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OBJECTIVES: This study was designed to: 1) evaluate and compare the evidence-practice gap (EPG) in minimal intervention dentistry (MID) in Japan and Brazil by measuring concordance between dentists' clinical practice and published evidence; and 2) identify dentists' factors associated with the EPG in both countries. METHODS: We performed a cross-sectional study using a web-delivered questionnaire among 136 Japanese and 110 Brazilian dentists. The questionnaire consisted of three questions concerning "restoration diagnosis and treatment," "deep caries diagnosis and treatment," and "caries risk assessment" regarding MID. A chi-square test was used to analyze differences in concordance among clinical practice and evidence from the literature between Japanese and Brazilian dentists. Logistic regression analyses were performed to analyze dentists' factors associated with overall concordance for all three questions. RESULTS: Overall concordance was significantly higher in Brazil (55%) than in Japan (38%) (p<0.01). Concerning how evidence was obtained, textbooks, nonacademic journals, and seminars and workshops were used as information sources more frequently by Japanese than Brazilian dentists (p<0.001), whereas scientific journal articles in English were used more frequently by Brazilian dentists (p<0.001). On logistic regression analysis, overall concordance was higher for Japanese dentists who frequently obtained evidence from scientific journal articles in English (p<0.05), whereas Brazilian dentists who frequently obtained evidence from the Internet were associated with lower overall concordance (p<0.05). CONCLUSIONS: Because overall concordance was significantly higher in Brazil than in Japan, Japan may have a greater EPG in MID practice. Specific characteristics of Japanese and Brazilian dentists showed significant associations with overall concordance.

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In vitro studies have shown that multidrug resistance protein 1 (MDR1) has an affinity for cortisol; however, in vivo association studies on the relationship between MDR1 gene polymorphisms and blood cortisol levels have produced inconsistent results. Therefore, we examined the effects of the C3435T polymorphism of the MDR1 gene on blood levels of hypothalamus-pituitary-adrenal (HPA) axis hormones such as cortisol and adrenocorticotropic hormone (ACTH) in healthy subjects. The subjects comprised 30 healthy Japanese males. Ten subjects were recruited for each of the C3435T MDR1 genotypes: C/C, C/T, and T/T. Blood samples were taken at 6:00 pm on two occasions with an interval of 2 weeks. Blood levels of cortisol and ACTH were determined by an electrochemiluminescence immunoassay. There were no significant differences in the blood levels of the HPA axis hormones among the MDR1 genotypes. The present study suggests that the C3435T MDR1 polymorphism does not affect blood levels of HPA axis hormones in healthy Japanese males.

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Holstein cattle dominate the global milk production industry because of their outstanding milk production, however, this breed is susceptible to tropical endemic pathogens and suffers from heat stress and thus fewer Holstein populations are raised in tropical areas. The bovine major histocompatibility complex (BoLA)-DRB3 class II gene is used as a marker for disease and immunological traits, and its polymorphism has been studied extensively in Holstein cattle from temperate and cold regions. We studied the genetic diversity of the BoLA-DRB3 gene in South American Holstein populations to determine whether tropical populations have diverged from those bred in temperate and cold regions by selection and/or crossbreeding with local native breeds. We specifically studied Exon 2 of this gene from 855 South American Holstein individuals by a polymerase chain reaction (PCR) sequence-based typing method. We found a high degree of gene diversity at the allelic (Na > 20 and He > 0.87) and molecular (π > 0.080) levels, but a low degree of population structure (FST = 0.009215). A principal components analysis and tree showed that the Bolivian subtropical population had the largest genetic divergence compared with Holsteins bred in temperate or cold regions, and that this population was closely related to Bolivian Creole cattle. Our results suggest that Holstein genetic divergence can be explained by selection and/or gene introgression from local germplasms. This is the first examination of BoLA-DRB3 in Holsteins adapted to tropical environments, and contributes to an ongoing effort to catalog bovine MHC allele frequencies by breed and location.

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Bovine leukocyte antigens (BoLAs) are used extensively as markers for bovine disease and immunological traits. In this study, we estimated BoLA-DRB3 allele frequencies using 888 cattle from 10 groups, including seven cattle breeds and three crossbreeds: 99 Red Angus, 100 Black Angus, 81 Chilean Wagyu, 49 Hereford, 95 Hereford × Angus, 71 Hereford × Jersey, 20 Hereford × Overo Colorado, 113 Holstein, 136 Overo Colorado, and 124 Overo Negro cattle. Forty-six BoLA-DRB3 alleles were identified, and each group had between 12 and 29 different BoLA-DRB3 alleles. Overo Negro had the highest number of alleles (29); this breed is considered in Chile to be an 'Old type' European Holstein Friesian descendant. By contrast, we detected 21 alleles in Holstein cattle, which are considered to be a 'Present type' Holstein Friesian cattle. Chilean cattle groups and four Japanese breeds were compared by neighbor-joining trees and a principal component analysis (PCA). The phylogenetic tree showed that Red Angus and Black Angus cattle were in the same clade, crossbreeds were closely related to their parent breeds, and Holstein cattle from Chile were closely related to Holstein cattle in Japan. Overall, the tree provided a thorough description of breed history. It also showed that the Overo Negro breed was closely related to the Holstein breed, consistent with historical data indicating that Overo Negro is an 'Old type' Holstein Friesian cattle. This allelic information will be important for investigating the relationship between major histocompatibility complex (MHC) and disease.

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Children with Kawasaki disease (n = 82), treated with intravenous immune globulin (IVIG) at a high dose, were classified as IVIG-responsive (defervescence within 5 days of starting IVIG, n = 69) or IVIG-non-responsive (consistent fever over a 6-day period since starting IVIG, n = 13). One patient in the IVIG-responsive group had a coronary artery abnormality during the acute phase (1. 4%) versus 5 in the IVIG-non-responsive group (38.5%). Age, duration of fever before the initiation of IVIG therapy, and laboratory data obtained on admission were tested by the Mann-Whitney U test. Serum levels of C-reactive protein, total bilirubin, lactate dehydrogenase, and gamma-glutamyltranspeptidase were significantly higher (P =.002, P <.001, P <.034, and P <.038, respectively), and the hemoglobin value was significantly lower (P =.025) in patients in the non-responsive group. A multivariate analysis showed that serum levels of C-reactive protein (P =.006), lactate dehydrogenase (P =. 035), and total bilirubin (P =.046) on admission were independent correlates of the success of IVIG therapy. By defining the predictive values, patients with a C-reactive protein level >10 mg/dL, LDH level >590 IU/L, and/or hemoglobin value <10 g/dL are considered non-responsive to IVIG. Additional therapy at an early stage of the disease should be considered for patients who are predicted to be IVIG-non-responsive.

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The MRP gene contributes to one form of multidrug resistance. To identify drugs interacting with MRP, we measured MRP mRNA expression by quantitative PCR in 60 cell lines of the National Cancer Institute Anticancer Drug Screen. Expression was detected in all cell lines (highest in lung carcinomas and central nervous system tumors) with a range of 14-fold. A mean graph of MRP mRNA levels was constructed to determine Pearson correlation coefficients (PCCs) with mean graphs of >40,000 compounds using the COMPARE analysis. Only 20 compounds had PCCs of >/=0.500. The PCCs for VP-16, doxorubicin, and vincristine were 0.008, 0.13, and 0.257, respectively. Initially, 36 compounds with PCCs of >/=0.428 were analyzed using two MRP-overexpressing cell lines; low levels of cross-resistance was demonstrated for 23 compounds (1.3-9.4-fold). Twenty-four compounds also were available for further studies. Using a fluorescence activated cell sorter assay to measure competition of calcein efflux from MRP-overexpressing cells, 10 compounds were found to increase calcein retention by >/=2-fold. Ten compounds also were able to reduce ATP-dependent [3H]LTC4 transport into vesicles from MRP-overexpressing cells. These results contrast with previous studies with MDR-1 in which high correlations were found and confirmed for a large number of compounds. Although other assays may be more revealing, in these unselected cell lines, MRP mRNA expression was a poor predictor of drug sensitivity. This raises the possibility that other factors, including conjugating enzymes, glutathione levels, or other transporters, confound the MRP effect.

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The mini-exon gene is unique and is tandemly repeated in the Leishmania genome. The transcribed region is highly conserved, but the non-transcribed spacer region is distinct in length and in sequence among different Leishmania species. The usefulness of PCR amplification of the Leishmania mini-exon gene was examined for molecular epidemiology of visceral and cutaneous leishmaniasis. We previously described a PCR method for amplification of the mini-exon gene and obtained positive amplification in bone marrow aspirates of patients with visceral leishmaniasis in China. In this study, we have cloned and sequenced two PCR products from the patients. The sequences of two products revealed 100% identity and showed more similarity to the mini-exon gene of L. donovani Indian strain than those of L. donovani complex in Africa and South America. We also applied this PCR method to the diagnosis of cutaneous leishmaniasis. We obtained positive PCR amplification in skin biopsy materials taken from patients with cutaneous leishmaniasis in Ecuador. Since this PCR amplification is simple and requires only a pair of primers to detect all Leishmania species distributed in Ecuador, the method may be a useful tool for the detection of parasites, not only from patients, but also from sandflies and reservoir animals in this area of endemicity.

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The methanol extract of Brazilian propolis was fractionated by HPLC, based on HuH13 (human hepatocellular carcinoma) cell cytotoxicity assay. Two isomers of diterpenoid with a molecular formula of C20H30O3 (MW: 318.46) were isolated. The structures of these colorless compounds were determined as clerodane diterpenoids (I, 15-oxo-3, 13Z-kolavadien-17-oic acid; II, 15-oxo-3Z, 13E-kolavadien-17-oic acid).

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Molecular karyotypes of Leishmania isolates from patients with cutaneous leishmaniasis in Ecuador were analyzed by pulsed-field gel electrophoresis (PFGE) and Southern blot hybridization. The DNA karyotypes of L. major-like parasites were similar between two human isolates from a lowland coastal and a highland Andean region, but were apparently different from those of eleven World Health Organization reference strains including L. major. The smallest chromosome of 240 kilobases in L. major-like parasites was found to belong to the 715-class of small linear chromosomal DNAs, which have been shown to appear in some lines of Leishmania. Chromosome banding patterns of L. mexicana isolates exhibited a novel, ordered, chromosomal ladder, and were identical among four human isolates and one canine isolate from a restricted geographic region in the Andes. On the other hand, minor chromosome size polymorphisms were observed among three L. panamensis isolates from different endemic regions near the Pacific Coast. Chromosomal locations of dihydrofolate reductase-thymidylate synthetase and P-glycoprotein genes revealed further differences in chromosomal organizations among these Leishmania species in Ecuador. These results indicate that karyotype analysis by PFGE is useful for epidemiologic studies of leishmaniasis in Ecuador.

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Infection with the nematode Nippostrongylus brasiliensis (NB) induces the intense production of specific and non-specific immunoglobulin E (IgE) in rats. In the present study, we analysed NB-derived allergenic substances and the variability of IgE antibody production in response to these allergens in outbred Sprague-Dawley rats. Two kinds of crude allergens were used: the excretory-secretory products (ES) of adult NB, and an extract of homogenized adult worms (AW). ELISA showed that IgE antibody titres to ES were more than five times higher than the titres to AW. In the homologous passive cutaneous anaphylaxis (PCA) reaction using serum from infected rats, as little as 50 micrograms of ES had a maximal PCA activity, while even 1 mg of AW still gave a slightly lower PCA titre. Thus, it appeared that ES contained more allergen than AW at the same amount of total proteins. By immunoblot analysis, at least six components were recognized by IgE antibodies from infected animals, and these components were exactly the same in both ES and AW. The results indicated that the allergenic components in ES and AW were the same molecules, and that only those molecules which could be excreted or secreted from living worms were allergenic. Among the array of allergens, 130,000 and 70,000 molecular weight (mw) molecules were commonly recognized by IgE from all serum samples examined, while other components of the allergens were recognized variably by IgE antibodies from individual animals. These findings suggested that individual animals varied considerably in their IgE antibody production to the different constituents of the nematode allergens.

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The localization of Plasmodium brasilianum antigens, common to merozoite micronemes and parasite-induced structures in the host erythrocyte, was determined by means of immunogold electron microscopy and monoclonal antibodies directed against blood stages of this parasite. All monoclonal antibodies reacted with micronemes. In addition, some reacted with either knob protrusions or caveolae of the host erythrocyte membrane; one reacted with a parasite-derived antigen present in the erythrocyte cytoplasm. Gold particles appeared over the membranes of ring-infected cells before the appearance of knobs and caveolae. We hypothesize that at least some knob- and caveolae-associated antigens of P. brasilianum are inserted into the erythrocyte membrane at the time of merozoite invasion.

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The localization of Plasmodium brasilianum-derived antigens in short and long clefts within the cytoplasm of infected erythrocytes and in association with knobs of the host cell membrane was demonstrated by immunoelectron microscopy with monoclonal antibodies. Our results document that malaria-induced short and long clefts, previously distinguishable only by morphology, differ also in antigenic composition. Another parasite-derived antigen was found to be associated with the parasitophorous vacuole space in schizonts. In segmenters, this antigen was present in large amounts between merozoites and in the cytoplasm of infected cells. These antigens were characterized by biosynthetic labeling and gel electrophoresis.

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