Expression, purification and molecular analysis of the human ZNF706 protein

Background: The ZNF706 gene encodes a protein that belongs to the zinc finger family of proteins and was found to be highly expressed in laryngeal cancer, making the structure and function of ZNF706 worthy of investigation. In this study, we expressed and purified recombinant human ZNF706 that was suitable for structural analysis in Escherichia coli BL21(DH3). Findings. ZNF706 mRNA was extracted from a larynx tissue sample, and cDNA was ligated into a cloning vector using the TOPO method. ZNF706 protein was expressed according to the E. coli expression system procedures and was purified using a nickel-affinity column. The structural qualities of recombinant ZNF706 and quantification alpha, beta sheet, and other structures were obtained by spectroscopy of circular dichroism. ZNF706's structural modeling showed that it is composed of α-helices (28.3%), β-strands (19.4%), and turns (20.9%), in agreement with the spectral data from the dichroism analysis. Conclusions: We used circular dichroism and molecular modeling to examine the structure of ZNF706. The results suggest that human recombinant ZNF706 keeps its secondary structures and is appropriate for functional and structural studies. The method of expressing ZNF706 protein used in this study can be used to direct various functional and structural studies that will contribute to the understanding of its function as well as its relationship with other biological molecules and its putative role in carcinogenesis. © 2013 Colombo et al.; licensee BioMed Central Ltd.

Encuesta latinoamericana sobre capacitacion en materia de asentamientos humanos; analisis de la situacion actual y propuestas para un programa de accion regional = Latin American survey on human settlements training; analysis of present situation and proposals for a regional programme

Incluye Bibliografía

Paleogenetic and taphonomic analysis of human bones from Moa, Beirada, and Zé Espinho Sambaquis, Rio de Janeiro, Brazil

The present paper discusses mtDNA and taphonomy of human remains from Moa, Beirada, and Zé Espinho sambaquis of Saquarema, state of Rio de Janeiro, Brazil. New human bone dating by 14C-AMS for Moa archeological site (3810+50 BP - GX-31826-AMS) is included. Preservation of microscopic lamellae and DNA is not related to the macroscopic integrity of the bones. Results here suggest that the preservation of amplifiable DNA fragments may have relation to the preservation of the lamellar arrangement as indicated by optical microscopic examination (polarized light). In 13 human bone fragments from Moa, Beirada, and Zé Espinho it was possible to sequence mtDNA from the 3 individuals of Moa, and from 1 of 4 individuals of Beirada, whose bones also show extensive areas with preserved lamellar structures. The 6 human bone fragments of Zé Espinho and 3 of the 4 fragments of Beirada showed extensive destruction of cortical microstructure represented by cavities, intrusive minerals, and agglomerated microscopic bodies of fungi and bacteria; it was not possible to extract mtDNA from these samples. The results support the hypothesis that the preservation of the microscopic osteon organization is a good predictor for DNA preservation. It was also confirmed the C haplogroup antiquity in Brazil.

Molecular analysis of three FUT3 gene single nucleotide polymorphisms and their relationship with the lewis erythrocytary phenotype in a human population of japanese-ancestry living in Tomé Açu, a town in the Brazilian Amazon

The Lewis blood group system involves two major antigens, Le^ª and Le^b. Their antigenic determinants are not primary gene products but are synthesized by the transfer of sugar subunits to a precursory chain by a specific enzyme which is the product of the FUT3 gene (Lewis gene). The presence of three FUT3 gene single nucleotide polymorphisms (SNPs) (59T > G; 508G > A and 1067T > A) was related to the Lewis phenotype of erythrocytes from 185 individuals of Japanese ancestry living in the town of Tomé-Açu in the Brazilian Amazon region. This relationship was detected using a serological hemagglutination test and the Dot-ELISA assay along with the molecular technique PCR-RFLP. We found that the three SNPs investigated in this study only accounted for a proportion of the Lewis-negative phenotype of the erythrocytes.

Molecular analysis of three FUT3 gene single nucleotide polymorphisms and their relationship with the lewis erythrocytary phenotype in a human population of japanese-ancestry living in Tomé Açu, a town in the Brazilian Amazon

The Lewis blood group system involves two major antigens, Le^a and Le^b. Their antigenic determinants are not primary gene products but are synthesized by the transfer of sugar subunits to a precursory chain by a specific enzyme which is the product of the FUT3 gene (Lewis gene). The presence of three FUT3 gene single nucleotide polymorphisms (SNPs) (59T > G; 508G > A and 1067T > A) was related to the Lewis phenotype of erythrocytes from 185 individuals of Japanese ancestry living in the town of Tomé-Açu in the Brazilian Amazon region. This relationship was detected using a serological hemagglutination test and the Dot-ELISA assay along with the molecular technique PCR-RFLP. We found that the three SNPs investigated in this study only accounted for a proportion of the Lewis-negative phenotype of the erythrocytes.

Analysis of rocuronium in human plasma by liquid chromatography-tandem mass spectrometry with application in clinical pharmacokinetics

Rocuronium (ROC) is a neuromuscular blocking agent used in surgical procedures which is eliminated primarily by biliary excretion. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for analysis of ROC in human plasma. Separation of ROC and IS (verapamil) was performed using an endcapped C-18 column and a mixture of water:acetonitrile:trifluoracetic acid (50:50:0.1, v/v) as mobile phase. Aliquots of 100 mu L of human plasma were extracted at pH 3, using dichloromethane. The lower limit of quantification of 5 ng/mL shows the high sensitivity of this method. Intra- and inter-assay precision (as relative standard deviation) was all <= 14.2% and accuracy (as relative standard error) did not exceed 10.1%. The validated method was successfully applied to quantify ROC concentrations in patients under surgical procedures up to 6 h after the administration of the 0.4-0.9 mg/kg ROC. The pharmacokinetic parameter estimations of ROC showed AUC/dose of 563 mu g min/mL, total clearance of 2.5 mL/min/kg, volume of distribution at steady state of 190 mL/kg and mean residence time of 83 min. (C) 2013 Elsevier B.V. All rights reserved.

Defining suitable reference genes for RT-qPCR analysis on human sertoli cells after 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Analysis of fluoxetine and norfluoxetine in human plasma by HPLC-UV using a high purity C18 silica-based SPE sorbent

This paper reports on the development and validation of a simple and sensitive method that uses solid phase extraction (SPE) and liquid chromatography with ultraviolet detection to analyze fluoxetine (FLX) and norfluoxetine (NFLX) in human plasma samples. A lab-made C18 SPE phase was synthesized by using a sol–gel process employing a low-cost silica precursor. This sorbent was fully characterized by nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FT-IR), and scanning electron microscopy (SEM) to check the particles' shape, size and C18 functionalization. The lab-made C18 silica was used in the sample preparation step of human plasma by the SPE-HPLC-UV method. The method was validated in the 15 to 500 ng mL 1 range for both FLX and NFLX using a matrix matched curve. Detection limits of 4.3 and 4.2 ng mL 1 were obtained for FLX and NFLX, respectively. The repeatability and intermediary precision achieved varied from 7.6 to 15.0% and the accuracy ranged from 14.9 to 9.1%. The synthesized C18 sorbent was compared to commercial C18 sorbents. The average recoveries were similar (85–105%), however the lab-made C18 silica showed fewer interfering peaks in the chromatogram. After development and validation, the method using the lab-made C18 SPE was applied to plasma samples of patients under FLX treatment (n ¼ 6). The concentrations of FLX and NFLX found in the samples varied from 46.8–215.5 and 48.0–189.9 ng mL 1 , respectively.

Analysis of acoustic frequency and wave amplitude generated by the Oscillatory Thoracic Thixotropic Device (Diottix(r)) in human chest

Diottix(r) was calibrated at 25 Hz to achieve the frequency indicated in literature as being effective to mobilize the airways secretions. However, the amplitude and frequency of the waves generated by the equipment in different regions of the chest still need to be investigated. The objective of this study was to analyze the frequency and amplitude of waves generated by Diottix(r) in chests of healthy subjects. Diottix(r) was used in the anterior and posterior regions of the chest. The mechanical waves were captured using stethoscopes connected to electret microphones, which were connected to a digital oscilloscope. Frequency and amplitude data were recorded by the stethoscope, positioned in six points in the anterior region and six in the posterior region of the chest, following the positions commonly used in pulmonary auscultation. Signals were recorded and transferred to a computer with software for their analysis. The frequency of waves did not present a significant change (from 24.9 to 26.4 Hz). The wave amplitude in the anterior versus the posterior region in each area of the lung, the upper, middle and lower, had differences. Diottix(r) produces frequencies in the chest according to the calibrated; thus, it can be a complementary resource to bronchial hygiene maneuvers. The amplitudes of waves seem to be affected by other structures like bone parts and heart.

Human development and data envelopment analysis: a structured literature review

Given the importance the concept of productive efficiency has on analyzing the human development process, which is complex and multidimensional, this study conducts a literature review on the research works that have used the data envelopment analysis (DEA) to measure and analyze the development process. Therefore, we researched the databases of Scopus and Web of Science, and considered the following analysis dimensions: bibliometrics, scope, DEA models and extensions used, interfaces with other techniques, units analyzed and depth of analysis. In addition to a brief summary, the main gaps in each analysis dimension were assessed, which may serve to guide future researches. (C) 2015 Elsevier Ltd. All rights reserved.

Computer graphics applied to anatomy: a study of two bio-cad modeling methods on finite element analysis of human edentulous hemi-mandible

Modeling is a step to perform a finite element analysis. Different methods of model construction are reported in literature, as the Bio-CAD modeling. The purpose of this study was to perform a model evaluation and application using two methods of Bio-CAD modeling from human edentulous hemi-mandible on the finite element analysis. From CT scans of dried human skull was reconstructed a stereolithographic model. Two methods of modeling were performed: STL conversion approach (Model 1) associated to STL simplification and reverse engineering approach (Model 2). For finite element analysis was used the action of lateral pterygoid muscle as loading condition to assess total displacement (D), equivalent von-Mises stress (VM) and maximum principal stress (MP). Two models presented differences on the geometry regarding surface number (1834 (model 1); 282 (model 2)). Were observed differences in finite element mesh regarding element number (30428 nodes/16683 elements (model 1); 15801 nodes/8410 elements (model 2). D, VM and MP stress areas presented similar distribution in two models. The values were different regarding maximum and minimum values of D (ranging 0-0.511 mm (model 1) and 0-0.544 mm (model 2), VM stress (6.36E-04-11.4 MPa (model 1) and 2.15E-04-14.7 MPa (model 2) and MP stress (-1.43-9.14 MPa (model 1) and -1.2-11.6 MPa (model 2). From two methods of Bio-CAD modeling, the reverse engineering presented better anatomical representation compared to the STL conversion approach. The models presented differences in the finite element mesh, total displacement and stress distribution.

cDNA representational difference analysis used in the identification of genes expressed by Trichophyton rubrum during contact with keratin

Dermatophytes are adapted to infect skin, hair and nails by their ability to utilize keratin as a nutrient source. Trichophyton rubrum is an anthropophilic fungus, causing up to 90% of chronic cases of dermatophytosis. The understanding of the complex interactions between the fungus and its host should include the identification of genes expressed during infection. To identify the genes involved in the infection process, representational difference analysis (RDA) was applied to two cDNA populations from T. rubrum, one transcribed from the RNA of fungus cultured in the presence of keratin and the other from RNA generated during fungal growth in minimal medium. The analysis identified differentially expressed transcripts. Genes related to signal transduction, membrane protein, oxidative stress response, and some putative virulence factors were up-regulated during the contact of the fungus with keratin. The expression patterns of these genes were also verified by real-time PCR, in conidia of T. rubrum infecting primarily cultured human keratinocytes in vitro, revealing their potential role in the infective process. A better understanding of this interaction will contribute significantly to our knowledge of the process of dermatophyte infection.

The oxidation of p-phenylenediamine, an ingredient used for permanent hair dyeing purposes, leads to the formation of hydroxyl radicals: oxidative stress and DNA damage in human immortalized keratinocytes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Development and validation of an LC-MS/MS method for quantitative analysis of mirtazapine in human plasma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

A general framework for multi-compartmental analysis of drug chemotherapy dynamics in human immunodeficiency virus type-1 infected individuals

An optimal control strategy for the highly active antiretroviral therapy associated to the acquired immunodeficiency syndrome should be designed regarding a comprehensive analysis of the drug chemotherapy behavior in the host tissues, from major viral replication sites to viral sanctuary compartments. Such approach is critical in order to efficiently explore synergistic, competitive and prohibitive relationships among drugs and, hence, therapy costs and side-effect minimization. In this paper, a novel mathematical model for HIV-1 drug chemotherapy dynamics in distinct host anatomic compartments is proposed and theoretically evaluated on fifteen conventional anti-retroviral drugs. Rather than interdependence between drug type and its concentration profile in a host tissue, simulated results suggest that such profile is importantly correlated with the host tissue under consideration. Furthermore, the drug accumulative dynamics are drastically affected by low patient compliance with pharmacotherapy, even when a single dose lacks. (C) 2012 Elsevier Inc. All rights reserved.