent predictor for vascular complications and bleeding events. CONCLUSIONS: Blood type A is significantly more prevalent in TAVI patients than in the general population and is related to higher rates of vascular and bleeding complications.

[Impact of long-term glycemic control on changes of lipid profile in children and adolescents with 1 type diabetes mellitus]. / Vplyv dlhodobej glykemickej kompenzácie na zmeny v lipidovom profile detí a adolescentov s diabetes mellitus 1. typu.

INTRODUCTION: Abnormalities in lipid metabolism contribute significantly to the increased occurrence of cardiovascular events in individuals with T1DM compared to healthy subjects. Disorder of lipid metabolism in T1DM is heavily dependent on maintaining of blood glucose values near the physiological range. DCCT study confirmed that patients with well compensated diabetes have similar lipid spectrum to the healthy subjects one. AIMS: We aimed to study relations of lipid profile parameters (cholesterol of high density HDL, cholesterol of low density LDL, total cholesterol - TC, triglycerides - TAG) to age, duration of T1DM (DD), blood glucose, HbA1c and if the blood pressure (BP), BMI corrected for age (BMIc) and daily insulin doses per kilogram (DI) in 30 patients with T1DM with good long-term glycemic compensation. We aimed also to find mathematical models of lipid profile parameters dependence of the parameters of glycemic control, age, duration of DM1T, blood pressure (systolic and diastolic BPs, BPd, respectively) BMIc and DI. RESULTS: HbA1c levels were significantly higher in diabetic patients compared to controls (p < 0.01), HDL were higher in diabetics than in controls (not significantly). LDL levels were in diabetics similar to controls. TAG was significantly lower in diabetics than in controls (p < 0.01). HDL significantly positively correlated with HbA1c (r = 0.372, p < 0.05) and negatively with BPs (r = -0.373, p < 0.05), TAG correlated with age (r = 0.546, p < 0.01), DD (r = 0.577, p < 0.001) and BPs (r = 0.407, p < 0.05). We also found a statistically appropriate mathematical models of the relationship of HDL and TAG with the parameters: age, DD, glucose, HbA1c, BP, BMIc, DI (r = 0.785, r2 = 0.616, p < 0.01, R = 0.758; r2 = 0.574, p < 0.05, respectively). CONCLUSION: The changes in HDL and TAG values in juvenile diabetics are significantly affected by particularly long-term glycemic control and insulin therapy.

Comparative study of serum/plasma glycation and lipid peroxidation of young patients with type 1 diabetes mellitus in relation to glycemic compensation and the occurrence of diabetic complications.

AIM OF THE STUDY: We tried to investigate whether the AGEs in serum and lipoperoxides (LPO) monitoring were suitable for an early prediction of diabetic complications (DC) development in diabetological practice. We wanted to find whether it is better to divide the file according to the presence of DC or in terms of glycemic compensation in this study. PATIENTS AND METHODS: 79 diabetic patients with duration of disease for at least 5 years were divided in respect to DC presence/absence and also to long-time glycemic compensation. HbA1c was measured by LPLC in fair capillary blood, s-AGEs were estimated spectrofluorimetrically and LPO iodimetrically and spectrophotometrically in serum. RESULTS: HbA1c, s-AGEs and LPO were significantly higher in the group with DC (+DC) vs. controls and also in -DC vs. controls. HbA1c and s-AGEs were significantly higher in +DC vs. patients without DC (-DC). HbA1c, s-AGEs and LPO were significantly higher in patients with poor glycemic compensation (PGC) compared to controls and HbA1c and LPO in patients with good glycemic compensation (GGC) compared to controls. HbA1c and s-AGEs were significantly higher in PGC vs. GGC. In the group of GGC we have found interesting significant correlations of HbA1c with HDL (r=0.451, p<0.05) and with LDL (r=-0.450, p<0.05). CONCLUSIONS: Our findings suggest that the monitoring of s-AGEs in poorly compensated diabetic patients and LPO in all may be very useful to recognize the risk of complications. The dividing of patient file in terms of long time glycemic compensation is more reliable for research of this issue (Tab. 3, Fig. 6, Ref. 41). Full Text in free PDF www.bmj.sk.

Glycation and lipid peroxidation in children and adolescents with type 1 diabetes mellitus with and without diabetic complications.

Glycation and oxidative stress lead to formation of compounds that have several biological effects and contribute to the development of the complications of diabetes mellitus. All steps of glycoxidation generate oxygen free radicals, some of them in common with lipid peroxidation pathways. Some oxidation or lipid peroxidation products may bind to proteins and amplify glycoxidation-generated lesions. The aim of this study was to measure glycation and lipid peroxidation parameters and examine the relationship between them in patients with type 1 diabetes mellitus (DM1) with (+DC) and without (-DC) diabetic complications. Fifty patients with DM1 aged from 7-19 years and with duration of DM1 (DD) at least 5 years were included. Twenty-four patients were -DC and 26 were +DC. Twelve healthy children formed a control group. There were significantly higher values of fructosamine (FAM), HbA(1c), serum advanced glycation endproducts (s-AGEs) and lipid peroxides (LPO) in the +DC group compared with -DC, and significantly higher values of HbA(1c), FAM and LPO in both diabetic groups than in controls. The s-AGEs level in the -DC group was similar to that in controls. In the total diabetic group, regardless of DC, there was a significant negative correlation between LPO and HDL-C (r = -0.379; p <0.05), and a positive correlation between LPO and triacylglycerol (TAG) (r = 0.852; p <<0.05), FAM (r = 0.414; p <0.05) and s-AGEs (r = 0.454; p <0.05). In the +DC group LPO correlated negatively with HDL-C (r = -0.392, p <0.05) and positively with TAG (r = 0.848; p <<0.05), FAM (r = 0.457; p = 0.02), and s-AGEs (r = 0.516, p = 0.02), whereas in the -DC group LPO correlated only with HDL-C (r = -0.441; p = 0.03) and TAG (r = 0.769; p <<0.05). We demonstrated a linkage between enhanced formation of AGEs and lipid peroxidation products and the presence of diabetic complications. Thus, the overproduction of glycation and lipid peroxidation products may take part in DC development as early as in childhood and adolescence.

HbA1c and serum levels of advanced glycation and oxidation protein products in poorly and well controlled children and adolescents with type 1 diabetes mellitus.

Diabetes mellitus is associated with hyperglycemia and with accelerated non-enzymatic glycation, increased oxidative stress and free radical production. The aim of the present study was to evaluate the levels of proteins glycation and oxidation parameters, compare them between poorly and well controlled children with type 1 diabetes mellitus, and determine the impact of glycemic control on these parameters. Blood and serum were obtained from 81 patients with type 1 diabetes mellitus (DM1) (20 patients had long-term good glycemic control [GGC], 61 patients had long-term poor glycemic control [PGC]). Thirty-one healthy children were used as controls. Fructosamine (FAM) was determined by a spectrophotometric method, HbA1c was measured by LPLC, serum advanced glycation end-products (s-AGEs) were determined fluorimetrically, and advanced oxidation protein products (AOPP) were measured spectrophotometrically. We observed significantly higher FAM, HbA1c, s-AGEs and AOPP levels in the patients with DM1 compared with controls, and significantly higher FAM, HbA1c and sAGEs levels in the PGC group compared with the GGC group. AOPP was higher in the PGC group than in the GGC group, but not significantly. In the PGC group we observed significant correlations between HbA1c and HDL-C (r = -0.306, p = 0.01), HbA1c and s-AGEs (r = 0.486, p < 0.001), and HbA1c and AOPP (r = 0.447, p < 0.01). s-AGEs significantly correlated with triacylglycerols (TAG) (r = 0.537, p < 0.001) and AOPP with HDL-C (r = -0.336, p < 0.05), TAG (r = 0.739, p < 0.001) and s-AGEs (r = 0.577, p < 0.001). In conclusion, our results showed both glycative and oxidative stress are increased in the PGC diabetic group compared with controls, they are linked with glycemic control, and probably contribute to the development of diabetic complications. We suggest that the measurement of not only HbA1c but also s-AGEs and AOPP may be useful to predict the risk of development of diabetic complications.

Identification of a novel beta-cell glucokinase (GCK) promoter mutation (-71G>C) that modulates GCK gene expression through loss of allele-specific Sp1 binding causing mild fasting hyperglycemia in humans.

OBJECTIVE: Inactivating mutations in glucokinase (GCK) cause mild fasting hyperglycemia. Identification of a GCK mutation has implications for treatment and prognosis; therefore, it is important to identify these individuals. A significant number of patients have a phenotype suggesting a defect in glucokinase but no abnormality of GCK. We hypothesized that the GCK beta-cell promoter region, which currently is not routinely screened, could contain pathogenic mutations; therefore, we sequenced this region in 60 such probands. RESEARCH DESIGN AND METHODS: The beta-cell GCK promoter was sequenced in patient DNA. The effect of the identified novel mutation on GCK promoter activity was assessed using a luciferase reporter gene expression system. Electrophoretic mobility shift assays (EMSAs) were used to determine the impact of the mutation on Sp1 binding. RESULTS: A novel -71G>C mutation was identified in a nonconserved region of the human promoter sequence in six apparently unrelated probands. Family testing established cosegregation with fasting hyperglycemia (> or = 5.5 mmol/l) in 39 affected individuals. Haplotype analysis in the U.K. family and four of the Slovakian families demonstrated that the mutation had arisen independently. The mutation maps to a potential transcriptional activator binding site for Sp1. Reporter assays demonstrated that the mutation reduces promoter activity by up to fourfold. EMSAs demonstrated a dramatic reduction in Sp1 binding to the promoter sequence corresponding to the mutant allele. CONCLUSIONS: A novel beta-cell GCK promoter mutation was identified that significantly reduces gene expression in vitro through loss of regulation by Sp1. To ensure correct diagnosis of potential GCK-MODY (maturity-onset diabetes of the young) cases, analysis of the beta-cell GCK promoter should be included.

Monitoring of early and advanced glycation in relation to the occurrence of microvascular complications in children and adolescents with type 1 diabetes mellitus.

The authors aimed to evaluate if the monitoring of serum advanced glycation end-products (s-AGEs) could help to predict a development of diabetic complications. Clinical and biochemical parameters including fructosamine (FAM), glycated hemoglobin (HbA1c) and serum AGEs were investigated in children and adolescents with 1 type diabetes with (+DC) and without (-DC) complications. FAM levels (in mmol/l) were significantly elevated in +DC diabetic group compared to -DC one (3.043+/-0.459 vs. 2.614+/-0.430; p<0.001) or to controls (3.043+/-0.459 vs. 1.620+/-0.340; p<0.001) as well as in -DC compared to controls (2.614+/-0.430 vs. 1.620+/-0.340; p<0.001). HbA1c (in %) were significantly elevated in +DC diabetic group compared to -DC one (10.48+/-1.83 vs. 8.41+/-1.19; p<<0.001) or to controls (10.48+/-1.83 vs. 5.0+/-0.38, p<<0.001) and also in -DC compared to controls (8.41+/-1.19 vs. 5.0+/-0.38; p<0.001). Serum AGEs levels (in A. U.) were significantly higher in +DC group than in -DC (73.0+/-14.09 vs. 65.8+/-9.05; p<0.05) and in group +DC than in controls (73.0+/-14.09 vs. 60.17+/-13.78; p<0.05), whereas there was no difference between -DC and controls. FAM correlated with HbA1c in both diabetic groups (+DC: r=0.374; p<0.05; -DC: r=0.719; p<0.001), but not in controls. Serum AGEs were correlated with HbA1c (r=0.478; p=0.003) in +DC, but not in -DC or controls. Enhanced serum AGEs levels show that they could be not only an attendant phenomenon of microangiopathies, but also a predictor of their development.

Regulation of M2-type pyruvate kinase mediated by the high-affinity IgE receptors is required for mast cell degranulation.

BACKGROUND AND PURPOSE: M2-type pyruvate kinase (M2PK) was found to interact directly with the 'ITAM' region of the gamma chain of the high-affinity IgE receptor (FcvarepsilonRI). Our hypothesis was that mast cell degranulation might require the FcvarepsilonRI-mediated inhibition of M2PK activity. EXPERIMENTAL APPROACH: In rat basophilic leukaemia (RBL-2H3) cells, the effects of directly inhibiting M2PK or preventing the FcvarepsilonRI-mediated inhibition of M2PK (disinhibition) on degranulation was measured by hexosaminidase release. Effects of blocking the FcvarepsilonRI-mediated inhibition of M2PK was also assessed in vivo in a mouse model of allergen-induced airway hyper-responsiveness. KEY RESULTS: Activation of FcvarepsilonRI in RBL-2H3 cells caused the rapid phosphorylation of tyrosine residues in M2PK, associated with a decrease in M2PK enzymatic activity. There was an inverse correlation between M2PK activity and mast cell degranulation. FcvarepsilonRI-mediated inhibition of M2PK involved Src kinase, phosphatidylinositol 3-kinase, PKC and calcium. Direct inhibition of M2PK potentiated FcvarepsilonRI-mediated degranulation and prevention of the FcvarepsilonRI-mediated inhibition of M2PK attenuated mast cell degranulation. Transfection of RBL-2H3 cells with M1PK which prevents FcvarepsilonRI-induced inhibition of M2PK, markedly reduced their degranulation and exogenous M1PK (i.p.) inhibited ovalbumin-induced airway hyper-responsiveness in vivo. CONCLUSIONS AND IMPLICATIONS: We have identified a new control point and a novel biochemical pathway in the process of mast cell degranulation. Our study suggests that the FcvarepsilonRI-mediated inhibition of M2PK is a crucial step in responses to allergens. Moreover, the manipulation of glycolytic processes and intermediates could provide novel strategies for the treatment of allergic diseases.

CTLA-4 gene polymorphisms predispose to autoimmune endocrinopathies but not to celiac disease.

OBJECTIVES: The aim of the study was to determine the association of two CTLA-4 gene polymorphisms (CT60, +49 A/G) with Hashimoto thyroiditis (HT), type 1 diabetes mellitus (T1DM) and celiac disease (CD) as well as with the occurrence of multi-organ involvement by autoimmunity in children. METHODS: Genotyping was done by RFLP analysis in Slovak children with HT (n=63) and CD (n=120) and both Slovak and Slovene children with T1DM (n=320) and healthy controls (n=231). RESULTS: We found a significant association of the G allele of the CT60 polymorphism with HT (p<0,0005) in the Slovak population and T1DM in both Slovak (p<0.01) and Slovene populations (p<0.005). The G allele of the +49A/G polymorphism was significantly, though less strongly, associated with T1DM (p<0.05) and HT (p<0.05). Distribution of genotypes of CTLA-4 gene polymorphisms in CD patients did not differ significantly from controls. None of the polymorphisms was associated with multi-organ involvement by autoimmunity. CONCLUSION: The G allele of both examined CTLA-4 gene polymorphisms predisposes to HT and T1DM, but not to CD. No association with multi-organ involvement was found. The GG genotype of the CT60 polymorphism may identify CD patients at an increased risk for concomitant T1DM and HT. Further studies to assess the predictive value of CTLA-4 polymorphisms for the co-occurrence of HT and T1DM in CD patients are needed.

Associations between HLA class II alleles and type 1 diabetes mellitus in the Slovak population.

OBJECTIVES: Several associations between HLA complex and diabetes mellitus type IA were found in various groups of patients of Caucasoid population. This study was therefore prompted to be conducted in Slovak population, since any such has not yet been performed in Slovak population. METHODS: Patients suffering from DM-1A originated from all regions of Slovakia. Their age ranged from 1 to 42 years; but the criterion for including the subject to the study was the definition of diagnosis in older patients before their age of 15 (Table 1). The diagnosis was set up according to internationally accepted criteria. A total of 460 patients was typed for HLA-DQB1 alleles, among them 97 also for HLA-DQA1 and 146 for HLA-DRB1 alleles. HLA-typing was performed by a PCR-SSP method. Control group consisted of 196 (DQA), 143 (DQB1) and 130 (DRB1) unrelated blood donors aged 19-55 years old irrespective of their age or sex. The data obtained were expressed in a 2 x 2 contingency table and statistical significance was calculated by the Fisher exact test. RESULTS: Among 11 HLA-DQB1 alleles tested DOB1*0302 was the most frequent in DM-1A patients (30.33% vs. 5.59% in healthy subjects (HS), followed by DQB1*0201 (22.93% vs. 12.94%, respectively). In contrast, the frequency rate of DQB1*0301 (10.66% vs. 24.48%), DOB1*0602 (2.17% vs. 10.14%) and DQB1*0603 (2.5% vs. 8.39 %) were decreased in DM-1A patients. Out of 14 DQA1 alleles the highest occurrence rate showed DQA1*0301 (30.93% vs. 17.09) and DQA1*0501 (34.02% vs. 25.76%), while DQA1*0102 (8.76% vs. 16.58%) and DQA1*0201 (6.18 % vs. 13.51%7), respectively, were found to be the least frequent. Among 13 HLA-DRB1 alleles tested, the most common occurrence rates showed DRB1*03 (26.37% vs. 9.62%) and DRB1*04 (7.19% vs. 14.23%), while the least frequent alleles were DRB 1*15 (2.74% vs. 12.31%), DRB1*07 (7.19% vs. 14.23%), and DRB1*11 (2.74% vs. 20.38%). The alleles DQB1*0302 and DQA1*0301, respectively, were present in the same individual in all DRB1*04 positive patients, suggesting that they belong to the haplotype. Similar situation was observed with the alleles DQB1*0201, DQA1*0501, and DRB*0301, respectively, forming the second HLA haplotype so characteristic for DM1A.

Association of HLA-DPB1 alleles with type I diabetes mellitus in Slovak population.

BACKGROUND: Genes of HLA complex on chromosome 6p21 principally contribute to the genetic risk of insulin-dependent diabetes mellitus type I (T1 DM). Associations of HLA class II loci allelic variants with T1 DM are well established. Another prime candidate, particularly the polymorphic DPB1 gene, has been reported as probably contributing to the disorder, but its relative contribution to the predisposition to the disease is difficult to assess due to strong linkage disequilibrium of HLA alleles. DPB1*0301 and DPB1*0202 have been reported as positively and DPB1*0402 as negatively associated alleles in different Caucasoid populations (predisposing versus protective alleles, respectively). OBJECTIVES: The aim of this study was to establish the occurrence rates of HLA-DPB1 alleles in patients suffering from T1 DM and to compare them with those in healthy subjects. METHODS: A PCR-SSP method was performed to identify HLA-DPB1 alleles in 61 patients and 160 healthy controls. The exact Fisher's test was used to determine the statistical significance of allele frequency differences between patients and control subjects. RESULTS: The analysis of obtained results has shown a significantly decreased frequency of DPB1*0402 and slightly increased occurrence rates of DPB1*0101 and DPB1*1301, respectively in the investigated group of patients. Neither DPB1*0301 nor DPB1*0202 were observed to be over-represented. CONCLUSIONS: The expected significant decrease in the frequency of DPB1*0402 was confirmed, whereas positive associations with DPB1*0301 and DPB1*0202, did not prove to be true, respectively (Tab. 1, Ref: 19).

HLA class II allele frequencies in type 1A diabetes mellitus Slovak patients.

BACKGROUND: Diabetes mellitus type 1A (DM-1A) is an autoimmune disease in which the immune response is directed to pancreatic islet cells. DM-1A occurs in genetically predisposed individuals. Among type 1A diabetes associated genes, those of the HLA region have the greatest effect. OBJECTIVES: The aim of our study was to obtain a comprehensive survey of the HLA-DRB1 and HLA-DQB1 allele frequencies in Slovak patients suffering from DM-1A. METHODS: HLA class II genotyping was performed on genomic DNA by the PCR-SSP method according to the 12th Workshop protocol. RESULTS: Our report gives the first presentation of the distribution of HLA-DRB1 alleles (including complete DRB1*04 subtypes) and that of HLA-DQB1 alleles in the Slovak diabetic patients diagnosed at 0-18 years of age. Susceptibility is significantly associated with the alleles DQB1*0302 (OR = 7.8), DRB1*04 (OR = 4.9), DRB1*0301 (OR = 4.2) and DQB1*02 (OR = 2.2), whereas the alleles DQB*0602 (OR = 0.05), DRB1*11 (OR = 0.2), DRB1*15 (OR = 0.2) and DQB1*0301 (OR = 0.3) were found to be protective. CONCLUSIONS: Our results, consistent with other studies, show increased frequencies of known positively associated HLA class II alleles in our type 1A diabetes mellitus patients compared to the general (nondiabetic) population. The protective effect of previously reported alleles was confirmed as well. Results of our population-based study serve in clinical practice for the identification of subjects at risk of developing DM-1A among the first-degree relatives (Tab. 2, Ref. 12).

Relative opioid efficacy is determined by the complements of the G protein-coupled receptor desensitization machinery.

G protein-coupled receptor regulation by G protein-coupled receptor kinases and beta-arrestins can lead to desensitization and subsequent internalization of the receptor. In in vitro and cellular systems, beta-arrestins do not seem to play a major role in regulating micro opioid receptor (microOR) responsiveness. Removal of the betaarrestin2 (betaarr2) gene in mice leads paradoxically to enhanced and prolonged microOR-mediated antinociception. The betaarr2 knockout (betaarr2-KO) mice also fail to develop morphine antinociceptive tolerance in the hot-plate test, further indicating that the betaarr2 protein plays an essential role in microOR regulation in vivo. In this study, the contribution of betaarr2 to the regulation of the microOR was examined in both human embryonic kidney 293 cells and in betaarr2-KO mice after treatment with several opiate agonists. A green fluorescent protein tagged betaarr2 was used to assess receptor-betaarr2 interactions in living cells. Opiate agonists that induced robust betaarr2-green fluorescent protein translocation produced similar analgesia profiles in wild-type and betaarr2-KO mice, whereas those that do not promote robust betaarr2 recruitment, such as morphine and heroin, produce enhanced analgesia in vivo. In this report, we present a rationale to explain the seemingly paradoxical relationship between beta-arrestins and microOR regulation wherein morphine-like agonists fail to promote efficient internalization and resensitization of the receptor.

Clinically unapparent central motor pathways lesion in patients with type I diabetes mellitus. A transcranial magnetic stimulation study.

OBJECTIVE: There is an evidence of central nervous system (CNS) involvement in diabetic patients. The aim of the study was to determine a conduction slowing in CNS pathways using a transcranial magnetic stimulation (TMS) and F-wave latency measurement. METHODS: Diabetic patients and a control group, both without clinical symptoms and signs of CNS lesion were evaluated. Motor evoked potentials were recorded from upper and lower extremities and central conduction time (CCT) was calculated according to formula: CCT = MEP-[0.5x(F-M-1)+M]. Obtained results and data from literature were compared. RESULTS: There was a significant prolongation of CCT recorded from lower extremities. The prolongation of CCT recorded from upper extremities was not statistically significant. Our results correlate with previously published data. CONCLUSION: In spite of missing clinical signs of CNS lesion in diabetic patients, a significant prolongation of CCT compared to control group and literature data was recorded. We assume a presence of diffuse subclinical CNS lesion induced by metabolic changes in DM. Difference between CCT obtained from upper and lower extremities implicate, that changes are analogical to peripheral neuropathies ("central length-dependent injury?"). Measurement of CCT using TMS could become a complementary electrophysiological method for assessment of subclinical CNS involvement in diabetic patients. (Tab. 4, Ref. 17.)

Differential regulation of the dopamine D2 and D3 receptors by G protein-coupled receptor kinases and beta-arrestins.

The D(2) and D(3) receptors (D(2)R and D(3)R), which are potential targets for antipsychotic drugs, have a similar structural architecture and signaling pathway. Furthermore, in some brain regions they are expressed in the same cells, suggesting that differences between the two receptors might lie in other properties such as their regulation. In this study we investigated, using COS-7 and HEK-293 cells, the mechanism underlying the intracellular trafficking of the D(2)R and D(3)R. Activation of D(2)R caused G protein-coupled receptor kinase-dependent receptor phosphorylation, a robust translocation of beta-arrestin to the cell membrane, and profound receptor internalization. The internalization of the D(2)R was dynamin-dependent, suggesting that a clathrin-coated endocytic pathway is involved. In addition, the D(2)R, upon agonist-mediated internalization, localized to intracellular compartments distinct from those utilized by the beta(2)-adrenergic receptor. However, in the case of the D(3)R, only subtle agonist-mediated receptor phosphorylation, beta-arrestin translocation to the plasma membrane, and receptor internalization were observed. Interchange of the second and third intracellular loops of the D(2)R and D(3)R reversed their phenotypes, implicating these regions in the regulatory properties of the two receptors. Our studies thus indicate that functional distinctions between the D(2)R and D(3)R may be found in their desensitization and cellular trafficking properties. The differences in their regulatory properties suggest that they have distinct physiological roles in the brain.