Appropriate BMI cut-off values for identification of metabolic risk factors: Third national surveillance of risk factors of non-communicable diseases in Iran (SuRFNCD-2007)

Aim: To determine the appropriate threshold of body mass index (BMI) associated with increased risk of cardiovascular diseases in a large representative sample of an Iranian population.

Subjects and methods: Data of third national surveillance of risk factors of non-communicable diseases (SuRFNCD-2007) were used in this study. Sensitivity, specificity, and shortest distance on the receiver-operating characteristic (ROC) curves were used to determine gender-specific optimal cut-offs of BMI for cardiometabolic risk factors including elevated blood pressure, low high-density lipoprotein cholesterol, high triglycerides, high fasting plasma glucose and for ≥ 2 of the aforementioned risk factors.

Results: There was a continuous increase in the prevalence of cardiometabolic risk factors with increasing BMI (p < 0.001). At the BMI of 25–29 kg/m² men were at higher risk of cardiovascular diseases compared to women (p < 0.001). The appropriate BMI cut-offs ranged from 24.6–26.1 kg/m² for men and from 26.9–28.8 kg/m² for women. The optimal BMI cut-offs for identifying any two or more of those risk factors were 25.2 and 27.3 kg/m² in men and women, respectively.

Conclusion: In men the appropriate BMI cut-offs are ～25 kg/m², while in women higher BMI values are associated with risk of cardiovascular diseases.     

Fluoro-immuno-cytoadherence (FICA): A new method for the identification and enumeration of antigen-binding cells.

Antigen-coated particles of cross-linked dextran may be used for affinity chromatography of antibodies and for the fractionation of lymphoid cells with appropriate surface receptors. Furthermore, such particles serve as convenient substrates for quantitative immunofluorescence tests. The fluoro-immuno-cyto-adherence (FICA) is a simple technique which combines affinity chromatography and immunofluorescence, provides durable antigen-coated substrates and allows the identification, enumeration and characterization of lymphoid cells capable of binding an antigen, covalently linked via a spacer onto the surface of dextran beads. In the present study chicken thyroglobulin (TG) or bovine serum albumin (BSA) were coupled onto fluorescein and rhodamin-labelled or unlabelled Sephadex G-25 beads by means of spacer molecules. The specificity and degree of antigen-coating were controlled by indirect immunoflourescence. For the study of antigen-binding cells the different antigen-coated beads were mixed with suspensions of peripheral blood lymphoid cells from Obese strain (OS) chickens with spontaneous hereditary autoimmune thyroiditis, or with cells from BSA-immunized or unimmunized normal White Leghron chickens. Specific adherence of OS lymphocytes to TG-coated beads and of lymphocytes from BSA-immunized chickens to BSA-beads was found. The test and control preparations are observed simultaneously under the fluorescence microscope where the distinction of beads coated with different antigens can be made on the basis of the color of their fluorescence. Results obtained with the FICA technique are in good agreement with those of conventional rosette tests.     

In vitro evidence for bakuchiol's influence towards drug metabolism through inhibition of UDP-glucuronosyltransferase (UGT) 2B7

Background

Inhibition of drug-metabolizing enzymes (DMEs) has been regarded as one of the most important reason for clinical drug-drug interaction.

Aim

The aim of the present study is to evaluate the inhibition of bakuchiol towards UDP-glucuronosyltransferase (UGT) 2B isoforms.

Methods

In vitro recombinant UGT2B-catalyzed 4-methylumbelliferone glucuronidation was used as the probe reaction. Dixon plot and Lineweaver-Burk plot were employed to determine the inhibition kinetic type, and nonlinear regression of data was utilized to calculate the inhibition kinetic parameter (Ki). In vitro-in vivo extrapolation (IVIVE) was carried out to predict in vivo inhibition magnitude.

Results

Among the tested UGT2B isoforms, UGT2B7 was inhibited by the strongest intensity. The noncompetitive inhibition was demonstrated by the results obtained from Dixon plot and Lineweaver-Burk plot. The K_i value was calculated to be 10.7 µM. In combination with the reported concentration after an intravenous administration of bakuchiol (15 mg/kg) in rats, the high risk of in vivo inhibition of bakuchiol towards UGT2B7-catalyzed metabolism of drugs was indicated.

Conclusion

All these results provide an important information for the risk evaluation of the clinical utilization of bakuchiol.     

Lack of satiety effect of cholecystokinin (CCK) in a new rat model not expressing the CCK-A receptor gene

This work expands recent observations that Otsuka Long-Evans Tokushima Fatty (OLETF) rats show little or no pancreatic expression of the cholecystokinin (CCK)-A receptor gene. We examined whether the CCK-A and -B receptor genes were expressed in the brain (hypothalamus) of OLETF rats in comparison with control (Long-Evans Tokushima Otsuka = LET) rats. CCK-A receptor mRNA was detected in the hypothalamus of LETO rats but not OLETF rats. The CCK-B receptor gene was expressed in the hypothalamus in both strains. Cerebroventricular administration of CCK-8 sulfate inhibited daily food intake in LETO rats, but not in OLETF rats. These results show that in OLETF rats the absence of CCK-A receptor gene expression in the hypothalamus results in hyperphagia because of lack of satiety.     

人CD226 (PTA1)分子胞膜外区截短体真核表达载体的构建、表达及其识别结构域的分析

目的：确定一套CD226单克隆抗体（McAb）识别CD226分子胞膜外区结构域的部位，方法：应用计算机软件分析CD226子蛋白质序列疏水性，确定其亲水性区域。采用PCR方法扩增CD226分子基因序列，分别缺失相应亲水性区域，构建CD226截短体重组真核组细胞表达载体pcDNA3-PTA1T1和pcDNA3-PTA1T2。测序正确后，转染COS7细胞，抗CD226分子克隆抗体间接免疫荧光染色，流式细胞术检测CD226分子截短体的表达，表达成功后，采用间接免疫荧光染色和流式细胞术分析，用一套CD226McAb检测与截短突变体的免疫反应性，从而确定CD226McAb识别CD226分子结构域的部位。结果：PCR扩增出CD226分子相应目的片段序列，定向克隆入pcDNA3真核表达。CD226McAb Leo A1,New E1T -7均可识别CD226全长分子；Leo A1,New E1,FMU1,FMU2,FMU4和MFU5与截短突变体PTA1T1和PTA1T2均无反应性，FMU3可结合PTA1T1分子，不结合PTA1T2分子；FMU6和FMU7均可结合PTA1T2均无反应性；FMU3可结合PTA1T1分子，不结合PTA1T2分子；FMU6和FMU7均可结合PTA1T1及PTA1T2分子，结论：CD226McAb Leo A1,New E1,FMU1,FMU2,FMU3,FMU4和FMU5识别CD226分子膜膜外区D1结构域，其中FMU3识别的位点位于V1和V2环之间；FMU6和FMU7识别D2结构域。     

Comparative sequence analysis (CSA): a new sequence-based method for the identification and characterization of mutations in DNA

Direct sequencing analysis is largely used to confirm and characterize mutations previously detected by more rapid tests. We have developed a method-Comparative Sequence Analysis (CSA)-that simplifies the analysis of sequencing data facilitating its use as a first screen for mutation detection. Sequence data were split into their component electrophoretograms and the use of a size standard enabled equivalent traces from different individuals to be overlaid. This allowed simple and rapid visual analysis of the results. Using this technique in a blind study, we tested 576 samples for mutations in the Von Hippel-Lindau tumor suppressor gene, VHL. We were able to identify and characterize all 78 known mutations present within the sample set (100% sensitivity and specificity).     

Comparative evaluation of the new Titertek Enterobac Rapid Automated System (TTE-RAS) for identification of members of the family Enterobacteriaceae.

The Titertek Enterobac Rapid Automated System (TTE-RAS; Flow Laboratories, SpA, Milan, Italy), a new semiautomated system for the identification of members of the family Enterobacteriaceae, was compared with the API 20E system (API System P.A., Montalieu Vercieu, France) by using 284 clinically isolated strains that were previously identified by conventional methods. Six strains from the American Type Culture Collection (Rockville, Md.) were included to evaluate the reproducibility of identification by both systems. Correct identifications at the species level were 93.7% with TTE-RAS and 96.1% with API 20E. Although some of the features of the TTE-RAS data base were not satisfactory, we consider this new miniaturized system to be a very valuable tool for the rapid identification of the most frequently isolated opportunistic bacteria.     

Identifying sputum specimens of high priority for examination by enhanced mycobacterial detection, identification, and susceptibility systems (EMDISS) to promote the rapid diagnosis of infectious pulmonary tuberculosis

AIMS: To compare clinical information and sputum microscopy as methods for the selection of samples for enhanced mycobacterial detection, identification, and susceptibility systems (EMDISS) to promote the rapid diagnosis of patients with infectious pulmonary tuberculosis. METHODS: Two thousand, two hundred and sixty four specimen request forms were examined for clinical details, which were then used to identify specimens likely to yield Mycobacterium tuberculosis on culture. These results were compared with the results of sputum microscopy for acid fast bacilli (AFB). Both methods were assessed against the results of culture using a combination of continuous automated mycobacterial liquid culture (CAMLiC) and conventional solid culture. RESULTS: Classification based on clinical details was an inefficient method of identifying high priority specimens for EMDISS. Although, when given, clinical details were often consistent, a substantial proportion of specimens arrived with no details. This approach would result in the referral of at least 16% of the workload but lead to the detection by culture of only 46% of the M tuberculosis present within it. In contrast, microscopy for AFB defined a much smaller number of specimens (4.8% of the total), which contained 90% of the M tuberculosis isolates. CONCLUSIONS: Microscopy for AFB is the most efficient method for defining sputum specimens suitable for referral for enhanced mycobacteriological techniques. However, it is essential that the methods used for smear preparation and microscopy are of the highest possible standard, otherwise some patients with infectious pulmonary tuberculosis will be denied, unnecessarily, the benefits of important advances in mycobacteriology.     

宫颈癌易感基因单核苷酸多态性研究进展

宫颈癌是严重危害妇女健康的一种疾病,研究其易感基因单核苷酸多态性(SNP),对其基因诊断提供分子标记有十分重要的意义。现就近两年宫颈癌易感基因的SNP研究做一综述。     

A new genetic test for the rapid identification of shiga-toxines producing (STEC), enteropathogenic (EPEC) E. coli isolates from children

Routine bacteriological techniques do not allow detection of the most frequent enteric pathogens in young children: enteropathogenic Escherichia coli (EPEC) and shigatoxinogenic E. coli (STEC/EHEC). Since there is no correlation between serotype and pathotype, a genotypic determination is therefore necessary for the identification of these pathogenic strains. We evaluated the Genotype EHEC test (Hain Life Science, Germany), a new rapid system based on DNA multiplex amplification and further hybridization for the detection of shigatoxin stx1, stx2 genes, intimin eae gene and invasin ipaH gene harbored by Shigella and enteroinvasive E. coli (EIEC). E. coli strains of various serogroups isolated from children with acute gastroenteritis, hemorrhagic colitis or hemolytic-uremic syndrome were tested. Their genotypes were first determined by standard in-house PCR. The strains collection included 11 STEC/EHEC (serogroups O157, O111, O26, O91, O-untypable) and nine EPEC (serogroups O26, O157, O55, O126, O127, O-untypable). The same strains were tested with Genotype EHEC. For all the strains, the hybridization banding pattern obtained by Genotype EHEC correlated with their expected genotypic characteristics. No specific equipment is required, except a thermocycler. Absence of electrophoresis system, of ethidium bromide staining and imaging system is a clear-cut advantage of Genotype EHEC. In addition, the short testing time (less than 2 h) optimizes treatment orientation. The Genotype EHEC test allows an easy and reliable identification of EHEC, STEC, EPEC and also EIEC. As such, it is a useful tool for the rapid diagnosis of diarrheal diseases.     

Cellular distribution of the new growth factor Pleiotrophin (HB-GAM) mRNA in developing and adult rat tissues

Summary Pleiotrophin (PTN), also known as HBGAM, belongs to an emerging cytokine family unrelated to other growth factors. We report here the first comprehensive study using in situ hybridization on the cellular distribution of this new heparin-binding growth factor mRNA in rat tissues. PTN mRNA was developmentally expressed in many — but not all — neuroectodermal and mesodermal lineages, whilst no PTN mRNA was detected in endoderm, ectoderm and trophoblast. PTN mRNA was found in the nervous system throughout development, with a post-natal peak of expression. In the adult nervous system, significant expression persisted in hippocampal CA1 pyramidal neurons and in cortical neurons, but also in different non-neuronal cells types in various locations (olfactory nerve, cerebellar astrocytes, pituicytes, Schwann cells surrounding the neurons in sensory ganglia). PTN mRNA was also found during development in the mesenchyme of lung, gut, kidney and reproductive tract, in bone and cartilage progenitors, in dental pulp, in myoblasts, and in several other sites. Expression was differently regulated in each location, but usually faded around birth. In the adult, PTN mRNA was still present in the meninges, the iris, the Leydig cells of the testis and in the uterus. PTN mRNA was also strongly expressed in the basal layers of the tongue epithelium, which is the only epithelium and ectodermal derivative to express PTN mRNA, and this only after birth. PTN is known to be a growth factor for perinatal brain neurons and a mitogen for fibroblasts in vitro. Recently, trophic effects on epithelial cells and a role as a tumour growth factor have been reported. The mechanisms of regulation and the functions of PTN are however still uncertain. Its expression pattern during development suggests important roles in growth and differentiation. Moreover, the presence of PTN mRNA in several adult tissues and the up-regulation of PTN mRNA expression in the gravid uterus indicate that PTN also has physiological functions during adulthood.     

Comparison of a new colorimetric assay with the NCCLS broth microdilution method (M-27A) for antifungal drug MIC determination

We evaluated a new microtiter assay for antifungal susceptibility testing based on a colorimetric reaction to monitor fungal substrate utilization. This new method (rapid susceptibility assay [RSA]) provides quantitative endpoint readings in less than 8 h compared with visual determination of MIC by the National Committee for Clinical Laboratory Standards (NCCLS) broth microdilution method, which requires a minimum of 48 h of incubation. In this study, we tested clinical isolates from each of the following species: Candida albicans (20 isolates), C. glabrata (20 isolates), C. krusei (19 isolates), C. tropicalis (19 isolates), and C. parapsilosis (28 isolates). RSA and NCCLS broth dilution methods were used to determine the MICs of amphotericin B, fluconazole, itraconazole, and 5-flucytosine for all 106 isolates. RPMI 1640 medium buffered with morpholinopropanesulfonic acid was used for both methods; however, glucose and inoculum concentrations in the RSA were modified. RSA MICs were determined as the lowest drug concentration that prevented glucose consumption by the organism after 6 h of incubation. MICs obtained from the RSA were compared with those obtained from the NCCLS M-27A method read at 24 and 48 h. MIC pairs were considered in agreement when the difference between the pairs was within 2 twofold dilutions. For the 106 isolates tested, amphotericin B and 5-flucytosine demonstrated the highest agreement in MICs between the two methods (100 and 98%, respectively), whereas fluconazole and itraconazole produced less favorable MIC agreement (63.2 and 61.3%, respectively). The azole MIC differences between the two methods were significantly reduced when lower inocula were used with a prolonged incubation time. This preliminary comparison suggests that this rapid procedure may be a reliable tool for the in vitro determination of MICs of amphotericin B and 5-flucytosine and warrants further evaluation.     

Characterization of woodchuck apolipoprotein A-I: a new tool for drug delivery and identification of altered isoforms in the woodchuck chronic hepatitis model

Apolipoprotein A-I (ApoA-I) is the major protein component of high density lipoprotein (HDL) particles in serum, and participates in the reverse transport of cholesterol from tissues to the liver for excretion. The natural HDL tropism to the liver and cancer cells has been used extensively to target encapsulated drugs. The alteration of the plasmatic isoforms of ApoA-I is a hallmark of chronic hepatitis and hepatocarcinoma in mice and humans. Woodchucks infected with the woodchuck hepatitis virus (WHV) represent the best animal model for the study of chronic viral hepatitis B and viral induced hepatocarcinoma (HCC). WHV-infected woodchuck represents a clinically relevant animal model under which new treatment strategies can be evaluated and optimized. Therapeutic efficacy in this model is likely to be translated into a successful therapy for patients infected with HBV. The present study describes, for the first time, the cloning and characterization of woodchuck ApoA-I. The open reading frame (ORF) of the woodchuck ApoA-I is 795 bp long, coding for 264 amino acids. Unexpectedly, phylogenetic analysis revealed that the closest sequences are those of human and macaque. Woodchuck HDLs were isolated successfully from sera by density gradient ultracentrifugation. A commercial antibody that recognized the woodchuck ApoA-I was also identified. Finally, taking advantage of the techniques and tools developed in this study, two potential applications of woodchuck HDLs are illustrated: drug delivery to a woodchuck hepatocarcinoma cell line and the use of isoelectrofocusing to identify ApoA-I isoforms.     

A new species of the medicinal leech (Oligochaeta, Hirudinida, Hirudo) from Transcaucasia and an identification key for the genus Hirudo

A recent molecular phylogenetic study has suggested that the genus Hirudo contains a neglected species previously known as the orientalis coloration type of the medicinal leech Hirudo medicinalis. In this paper, the new species is formally described as Hirudo orientalis sp. n. It can most readily be identified by the grass green coloration of the dorsum, segmentally arranged pairs of black quadrangular or rounded dots on its paramarginal dorsal stripes and similarly arranged, but less regular light-colored markings on the predominantly black venter. It has medium-sized epididymes and an evenly coiled vagina. H. orientalis is known from Transcaucasia, Iran, and Uzbekistan. It is widely used in medicine as the “medicinal leech.” Very little is known about its exact distribution, specific habitat, and conservation status. The paper contains an identification key to all species of the genus Hirudo.     

Complementation studies in the cblA class of inborn error of cobalamin metabolism: evidence for interallelic complementation and for a new complementation class (cblH)

AIM—To investigate genetic heterogeneity within the cblA class of inborn error of cobalamin metabolism.
CONTEXT—The cblA disorder is characterised by vitamin B12 (cobalamin) responsive methylmalonic aciduria and deficient synthesis of adenosylcobalamin, required for activity of the mitochondrial enzyme methylmalonyl CoA mutase. The cblA gene has not been identified or cloned. We have previously described a patient with the clinical and biochemical phenotype of the cblA disorder whose fibroblasts complemented cells from patients with all known types of inborn error of adenosylcobalamin synthesis, including cblA.
METHODS—We have performed somatic cell complementation analysis of the cblA variant fibroblast line with a panel of 28 cblA lines. We have also performed detailed complementation analysis on a panel of 10 cblA fibroblast lines, not including the cblA variant line.
RESULTS—The cblA variant line complemented all 28 cell lines of the panel. There was evidence for interallelic complementation among the 10 cblA lines used for detailed complementation analysis; no cell line in this panel complemented all other members.
CONCLUSIONS—These results strongly suggest that the cblA variant represents a novel complementation class, which we have designated cblH and which represents a mutation at a distinct gene. They also suggest that the cblA gene encodes a protein that functions as a multimer, allowing for extensive interallelic complementation.


     

<Emphasis Type="Italic">c</Emphasis>hip <Emphasis Type="Italic">a</Emphasis>rtifact <Emphasis Type="Italic">CORRECT</Emphasis>ion (caCORRECT): A Bioinformatics System for Quality Assurance of Genomics and Proteomics Array Data

Quality assurance of high throughput "-omics" data is a major concern for biomedical discovery and translational medicine, and is considered a top priority in bioinformatics and systems biology. Here, we report a web-based bioinformatics tool called caCORRECT for chip artifact detection, analysis, and CORRECTion, which removes systematic artifactual noises that are commonly observed in microarray gene expression data. Despite the development of major databases such as GEO arrayExpress, caArray, and the SMD to manage and distribute microarray data to the public, reproducibility has been questioned in many cases, including high-profile papers and datasets. Based on both archived and synthetic data, we have designed the caCORRECT to have several advanced features: (1) to uncover significant, correctable artifacts that affect reproducibility of experiments; (2) to improve the integrity and quality of public archives by removing artifacts; (3) to provide a universal quality score to aid users in their selection of suitable microarray data; and (4) to improve the true-positive rate of biomarker selection verified by test data. These features are expected to improve the reproducibility of Microarray study. caCORRECT is freely available at: http://caCORRECT.bme.gatech.edu.     

A staging table for the embryonic development of the brownbanded bamboo shark (Chiloscyllium punctatum)

Background: Studying cartilaginous fishes (chondrichthyans) has helped us understand vertebrate evolution and diversity. However, resources such as genome sequences, embryos, and detailed staging tables are limited for species within this clade. To overcome these limitations, we have focused on a species, the brownbanded bamboo shark (Chiloscyllium punctatum), which is a relatively common aquarium species that lays eggs continuously throughout the year. In addition, because of its relatively small genome size, this species is promising for molecular studies. Results: To enhance biological studies of cartilaginous fishes, we establish a normal staging table for the embryonic development of the brownbanded bamboo shark. Bamboo shark embryos take around 118 days to reach the hatching period at 25°C, which is approximately 1.5 times as fast as the small‐spotted catshark (Scyliorhinus canicula) takes. Our staging table divides the embryonic period into 38 stages. Furthermore, we found culture conditions that allow early embryos to grow in partially opened egg cases. Conclusions: In addition to the embryonic staging table, we show that bamboo shark embryos exhibit relatively fast embryonic growth and are amenable to culture, key characteristics that enhance their experimental utility. Therefore, the present study is a foundation for cartilaginous fish research. Developmental Dynamics 247:712–723, 2018. © 2017 Wiley Periodicals, Inc.