Silencing of HTLV-1 gag and env genes by small interfering RNAs in HEK 293 cells

Since the discovery of RNAi technology, several functional genomic and disease therapy studies have been conducted using this technique in the field of oncology and virology. RNAi-based antiviral therapies are being studied for the treatment of retroviruses such as HIV-1. These studies include the silencing of regulatory, infectivity and structural genes. The HTLV-1 structural genes are responsible for the synthesis of proteins involved in the entry, assembly and release of particles during viral infection. To examine the possibility of silencing HTLV-1 genes gag and env by RNA interference technology, these genes were cloned into reporter plasmids. These vectors expressed the target mRNAs fused to EGFP reporter genes. Three small interference RNAs (siRNAs) corresponding to gag and three corresponding to env were designed to analyze the effect of silencing by RNAi technology. The plasmids and siRNAs were co-transfected into HEK 293 cells. The results demonstrated that the expression of the HTLV-1 gag and env genes decreased significantly in vitro. Thus, siRNAs can be used to inhibit HTLV-1 structural genes in transformed cells, which could provide a tool for clarifying the roles of HTLV-1 structural genes, as well as a therapy for this infection. (C) 2011 Elsevier B.V. All rights reserved.

Analysis of Intracellular Substrates and Products of Thimet Oligopeptidase in Human Embryonic Kidney 293 Cells

Thimet oligopeptidase (EC 3.4.24.15; EP24.15) is an intracellular enzyme that has been proposed to metabolize peptides within cells, thereby affecting antigen presentation and G protein-coupled receptor signal transduction. However, only a small number of intracellular substrates of EP24.15 have been reported previously. Here we have identified over 100 peptides in human embryonic kidney 293 (HEK293) cells that are derived from intracellular proteins; many but not all of these peptides are substrates or products of EP24.15. First, cellular peptides were extracted from HEK293 cells and incubated in vitro with purified EP24.15. Then the peptides were labeled with isotopic tags and analyzed by mass spectrometry to obtain quantitative data on the extent of cleavage. A related series of experiments tested the effect of overexpression of EP24.15 on the cellular levels of peptides in HEK293 cells. Finally, synthetic peptides that corresponded to 10 of the cellular peptides were incubated with purified EP24.15 in vitro, and the cleavage was monitored by high pressure liquid chromatography and mass spectrometry. Many of the EP24.15 substrates identified by these approaches are 9-11 amino acids in length, supporting the proposal that EP24.15 can function in the degradation of peptides that could be used for antigen presentation. However, EP24.15 also converts some peptides into products that are 8-10 amino acids, thus contributing to the formation of peptides for antigen presentation. In addition, the intracellular peptides described here are potential candidates to regulate protein interactions within cells.

Fratura de mandíbula: análise de 293 pacientes tratados no Hospital de Clínicas da Universidade Federal de Uberlândia

A fratura de mandíbula ocupa o segundo lugar entre as fraturas dos ossos da face, tendo havido aumento significativo de casos nos últimos anos. A não-identificação e o tratamento inadequado podem levar à deformidade estética ou funcional permanente. OBJETIVO: Avaliar os casos submetidos à redução de fratura de mandíbula no Hospital de Clínicas da Universidade Federal de Uberlândia, entre janeiro de 1974 e dezembro de 2002. FORMA DE ESTUDO: coorte historica. PACIENTES E MÉTODO: Duzentos e noventa e três pacientes foram submetidos à redução de fratura de mandíbula e retrospectivamente foram avaliados segundo fatores relacionados a: paciente, trauma, quadro clínico e tratamento cirúrgico. RESULTADOS: Houve uma clara tendência de aumento do número de fraturas de mandíbula ao longo dos anos. Houve um predomínio no sexo masculino (4:1), com pico de ocorrência entre 20 a 29 anos. As principais causas de fratura da mandíbula neste estudo foram acidentes de trânsito e violência, perfazendo juntas 72,4%. Cento e trinta e cinco pacientes apresentavam fratura única. Os sítios mais acometidos foram, em ordem decrescente, sínfise, côndilo, ângulo, corpo, ramo e processo coronóide. Foram realizadas redução incruenta (28), cruenta (213) e associação das duas (11 pacientes), sendo que 56,8% dos pacientes foram tratados nos primeiros 3 dias e 50,4% recebeu a alta hospitalar até o primeiro pós-operatório. Cerca de 10% dos pacientes apresentaram complicações, sendo osteomielite a mais freqüente. CONCLUSÃO: A incidência de fraturas de mandíbula foi marcadamente maior no sexo masculino, durante a terceira década de vida. A causa mais comum foi o acidente de trânsito e as regiões mais atingidas foram sínfise e côndilo. As fraturas isoladas de mandíbula ocorreram em mais de metade dos casos. A maioria dos pacientes foi tratada nos primeiros três dias e recebeu alta até o primeiro pós-operatório. A redução cruenta foi tratamento mais comumente empregado. A complicação mais freqüente foi a osteomielite.

Jornal da UNESP, 2013, ano XXVII, n. 293

Jornal elaborado pela Assessoria de Comunicação e Imprensa da Reitoria da UNESP

Density measurements of compressed dipropyl, dibutul, bis (2-ethylhexyl) adipates from (293 to 373K) at pressures up to about 68MPa

The article reports density measurements of dipropyl (DPA), dibutyl (DBA) and bis(2-ethylhexyl) (DEHA) adipates, using a vibrating U-tube densimeter, model DMA HP, from Anton Paar GmbH. The measurements were performed in the temperature range (293 to 373) K and at pressures up to about 68 MPa, except for DPA for which the upper limits were 363 K and 65 MPa, respectively. The density data for each liquid was correlated with the temperature and pressure using a modified Tait equation. The expanded uncertainty of the present density results is estimated as 0.2% at a 95% confidence level. No literature density data at pressures higher than 0.1 MPa could be found. DEHA literature data at atmospheric pressure agree with the correlation of the present measurements, in the corresponding temperature range, within +/- 0.11%. The isothermal compressibility and the isobaric thermal expansion were calculated by differentiation of the modified Tait correlation equation. These two parameters were also calculated for dimethyl adipate (DMA), from density data reported in a previous work. The uncertainties of isothermal compressibility and the isobaric thermal expansion are estimated to be less than +/- 1.7% and +/- 1.1%, respectively, at a 95% confidence level. Literature data of isothermal compressibility and isobaric thermal expansivity for DMA have an agreement within +/- 1% and +/- 2.4%, respectively, with results calculated in this work. (C) 2014 Elsevier B.V. All rights reserved.

Tris(2-Ethylhexyl) trimellitate (TOTM) a potential reference fluid for high viscosity. Part II: density measurements at temperatures from (293 to 373)K and pressures up to 85MPa

In Part I of the present work we describe the viscosity measurements performed on tris(2-ethylhexyl) trimellitate or 1,2,4-benzenetricarboxylic acid, tris(2-ethylhexyl) ester (TOTM) up to 65 MPa and at six temperatures from (303 to 373)K, using a new vibrating-wire instrument. The main aim is to contribute to the proposal of that liquid as a potential reference fluid for high viscosity, high pressure and high temperature. The present Part II is dedicated to report the density measurements of TOTM necessary, not only to compute the viscosity data presented in Part I, but also as complementary data for the mentioned proposal. The present density measurements were obtained using a vibrating U-tube densimeter, model DMA HP, using model DMA5000 as a reading unit, both instruments from Anton Paar GmbH. The measurements were performed along five isotherms from (293 to 373)K and at eleven different pressures up to 68 MPa. As far as the authors are aware, the viscosity and density results are the first, above atmospheric pressure, to be published for TOTM. Due to TOTM's high viscosity, its density data were corrected for the viscosity effect on the U-tube density measurements. This effect was estimated using two Newtonian viscosity standard liquids, 20 AW and 200 GW. The density data were correlated with temperature and pressure using a modified Tait equation. The expanded uncertainty of the present density results is estimated as +/- 0.2% at a 95% confidence level. Those results were correlated with temperature and pressure by a modified Tait equation, with deviations within +/- 0.25%. Furthermore, the isothermal compressibility, K-T, and the isobaric thermal expansivity, alpha(p), were obtained by derivation of the modified Tait equation used for correlating the density data. The corresponding uncertainties, at a 95% confidence level, are estimated to be less than +/- 1.5% and +/- 1.2%, respectively. No isobaric thermal expansivity and isothermal compressibility for TOTM were found in the literature. (C) 2014 Elsevier B.V. All rights reserved.

Agenda For Change - Frequently Asked Questions (PDF 293 KB)

Agenda For Change - Frequently Asked Questions

Infection kinetics of human adenovirus serotype 41 in HEK 293 cells

The purpose of this work was to acquire an overview of the infectious cycle of HAdV-41 in permissive HEK 293 cells and compare it to that observed with the prototype of the genus, Human adenovirus C HAdV-2. HEK 293 cells were infected with each virus separately and were harvested every 12 h for seven days. Infection kinetics were analysed using confocal and electronic microscopy. The results show that, when properly cultivated, HAdV-41 was not fastidious. It had a longer multiplication cycle, which resulted in the release of complete viral particles and viral stocks reached high titres. After 60 h of infection, the export of viral proteins from the infected cell to the extracellular milieu was observed, with a pattern similar to that previously described for HAdV-2 penton-base trafficking after 30 h of infection. HAdV-41 had a non-lytic cycle and the infection spread from the first infected cell to its neighbours. The release process of the viral particles is unknown. The results observed for HAdV-41 infection in HEK 293 cells show how different this virus is from the prototype HAdV-2 and provides information for the development of this vector for use in gene therapy.

Experimental Project No. 621, Pavement Instrumentation, HR-293, 1992

This document contains a discussion of the reasons why the project did not succeed. A detailed discussion of the steps taken by the Iowa Department of Transportation to make the experiment work are contained in this document, along with recommendations for future projects.

Pavement Instrumentation, HR-293, 1988

The report documents the development and installation of an instrumented pavement on I-80 in Iowa for the purposes of demonstration and answering current pavement questions. Its two primary objectives include documentation of the installation and verification of existing design procedures through monitoring of the continuous traffic stream reactions in the pavement. Some 120 instruments were installed in a forty foot segment of reconstructed pavement. The instruments included concrete strain gages, weldable strain gages on dowels, LVDT-deflection gages and temperature sensors in the concrete and base material. Five tubes were placed under three joints and two midslabs to measure the relative moisture and density at the interface between the pavement and base with atomic equipment. The instruments were placed ahead of the paving and over 92% of the instruments responded after paving. Planning requirements, problems encountered and costs of installation are presented. The site will use piezoelectric cables in a weigh-in-motion arrangement to trigger the data collection, a microcomputer controlled data acquisition system to analyze multiple sensors simultaneously, and telemetry to monitor the site remotely. Details provide the first time user of instrumentation with valuable information on the planning, problems, costs and coordination required to establish and operate such a site.