Comparison of the Effects of Methadone and Heroin on Human <Emphasis Type="Italic">ether</Emphasis>-<Emphasis Type="Italic">à</Emphasis>-<Emphasis Type="Italic">go</Emphasis>-<Emphasis Type="Italic">go</Emphasis>-Related Gene Channels

Torsades de pointes (TdP) is a life-threatening form of ventricular arrhythmia that occurs under conditions of delayed cardiac repolarization indicated by prolonged QT intervals in ECG recordings. The main mechanism of QT prolongation and TdP is block of the rapid component of the cardiac delayed rectifier K⁺ current (I_Kr), which is encoded by hERG (human ether-à-go-go-related gene). The opioid agonist methadone has previously been demonstrated to inhibit hERG currents, and there are reports of serious cardiac arrhythmias and deaths from TdP and ventricular fibrillation in patients taking methadone. The aim of the present study was to compare the effects of the opioid agonists methadone and heroin (3,6-diacetylmorphine) on hERG currents stably expressed in human embryonic kidney (HEK 293) cells using the whole-cell configuration of the patch-clamp technique. Both methadone and heroin inhibit hERG currents in a concentration-dependent manner. The following values were calculated for IC₅₀ (concentration causing half-maximal inhibition) and n (the Hill coefficient): 4.8 μM and 0.9 for methadone, 427 μM and 0.7 for heroin. In conclusion, the potency for block of hERG currents is about 100-fold lower for heroin when compared to methadone.     

In situ validation of VEGFR-2 and <Emphasis Type="Italic">α</Emphasis><Subscript><Emphasis Type="Italic">v</Emphasis></Subscript><Emphasis Type="Italic">ß</Emphasis><Subscript>3</Subscript> integrin as targets for breast lesion characterization

Vascular endothelial growth factor receptor 2 (VEGFR-2) and α _v ß ₃ integrin are the most frequently addressed targets in molecular imaging of tumor angiogenesis. In preclinical studies, molecular imaging of angiogenesis has shown potential to detect and differentiate benign and malignant lesions of the breast. Thus, in this retrospective clinical study employing patient tissues, the diagnostic value of VEGFR-2, α _v ß ₃ integrin and vascular area fraction for the diagnosis and differentiation of breast neoplasia was evaluated. To this end, tissue sections of breast cancer (n = 40), pre-invasive ductal carcinoma in situ (DCIS; n = 8), fibroadenoma (n = 40), radial scar (n = 6) and normal breast tissue (n = 40) were used to quantify (1) endothelial VEGFR-2, (2) endothelial α _v ß ₃ integrin and (3) total α _v ß ₃ integrin expression, as well as (4) the vascular area fraction. Sensitivity and specificity to differentiate benign from malignant lesions were calculated for each marker by receiver operating characteristics (ROC) analyses. Whereas vessel density, as commonly used, did not significantly differ between benign and malignant lesions (AUROC: 0.54), VEGFR-2 and α _v ß ₃ integrin levels were gradually up-regulated in carcinoma versus fibroadenoma versus healthy tissue. The highest diagnostic accuracy for differentiating carcinoma from fibroadenoma was found for total α _v ß ₃ integrin expression (AUROC: 0.76), followed by VEGFR-2 (AUROC: 0.71) and endothelial α _v ß ₃ integrin expression (AUROC: 0.68). In conclusion, total α _v ß ₃ integrin expression is the best discriminator between breast cancer, fibroadenoma and normal breast tissue. With respect to vascular targeting and molecular imaging of angiogenesis, endothelial VEGFR-2 appeared to be slightly superior to endothelial α _v ß ₃ for differentiating benign from cancerous lesions.     

Levels of (1→3)-β-D-glucan, <Emphasis Type="Italic">Candida</Emphasis> mannan and <Emphasis Type="Italic">Candida</Emphasis> DNA in serum samples of pediatric cancer patients colonized with <Emphasis Type="Italic">Candida</Emphasis>species

Background  

Surveillance cultures may be helpful in identifying patients at increased risk of developing invasive candidiasis. However, only scant information exists on the effect of Candida colonization on serum levels of diagnostic biomarkers. This prospective surveillance study determined the extent of Candida colonization among pediatric cancer patients and its possible impact on serum levels of (1-3)-β-D-glucan (BDG), Candida mannan and Candida DNA.     

Longevity-modulating effects of symbiosis: insights from <Emphasis Type="Italic">Drosophila</Emphasis>–<Emphasis Type="Italic">Wolbachia</Emphasis> interaction

Microbial communities are known to significantly affect various fitness components and survival of their insect hosts, including Drosophila. The composition of symbiotic microbiota has been shown to change with the host’s aging. It is unclear whether these changes are caused by the aging process or, vice versa, they affect the host’s aging and longevity. Recent findings indicate that fitness and lifespan of Drosophila are affected by endosymbiotic bacteria Wolbachia. These effects, however, are inconsistent and have been reported both to extend and shorten longevity. The main molecular pathways underlying the lifespan-modulating effects of Wolbachia remain unclear, however insulin/insulin-like growth factor, immune deficiency, ecdysteroid synthesis and signaling and c-Jun N-terminal kinase pathways as well as heat shock protein synthesis and autophagy have been proposed to play a role. Here we revise the current evidence that elucidates the impact of Wolbachia endosymbionts on the aging processes in Drosophila.     

<Emphasis Type="Italic">Oochoristica koubeki</Emphasis> n. sp. (Cestoda,Anoplocephalidae) from African <Emphasis Type="Italic">Chamaeleo senegalensis</Emphasis> (Chamaeleonidae) and emendation of the genus <Emphasis Type="Italic">Oochoristica</Emphasis> Lühe, 1898

During a survey of species diversity and ecology of selected West African vertebrates in the Niokolo Koba National Park, Senegal, two Chamaeleo senegalensis Daudin, 1802 (Chamaeleonidae) were collected and examined for parasites. A new species, Oochoristica koubeki n. sp., anoplocephalidian cestode of the genus Oochoristica Lühe, 1898 is described from the intestine of this host. The new species is related to O. theileri Fuhrmann, 1924; O. theileri Fuhrmann, 1924 f. major Baer, 1933; O. celebensis Yamaguti, 1954 and O. rostellata Zschokke, 1905 var. agamicolla Dollfus, 1957. All the forenamed species share characteristic strobila with proglottids wider than long. Oochoristica koubeki n. sp. differs from these species in different proportions of scolex, cirrus sac, ovarium and vitellarium, further differences are in number of testes and presence of reinforced genital atrium. Only O. theileri shares two last mentioned features, however it differs in shape of testes (rounded) and in distribution of testes (two clusters). The new species differs from all other species in extremely high number of egg’s capsules, different host species and zoogeographic distribution. Emendation of genus diagnosis sensu Beveridge (1994) is provided.     

Genetic epidemiology of MODY in the Czech republic: new mutations in the MODY genes <Emphasis Type="Italic">HNF-4α</Emphasis>, <Emphasis Type="Italic">GCK</Emphasis> and <Emphasis Type="Italic">HNF-1α</Emphasis>

AIMS/HYPOTHESIS: The aim of this study was to examine the prevalence and nature of mutations in HNF4alpha/MODY1, GCK/MODY2 and HNF-1alpha/MODY3 genes in Czech subjects with clinical diagnosis of MODY. METHODS: We studied 61 unrelated index probands of Czech origin (28 males, 33 females) with a clinical diagnosis of MODY and 202 family members. The mean age of probands was 22.7+/-12.0 years (range, 6-62) and the mean age at the first recognition of hyperglycaemia was 14.7+/-6.0 years (range, 1-25). The promotor and coding regions inclusive intron exon boundaries of the HNF-4alpha, GCK and HNF-1alpha genes were examined by PCR-dHPLC (HNF-1alpha and GCK) and direct sequencing. RESULTS: We identified 20 different mutations in the HNF-4alpha, GCK and HNF-1alpha in 29 families (48% of all families studied), giving a relative prevalence of 5% of MODY1, 31% of MODY2 and 11.5% of MODY3 among the Czech kindred with MODY. Three of 3, 10 of 11 and 1 of 6 of the mutations identified in HNF-4alpha, GCK and HNF-1alpha respectively, were new. CONCLUSION/INTERPRETATION: Of the families 48% carried mutations in the MODY1-3 genes and of the identified mutations 70% were new. In 52% of Czech families with clinical characteristics of MODY, no mutations were found in the analysed genes. This finding shows that the majority of MODY mutations in a central European population are local and that other MODY genes could be responsible for autosomal dominant transmission of diabetes mellitus.     

<Emphasis Type="Italic">Mycobacterium bovis</Emphasis> BCG-itis and Cervical Lymphadenitis due to <Emphasis Type="Italic">Salmonella enteritidis</Emphasis> in a Patient with Complete Interleukin-12/-23 Receptor <Emphasis Type="Italic">β</Emphasis>1 Deficiency

Mendelian susceptibility to mycobacterial disease (MSMD) is a rare disorder with predisposition to severe, sometimes lethal, disease caused by otherwise poorly virulent, non-tuberculous environmental mycobacteria and poorly virulent salmonellae. In patients with MSMD, mutations have been identified in five genes that encode for the proteins IL-12/IL-23p40, IL-12/ IL-23Rβ1, IFN-コR1, IFN-γR2 and STAT1. These proteins play important roles in the type-1 cytokine pathway, which is crucial for human host defence against intracellular pathogens such as mycobacteria and salmonellae. We report a girl with mild Mycobacterium bovis Bacille Calmette–Guérin (BCG) disease and Salmonella enteritidis cervical lymphadenitis. Despite treatment, she has remained a fecal carrier of S. enteritidis for the past 14 years. She was found to have complete IL-12/IL-23Rβ1 deficiency. A homozygous r.518G>C IL12RB1 mutation was identified, leading to a non-functional R173P substitution in the IL-12/IL-23Rβ1 protein. This mutation abrogated IL-12/IL-23Rβ1 cell-surface expression and resulted in complete lack of T cell responsiveness to both IL-12 and IL-23.     

Emerging causes of traveler’s diarrhea: <Emphasis Type="Italic">Cryptosporidium,Cyclospora, Isospora</Emphasis>, and <Emphasis Type="Italic">Microsporidia</Emphasis>

Travel is a risk factor for acquiring infection with a sporeforming protozoa: Cryptosopridium, Cyclospora, Microsporidia, and Isospora. Certain travel destinations have a high disease burden and intense exposure. Patients present with persistent diarrhea and a history of recent travel to a developing country in the tropics. Very mild infections may be underdiagnosed and may cause typical traveler’s diarrhea. In a patient with a history of travel and persistent diarrhea unresponsive to the usual antibiotic and antidiarrhea treatment, stool studies for all four of these protozoa infections should be performed. If immune status is normal and the disease is mild, symptomatic therapy may suffice. Effective treatment is available for Cyclospora, Microsporidia, and Isospora.     

Does Cyclophosphamide Play a Protective Role Against Neuronal Loss in Chronic <Emphasis Type="Italic">T.</Emphasis><Emphasis Type="Italic"> cruzi</Emphasis> Infection?

In this study, we verified the possible role of cyclophosphamide (CY) in protecting or not against neuronal losses in young and aged male Calomys callosus chronically infected with the MORC-1 strain of Trypanosoma cruzi through numerical quantification of neurons from the myenteric plexus of the colon and quantification of nitric-oxide concentration (NO) during the acute and chronic phase of infection. For this purpose, groups of young C. callosus were infected with the MORC-1 strain of T. cruzi. A group of infected animals received i.p. 0.2 mg/ml genuxal dissolved in distilled water treatment with CY. NO concentration in aged animals displayed reduced levels when compared to those found in young animals. No significant alterations in the number of neurons were observed in young animals, but for aged ones, a protective role of CY in reducing neuron loss was noted, in addition to enhancing the neuronal volume, area, and perimeter. These results suggest that CY administration, depending on the dose and time span, can act as a protective agent against neuronal losses.     

Post-translational protein modifications in type 1 diabetes: a role for the repair enzyme protein-<Emphasis Type="SmallCaps">l</Emphasis>-isoaspartate (<Emphasis Type="SmallCaps">d</Emphasis>-aspartate) <Emphasis Type="Italic">O</Emphasis>-methyltransferase?

Aims/hypothesis

Post-translational modifications, such as isomerisation of native proteins, may create new antigenic epitopes and play a role in the development of the autoimmune response. Protein-l-isoaspartate (d-aspartate) O-methyltransferase (PIMT), encoded by the gene PCMT1, is an enzyme that recognises and repairs isomerised Asn and Asp residues in proteins. The aim of this study was to assess the role of PIMT in the development of type 1 diabetes.

Materials and methods

Immunohistochemical analysis of 59 normal human tissues was performed with a monoclonal PIMT antibody. CGP3466B, which induces expression of Pcmt1, was tested on MIN6 and INS1 cells, to assess its effect on Pcmt1 mRNA and PIMT levels (RT-PCR and western blot) and apoptosis. Forty-five diabetes-prone BioBreeding (BB) Ottawa Karlsburg (OK) rats were randomised to receive 0, 14 or 500 μg/kg (denoted as the control, low-dose and high-dose group, respectively) of CGP3466B from week 5 to week 20.

Results

A high level of PIMT protein was detected in beta cells. CGP3466B induced a two- to threefold increase in Pcmt1 mRNA levels and reduced apoptosis by 10% in MIN6 cells. No significant effect was seen on cytokine-induced apoptosis or PIMT protein levels in INS1 cells. The onset of diabetes in the BB/OK rats was significantly delayed (85.6?±?9.0 vs 84.3?±?6.8 vs 106.6?±?13.5 days, respectively; p?p?p?

Conclusions/interpretation

The results support a role for post-translational modifications and PIMT in the development of type 1 diabetes in the diabetes-prone BB rat, and perhaps also in humans.

     

Association between <Emphasis Type="Italic">IL17A</Emphasis> and <Emphasis Type="Italic">IL17F</Emphasis> polymorphisms and risk of Henoch–Schonlein purpura in Chinese children

Previous studies suggested that interleukin-17 and Th17 cell play an important role in the pathogenesis of childhood Henoch–Schonlein purpura (HSP). The purpose of our study is to elucidate whether the IL17A and IL17F gene polymorphisms are susceptibility genes for the development of HSP in Chinese children. A total of 148 HSP patients and 202 controls were enrolled for analyzing the single nucleotide polymorphisms (SNP) of IL17A (rs2275913, rs8193037 and rs3819025) and IL17F (rs763780 and rs9463772). TaqMan Real-Time polymerase chain reaction method was used in SNP genotyping. Compared to the healthy controls, the IL17A rs2275913 variant allele A showed a significant association with HSP [odds ratio (OR) 0.70; 95 % CI 0.51–0.94, P = 0.018]. Genotyping analysis demonstrated rs2275913 was associated with a decreased HSP risk (G/A vs. G/G: OR 0.56; 95 % CI 0.33–0.95; A/A vs. G/G: OR 0.46; 95 % CI 0.24–0.86; P = 0.032). Also, our findings showed that the A allele of IL17A rs3819025 was associated with a higher risk of HSP nephritis (OR 1.61; 95 % CI 1.00–2.58; P = 0.047). In addition, a risk haplotype of IL17A (GGA) was found (OR 1.84; 95 % CI 1.17–2.88; P = 0.008). However, no significant differences between HSP patients and healthy controls were observed when comparing genotype, allele or haplotype frequencies of the IL17F rs763780 and rs9463772 polymorphisms. In this study, we confirmed that the rs2275913 polymorphism of the IL17A gene was associated with susceptibility to HSP in Chinese children. However, there was no relationship between IL17F rs763780 and rs9463772 polymorphisms and HSP susceptibility.     

Antimicrobial resistance in <Emphasis Type="Italic">Enterobacteriaceae</Emphasis> from healthy broilers in Egypt: emergence of colistin-resistant and extended-spectrum β-lactamase-producing <Emphasis Type="Italic">Escherichia coli</Emphasis>

Background

Poultry remains one of the most important reservoir for zoonotic multidrug resistant pathogens. The global rise of antimicrobial resistance in Gram-negative bacteria is of reasonable concern and demands intensified surveillance.

Methods

In 2016, 576 cloacal swabs were collected from 48 broiler farms located in five governorates in northern Egypt. Isolates of Enterobacteriaceae could be cultivated on different media and were identified by MALDI-TOF MS and PCR. Escherichia coli isolates were genotyped by DNA-microarray-based assays. The antimicrobial susceptibility to 14 antibiotics was determined and resistance-associated genes were detected. The VITEK-2 system was applied for phenotypical confirmation of extended-spectrum β-lactamase-producing isolates. The determination of colistin resistance was carried out phenotypically using E-test and genotypically using PCR for detection of the mcr-1 gene.

Results

Out of 576 samples, 72 representatives of Enterobacteriaceae were isolated and identified as 63 E. coli (87.5%), 5 Enterobacter cloacae (6.9%), 2 Klebsiella pneumoniae (2.8%) and 2 Citrobacter spp. (2.8%). Seven out of 56 cultivated E. coli (12.5%) were confirmed as ESBL-producing E. coli and one isolate (1.8%) as ESBL/carbapenemase-producing E. coli. Five out of 63 E. coli isolates (7.9%) recovered from different poultry flocks were phenotypically resistant to colistin and harboured mcr-1 gene.

Conclusions

This is the first study reporting colistin resistance and emergence of multidrug resistance in Enterobacteriaceae isolated from healthy broilers in the Nile Delta region, Egypt. Colistin-resistant E. coli in poultry is of public health significance. The global rise of ESBL- and carbapenemase-producing Gram-negative bacteria demands intensified surveillance. ESBL-producing E. coli in poultry farms in Egypt are of major concern that emphasizes the possibility of spread of such strains to humans. The results also reinforce the need to develop strategies and to implement specific control procedures to reduce the use of antibiotics.

     

Assessing gene–treatment interactions at the <Emphasis Type="Italic">FTO</Emphasis> and <Emphasis Type="Italic">INSIG2</Emphasis> loci on obesity-related traits in the Diabetes Prevention Program

Aims/hypothesis  The single nucleotide polymorphism (SNP) rs9939609 in the fat mass and obesity associated gene (FTO) and the rs7566605 SNP located 10 kb upstream of the insulin-induced gene 2 gene (INSIG2) have been proposed as risk factors for common obesity. Methods  We tested for genotype–treatment interactions on changes in obesity-related traits in the Diabetes Prevention Program (DPP). The DPP is a randomised controlled trial of 3,548 high-risk individuals from 27 participating centres throughout the USA who were originally randomised to receive metformin, troglitazone, intensive lifestyle modification or placebo to prevent the development of type 2 diabetes. Measures of adiposity from computed tomography were available in a subsample (n = 908). This report focuses on the baseline and 1 year results. Results  The minor A allele at FTO rs9939609 was positively associated with baseline BMI (p = 0.003), but not with baseline adiposity or the change at 1 year in any anthropometric trait. For the INSIG2 rs7566605 genotype, the minor C allele was associated with more subcutaneous adiposity (second and third lumbar vertebrae [L2/3]) at baseline (p = 0.04). During follow-up, CC homozygotes lost more weight than G allele carriers (p = 0.009). In an additive model, we observed nominally significant gene–lifestyle interactions on weight change (p = 0.02) and subcutaneous (L2/3 [p = 0.01] and L4/5 [p = 0.03]) and visceral (L2/3 [p = 0.02]) adipose areas. No statistical evidence of association with physical activity energy expenditure or energy intake was observed for either genotype. Conclusions/interpretation  Within the DPP study population, common variants in FTO and INSIG2 are nominally associated with quantitative measures of obesity, directly and possibly by interacting with metformin or lifestyle intervention. Trial registration: ClinicalTrials.gov NCT00004992 Funding: The study was funded by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), the National Institute on Aging (NIA), the National Institute of Child Health and Human Development (NICHD), the National Center on Minority Health and Health Disparities (NCMHD) and the Office of Research on Women’s Health (ORWH). Electronic supplementary material  The online version of this article (doi:) contains supplementary material, including a list of members of the Diabetes Prevention Program Research Group, which is available to authorised users. An erratum to this article can be found at     

Analysis of the 3′ Variable Region of the <Emphasis Type="Italic">cagA</Emphasis> Gene of <Emphasis Type="Italic">Helicobacter pylori</Emphasis> Isolated in Koreans

CagA protein of Helicobacter pylori is injected into epithelial cells, and it undergoes tyrosine phosphorylation, resulting in inducing cytoskeletal rearrangements. A few studies have suggested that the number of CagA tyrosine phosphorylation motifs (EPIYA) and subtypes of CagA were associated with gastric cancer. This study was performed to characterize the 3' variable regions of the cagA gene of H. pylori and to investigate whether or not there is any relationship between the diversities of cagA and the disease outcome in Korea. Seventy-nine patients (chronic gastritis, 15; duodenal ulcer, 27; benign gastric ulcer, 18; gastric cancer, 19) were enrolled. Biopsy specimens were taken from the antrum for H. pylori culture, and genomic DNA was extracted. PCR and DNA sequence analysis was carried out for the 3′ variable region of the cagA gene. Seventy-eight strains (98.8%) contained three EPIYA motifs and one strain (1.2%) isolated from a patient with duodenal ulcer contained four EPIYA motifs. Seventy-six strains (96.2%) were the East Asian type. In conclusion, there was no significant difference between the number of EPIYA motifs or CagA subtypes and various gastroduodenal diseases in Korea.     

<Emphasis Type="Italic">PAX3/7</Emphasis>–<Emphasis Type="Italic">FOXO1</Emphasis> fusion status in older rhabdomyosarcoma patient population by fluorescent in situ hybridization

Purpose  

In pediatric alveolar rhabdomyosarcoma, the PAX3–FOXO1 and PAX7–FOXO1 gene fusions are prognostic indicators, while little is known concerning this disease in older patients. To determine whether PAX3/7–FOXO1 fusion gene status correlates with outcome in adolescent, young adult, and adult rhabdomyosarcoma patients, the histological, immunohistochemical, and clinical characteristics of 105 patients followed at The University of Texas MD Anderson Cancer Center from 1957 to 2001 were evaluated.     

Hypertension Due to Loss of Clock: Novel Insight From the Molecular Analysis of <Emphasis Type="Italic">Cry1</Emphasis>/<Emphasis Type="Italic">Cry2</Emphasis>–Deleted Mice

In our consumer-oriented society, in which productivity requires around-the-clock activity and demanding shift work, the biologic system that regulates our internal rhythms is being compromised. Poor sleep patterns and hectic lifestyle are detrimental to harmonious physiological and metabolic body systems, with severe impact on public health. Over a trillion peripheral cellular clocks throughout the body, supervised by the master clock located in the hypothalamic suprachiasmatic nucleus, govern most aspects of physiology and behavior. To exemplify the importance of the biologic clock for health, we have recently demonstrated that mice that are arrhythmic because of the deletion of Cry1 and Cry2 clock genes suffer from salt-sensitive hypertension. In these mice, a novel 3β-hydroxyl-steroid dehydrogenase (3β–Hsd) gene under clock control is severely overexpressed specifically in aldosterone-producing cells in the adrenal cortex, leading to hyperaldosteronism and ultimately to salt-sensitive hypertension. The human homologue of this aldosterone-producing, cell-specific enzyme was also characterized and represents a new possibility in the pathogenesis of hypertension.     

Are <Emphasis Type="Italic">Thymidylate synthase</Emphasis> and <Emphasis Type="Italic">Methylene tetrahydrofolate reductase</Emphasis> genes linked with methotrexate response (efficacy,toxicity) in Indian (Asian) rheumatoid arthritis patients?

Methotrexate (MTX) is among the best-tolerated disease-modifying antirheumatic drugs for the treatment of rheumatoid arthritis (RA); major drawbacks of MTX therapy are the large interpatient variability in clinical response and the unpredictable appearance of a large spectrum of side effects. Several studies have demonstrated gene polymorphism that may regulate intracellular methotrexate metabolic pathway enzymes linked to drug efficacy and safety, but the evidence available is not yet conclusive. We decided to run a pilot study to determine the incidence of Methylene tetrahydrofolate (MTHFR; C677T, A1298C) and Thymidylate synthase (TS; 5′ UTR repeat, 3′ UTR deletion) gene polymorphism in rheumatoid arthritis patients in our community (Indian Asian) and further explore its association with MTX response (efficacy, toxicity). Thirty-four naïve RA patients on supervised MTX therapy and 139 healthy controls were genotyped for A1298C and C677T polymorphism of the MTHFR gene and 5′ UTR repeat and 3′ UTR deletion polymorphism of the TYMS gene by polymerase chain reaction-restriction fragment length polymorphism. Association, if any, between gene polymorphism and MTX response in RA patients was analyzed. The MTHFR A1298C ‘C’ allele incidence among RA patients (46%) was significantly higher (χ ²?=?4.24, P?相似文献