Identification of the alpha(6) integrin as a candidate receptor for papillomaviruses

Papillomaviruses (PVs) bind in a specific and saturable fashion to a range of epithelial and other cell lines. Treatment of cells with trypsin markedly reduces their ability to bind virus particles, suggesting that binding is mediated via a cell membrane protein. We have investigated the interaction bf human PV type 6b L1 virus-like particles (VLPs) with two epithelial cell lines, CV-1 and HaCaT, which bind VLPs, and a B-cell line (DG75) previously shown not to bind VLPs. Immunoprecipitation of a mixture of PV VLPs with [S-35]methionine-labeled cell extracts and with biotin-labeled cell surface proteins identified four proteins from CV-1 and HaCaT cells of 220, 120, 87, and 35 kDa that reacted with VLPs and were not present in DG75 cells. The alpha(6) beta(4) integrin complex has subunits corresponding to the VLP precipitated proteins, and the tissue distribution of this complex suggested that it was a candidate human PV receptor. Monoclonal antibodies (MAbs) to the alpha(6) or beta(4) integrin subunits precipitated VLPs from a mixture of CV-1 cell proteins and VLPs, whereas MAbs to other integrin subunits did not. An alpha(6) integrin-specific MAb (GoH3) inhibited VLP binding to CV-1 and HaCaT cells, whereas an anti-beta(4) integrin MAb and a range of integrin-specific and other MAbs did not. Furthermore, human laminin, the natural ligand for the alpha(6) beta(4) integrin, was able to block VLP binding. By use of sections of monkey esophagus, the distribution of alpha(6), integrin expression in the basal epithelium was shown to coincide with the distribution of bound VLPs. Taken together, these data suggest that VLPs bind specifically to the alpha(6) integrin subunit and that integrin complexes containing alpha(6) integrin complexed with either beta(1) or beta(4) integrins may act as a receptor for PV binding and entry into epithelial cells.

Recurrent Laryngeal Nerve A Plexus Rather Than a Nerve?

Objectives: To analyze the frequency of extralaryngeal branching (ELB) of the recurrent laryngeal nerve (RLN) in a consecutive series of patients undergoing thyroidectomy by the same group of surgeons during an extended period and to compare our findings with the data available in the literature. Design: Retrospective medical record study. Setting: Academic research. Patients: From March 1, 1983, to September 30, 2008, 2677 patients underwent thyroidectomy. Of these, 1638 patients had surgical information about at least 1 RLN. A total of 1081 patients underwent bilateral operations. During the last 5 years of the study, intraoperative laryngeal nerve monitoring was performed in selected patients using a commercially available system. Main Outcome Measures: Information was obtained regarding 2154 RLNs. Results: A total of 1390 RLNs (64.53%) had ELB. Among 447 patients in whom intraoperative laryngeal nerve monitoring was used, the anterior branches usually exhibited more electrophysiologic activity. Conclusions: Extralaryngeal branching was found in 64.53% of RLNs in this case series. In recent patients with intraoperative laryngeal nerve monitoring, electrophysiologic activity was observed in the branches, particularly the anteriorly situated ones. Recognition of this frequent anatomical configuration and meticulous preservation of all branches are of paramount importance to decrease postoperative morbidity associated with thyroidectomy.

Evaluation of serum trace element, biochemical and hematological data of a healthy elderly group residing in So Paulo city, Brazil

In this study, blood serum trace elements, biochemical and hematological parameters were obtained to assess the health status of an elderly population residing in So Paulo city, SP, Brazil. Results obtained showed that more than 93% of the studied individuals presented most of the serum trace element concentrations and of the hematological and biochemical data within the reference values used in clinical laboratories. However, the percentage of elderly presenting recommended low density lipoprotein (LDL) cholesterol concentrations was low (70%). The study indicated positive correlation between the concentrations of Zn and LDL-cholesterol (p < 0.06).

Identification of products formed by a fructan:fructan fructosyltransferase activity from Lolium rigidum

The products formed by a fructan:fructan fructosyltransferase (FFT) activity purified from Lolium rigidum Gaudin were identified after gas chromatography-mass spectrometry of partially methylated alditol acetates, electrospray ionization-mass spectrometry and reversed-phase high-performance liquid chromatography. The FFT activity synthesized oligofructans up to degree of polymerization (DP) 6, but did not synthesize fructans of DP > 6 even when assayed with (1,1,1)-kestopentaose for up to 10 h. The FFT activity when assayed with 1-kestose or 6(G)-kestose synthesized fructan with fructosyl residues almost exclusively linked by beta-2,1-glycosidic linkages. When assayed with 1-kestose, the FFT activity synthesized tetrasaccharides and pentasaccharides with an internal glucosyl residue. The predominant tetrasaccharide was (1&6(G))-kestotetraose and the predominant pentasaccharide was (1&6(G),1)-kestopentaose. By comparison, tetrasaccharides and pentasaccharides extracted from L. rigidum also contained predominantly beta-2,1-glycosidic linked fructans with an internal glucosyl residue. The only exception was that one of the pentasaccharides contained beta-2,1- and beta-2,6-glycosidic linked fructosyl residues. This pentasaccharide was not synthesized by the FFT activity. The role of this FFT activity in formation of oligofructans in L. rigidum is discussed.

Anti-Group A Streptococcal Vaccine Epitope STRUCTURE, STABILITY, AND ITS ABILITY TO INTERACT WITH HLA CLASS II MOLECULES

Streptococcus pyogenes infections remain a health problem in several countries due to poststreptococcal sequelae. We developed a vaccine epitope (StreptInCor) composed of 55 amino acids residues of the C-terminal portion of the M protein that encompasses both T and B cell protective epitopes. The nuclear magnetic resonance (NMR) structure of the StreptInCor peptide showed that the structure was composed of two microdomains linked by an 18-residue alpha-helix. A chemical stability study of the StreptInCor folding/unfolding process using far-UV circular dichroism showed that the structure was chemically stable with respect to pH and the concentration of urea. The T cell epitope is located in the first microdomain and encompasses 11 out of the 18 alpha-helix residues, whereas the B cell epitope is in the second microdomain and showed no alpha-helical structure. The prediction of StreptInCor epitope binding to different HLA class II molecules was evaluated based on an analysis of the 55 residues and the theoretical possibilities for the processed peptides to fit into the P1, P4, P6, and P9 pockets in the groove of several HLA class II molecules. We observed 7 potential sites along the amino acid sequence of StreptInCor that were capable of recognizing HLA class II molecules (DRB1*, DRB3*, DRB4*, and DRB5*). StreptInCoroverlapping peptides induced cellular and humoral immune responses of individuals bearing different HLA class II molecules and could be considered as a universal vaccine epitope.

A novel method for development of species and strain-specific DNA probes and PCR primers for identifying Burkholderia solanacearum (formerly Pseudomonas solanacearum)

Six Burkholderia solanacearum (formerly Pseudomonas solanacearum) genomic DNA fragments were isolated, using RAPD techniques and cloning, from the three genetically diverse strains: ACH092 (Biovar 4), ACH0158 (Biovar 2) and ACH0171 (Biovar 3) (1). One of these cloned fragments was selected because it was present constantly in all bacterial strains analysed. The remaining five clones were selected because Southern hybridisation revealed that each showed partial or complete specificity towards the strain of origin. A seventh genomic fragment showing a strain-specific distribution in Southern hybridisations was obtained by differential restriction, hybridisation and cloning of genomic DNA. Each of these clones was sequenced and primers to amplify the insert were designed. When DNA from the strain of origin was used as template, PCR amplification for each of these fragments yielded a single band on gel analysis. One pair of primers amplified the species-constant fragment of 281 bp from DNA of all B. solanacearum strains investigated, from DNA of the closely related bacterium which causes ''blood disease'' of banana (BDB) and in P. syzigii. The sensitivity of detection of B. solanacearum using these ubiquitous primers was between 1.3 and 20 bacterial cells. The feasibility and reliability of a PCR approach to detection and identification of B. solanacearum was tested in diverse strains of the bacterium in several countries and laboratories.

Clinical profile of patients enroled in a cell therapy trial for severe coronary artery disease

Aims and objectives. To compare the clinical profile of patients included in a clinical trial of autologous bone marrow cells as an adjunctive therapy to coronary artery bypass grafting with that of patients undergoing routine coronary artery bypass grafting. Background. The therapeutic potential of autologous bone marrow cells has been explored in the treatment of severe coronary artery disease. There are few data regarding the clinical and socio-economic profile of patients included in clinical trials using bone marrow cell. Design. Case-control study. Method. Sixty-seven patients (61 SD 9) years, 82% men) with multivessel coronary artery disease were divided into two groups: patients in the bone marrow cell group (n = 34) underwent incomplete coronary artery bypass grafting + intramyocardial injection of autologous bone marrow cells (lymphomonocytic fraction -2.0 (SD 0.2 x 108) cells/patient) in the ischaemic, non-revascularised myocardium, whereas patients in the coronary artery bypass grafting group (n = 33) underwent routine bypass surgery. Demographics, socio-economic status, clinical and echocardiographic data were collected. Statistical analysis included the Fisher`s exact test (categorical variables) and the Student`s t-test (continuous variables). Results. There were no significant differences between groups regarding age, gender, BMI, heart rate, blood pressure and echo data. There was a greater prevalence of obesity (65 vs. 33%; OR = 3.7 [1.3-10.1]), of previous myocardial infarction (68 vs. 39%; OR = 3.2 [1.2-8.8]) and prior revascularisation procedures (59 vs. 24%; OR = 4.5 [1.6-12.7]) in the autologous bone marrow cells group and of smokers in the coronary artery bypass grafting group (51 vs. 23%; OR = 3.5 [1.2-10.4]). Conclusions. Patients included in this clinical trial of autologous bone marrow cells for severe coronary artery disease presented a greater prevalence of myocardial revascularisation procedures, indicating a more severe clinical presentation of the disease. Fewer smokers in this group could be attributable to life style changes after previous cardiovascular events and/or interventions. Relevance to clinical practice. The knowledge of the clinical profile of patients included in cell therapy trials may help researchers in the identification of patients that may be enroled in future clinical trials of this new therapeutic strategy.

On the distance between distinct group Latin squares

In an article in 1992, Drapal addressed the question of how far apart the multiplication tables of two groups can be? In this article we continue this investigation; in particular, we study the interaction between partial equalities in the multiplication tables of the two groups and their subgroup structure. (C) 1997 John Wiley & Sons, Inc.

Congenic strains provide evidence that four mapped loci in chromosomes 2, 4, and 16 influence hypertension in the SHR

Aneas I, Rodrigues MV, Pauletti BA, Silva GJ, Carmona R, Cardoso L, Kwitek AE, Jacob HJ, Soler JM, Krieger JE. Congenic strains provide evidence that four mapped loci in chromosomes 2, 4, and 16 influence hypertension in the SHR. Physiol Genomics 37: 52-57, 2009. First published January 6, 2009; doi: 10.1152/physiolgenomics.90299.2008. - To dissect the genetic architecture controlling blood pressure (BP) regulation in the spontaneously hypertensive rat (SHR) we derived congenic rat strains for four previously mapped BP quantitative trait loci (QTLs) in chromosomes 2, 4, and 16. Target chromosomal regions from the Brown Norway rat (BN) averaging 13 - 29 cM were introgressed by marker-assisted breeding onto the SHR genome in 12 or 13 generations. Under normal salt intake, QTLs on chromosomes 2a, 2c, and 4 were associated with significant changes in systolic BP (13, 20, and 15 mmHg, respectively), whereas the QTL on chromosome 16 had no measurable effect. On high salt intake (1% NaCl in drinking water for 2 wk), the chromosome 16 QTL had a marked impact on SBP, as did the QTLs on chromosome 2a and 2c (18, 17, and 19 mmHg, respectively), but not the QTL on chromosome 4. Thus these four QTLs affected BP phenotypes differently: 1) in the presence of high salt intake (chromosome 16), 2) only associated with normal salt intake (chromosome 4), and 3) regardless of salt intake (chromosome 2c and 2a). Moreover, salt sensitivity was abrogated in congenics SHR. BN2a and SHR. BN16. Finally, we provide evidence for the influence of genetic background on the expression of the mapped QTLs individually or as a group. Collectively, these data reveal previously unsuspected nuances of the physiological roles of each of the four mapped BP QTLs in the SHR under basal and/or salt loading conditions unforeseen by the analysis of the F2 cross.

Human breast tumor slices: A model for identification of vitamin D regulated genes in the tumor microenvironment

While many studies have addressed the direct effects of 1 alpha,25(OH)(2)D(3) on breast cancer (BC) cells, stromal-epithelial interactions, which are important for the tumor development, have been largely ignored. In addition, high concentrations of the hormone, which cannot be attained in vivo, have been used. Our aim was to establish a more physiological breast cancer model, represented by BC tissue slices, which maintain epithelial-mesenchymal interactions, cultured with a relatively low 1 alpha,25(OH)(2)D(3) concentration, in order to evaluate the vitamin D pathway. Freshly excised human BC samples were sliced and cultured in complete culture media containing vehicle, 0.5 nM or 100 nM 1 alpha,25(OH)(2)D(3) for 24 h. BC slices remained viable for at least 24 h, as evaluated by preserved tissue morphology in hematoxylin and eosin (HE) stained sections and bromodeoxyuridine (BrdU) incorporation by 10% of tumor cells. VDR mRNA expression was detected in all samples and CYP24A1 mRNA expression was induced by 1 alpha,25(OH)(2)D(3) in both concentrations (but mainly with 100 nM). Our results indicate that the vitamin D signaling pathway is functional in BC slices, a model which preserves stromal-epithelial interactions and mimics in vivo conditions. (C) 2010 Elsevier Ltd. All rights reserved.

Use of SVM Methods with Surface-Based Cortical and Volumetric Subcortical Measurements to Detect Alzheimer`s Disease

Here, we examine morphological changes in cortical thickness of patients with Alzheimer`s disease (AD) using image analysis algorithms for brain structure segmentation and study automatic classification of AD patients using cortical and volumetric data. Cortical thickness of AD patients (n = 14) was measured using MRI cortical surface-based analysis and compared with healthy subjects (n = 20). Data was analyzed using an automated algorithm for tissue segmentation and classification. A Support Vector Machine (SVM) was applied over the volumetric measurements of subcortical and cortical structures to separate AD patients from controls. The group analysis showed cortical thickness reduction in the superior temporal lobe, parahippocampal gyrus, and enthorhinal cortex in both hemispheres. We also found cortical thinning in the isthmus of cingulate gyrus and middle temporal gyrus at the right hemisphere, as well as a reduction of the cortical mantle in areas previously shown to be associated with AD. We also confirmed that automatic classification algorithms (SVM) could be helpful to distinguish AD patients from healthy controls. Moreover, the same areas implicated in the pathogenesis of AD were the main parameters driving the classification algorithm. While the patient sample used in this study was relatively small, we expect that using a database of regional volumes derived from MRI scans of a large number of subjects will increase the SVM power of AD patient identification.

Synthesis, identification, characterization, stability, solubility, and protein binding of ester derivatives of salicylic acid and diflunisal

O-Acyl esters were prepared from salicylic acid and diflunisal by esterification with the appropriate acyl anhydride (in the presence of sulfuric acid at 80 degrees C) or acyl chloride (in the presence of pyridine at 0 degrees C). Synthesis, identification and characterization of these compounds is described. In vitro hydrolysis, solubility and protein binding studies of these O-acyl esters were performed. For the diflunisal esters, the melting points fell as the side chain was increased from ethyl to pentyl. The melting points showed no significant difference as the length of the side chain was increased from pentyl to heptyl. The aspirin analogues showed a similar trend, The relationship between solubility and carbon chain length agreed closely with that for the melting points with carbon chain length. In vitro non-enzymatic hydrolysis studies concluded that: (1) hydrolysis rate constants generally decreased with carbon chain length; (2) the diflunisal esters have shorter half lives compared with their salicylate counterparts; and (3) the in vitro hydrolysis of these compounds was retarded by the presence of bovine serum albumin. Protein binding experiments showed that the strength of binding of the aspirin and diflunisal analogues to bovine serum albumin increased with carbon chain length. (C) 1997 Elsevier Science B.V.

Hyperplane navigation: A method to set individual scores in fMRI group datasets

Recent studies have demonstrated that spatial patterns of fMRI BOLD activity distribution over the brain may be used to classify different groups or mental states. These studies are based on the application of advanced pattern recognition approaches and multivariate statistical classifiers. Most published articles in this field are focused on improving the accuracy rates and many approaches have been proposed to accomplish this task. Nevertheless, a point inherent to most machine learning methods (and still relatively unexplored in neuroimaging) is how the discriminative information can be used to characterize groups and their differences. In this work, we introduce the Maximum Uncertainty Linear Discrimination Analysis (MLDA) and show how it can be applied to infer groups` patterns by discriminant hyperplane navigation. In addition, we show that it naturally defines a behavioral score, i.e., an index quantifying the distance between the states of a subject from predefined groups. We validate and illustrate this approach using a motor block design fMRI experiment data with 35 subjects. (C) 2008 Elsevier Inc. All rights reserved.

Enolase from Paracoccidioides brasiliensis: isolation and identification as a fibronectin-binding protein

Paracoccidioides brasiliensis yeast cells can enter mammalian cells and may manipulate the host cell environment to favour their own growth and survival. Moreover, fibronectin and several other host extracellular matrix proteins are recognized by various components of the yeast cell extracts. The present study was designed to isolate and characterize a fibronectin-binding protein from P. brasiliensis. We also compared P. brasiliensis strain 18, tested before (Pb18a) and after (Pb18b) animal passage, in relation to its adhesion and invasion processes. Extracts from both samples, when cultured on blood agar solid medium, showed higher levels of protein expression than when the same samples were cultured on Fava-Netto solid medium, as demonstrated by two-dimensional electrophoresis and SDS-PAGE. Also, both Pb18a and Pb18b exhibited stronger adhesion to A549 epithelial cells when cultured on blood agar medium than when cultured on Fava-Netto medium. Ligand affinity binding assays revealed a protein of 54 kDa and pl 5.6 in P. brasiliensis cell-free extracts with the properties of a fibronectin-binding adhesin, which was characterized by tryptic digestion and mass spectroscopy as a homologue of enolase from P. brasiliensis. Antibody raised against this 54 kDa protein abolished 80 % of P. brasiliensis adhesion to A549 epithelial cells. Our results demonstrate that P. brasiliensis produces a fibronectin-binding adhesin, irrespective of the culture medium, and that this activity can be inhibited by a specific antibody and is involved in the adhesion of the fungus to pulmonary epithelial cells.

A high-resolution genetic map of the familial Mediterranean fever candidate region allows identification of haplotype-sharing among ethnic groups

Familial Mediterranean fever (FMF) is a recessive disorder of inflammation caused by mutations in a gene (designated MEFV) on chromosome 16p13.3, We have recently constructed a 1-Mb cosmid contig that includes the FMF critical region. Here we show genotype data for 12 markers from our physical map, including 5 newly identified microsatellites, in FMF families. Intrafamilial recombinations placed MEFV in the similar to 285 kb between D16S468/D16S3070 and D16S3376. We observed significant linkage disequilibrium in the North African Jewish population, and historical recombinants in the founder haplotype placed MEFV between D16S3082 and D16S3373 (similar to 200 kb). In smaller panels of Iraqi Jewish, Arab, and Armenian families, there were significant allelic associations only for D16S3370 and D16S2617 among the Armenians. A sizable minority of Iraqi Jewish and Armenian carrier chromosomes appeared to be derived from the North African Jewish ancestral haplotype. We observed a unique FMF haplotype common to Iraqi Jews, Arabs, and Armenians and two other haplotypes restricted to either the Iraqi Jewish or the Armenian population. These data support the view that a few major mutations account for a large percentage of the cases of FMF and suggest that same of these mutations arose before the affected Middle Eastern populations diverged from one another. (C) 1997 Academic Press.