Mammalian cell transfection: the present and the future

Transfection is a powerful analytical tool enabling study of the function of genes and gene products in cells. The transfection methods are broadly classified into three groups; biological, chemical, and physical. These methods have advanced to make it possible to deliver nucleic acids to specific subcellular regions of cells by use of a precisely controlled laser-microcope system. The combination of point-directed transfection and mRNA transfection is a new way of studying the function of genes and gene products. However, each method has its own advantages and disadvantages so the optimum method depends on experimental design and objective

Valproic acid inhibits adhesion of vincristine- and cisplatin-resistant neuroblastoma tumour cells to endothelium

Drug resistance to chemotherapy is often associated with increased malignancy in neuroblastoma (NB). In pursuit of alternative treatments for chemoresistant tumour cells, we tested the response of multidrug-resistant SKNSH and of vincristine (VCR)-, doxorubicin (DOX)-, or cisplatin (CDDP)-resistant UKF-NB-2, UKF-NB-3 or UKF-NB-6 NB tumour cell lines to valproic acid (VPA), a differentiation inducer currently in clinical trials. Drug resistance caused elevated NB adhesion (UKF-NB-2VCR, UKF-NB-2DOX, UKF-NB-2CDDP, UKF-NB-3VCR, UKF-NB-3CDDP, UKF-NB-6VCR, UKF-NB-6CDDP) to an endothelial cell monolayer, accompanied by downregulation of the adhesion receptor neural cell adhesion molecule (NCAM). Based on the UKF-NB-3 model, N-myc proteins were enhanced in UKF-NB-3VCR and UKF-NB-3CDDP, compared to the drug naïve controls. p73 was diminished, whereas the p73 isoform deltaNp73 was upregulated in UKF-NB-3VCR and UKF-NB-3CDDP. Valproic acid blocked adhesion of UKF-NB-3VCR and UKF-NB-3CDDP, but not of UKF-NB-3DOX, and induced the upregulation of NCAM surface expression, NCAM protein content and NCAM coding mRNA. Valproic acid diminished N-myc and enhanced p73 protein level, coupled with downregulation of deltaNp73 in UKF-NB-3VCR and UKF-NB-3CDDP. Valproic acid also reverted enhanced adhesion properties of drug-resistant UKF-NB-2, UKF-NB-6 and SKNSH cells, and therefore may provide an alternative approach to the treatment of drug-resistant NB by blocking invasive processes

CADM1 is a strong neuroblastoma candidate gene that maps within a 3.72 Mb critical region of loss on 11q23

<p>Abstract</p> <p>Background</p> <p>Recurrent loss of part of the long arm of chromosome 11 is a well established hallmark of a subtype of aggressive neuroblastomas. Despite intensive mapping efforts to localize the culprit 11q tumour suppressor gene, this search has been unsuccessful thus far as no sufficiently small critical region could be delineated for selection of candidate genes.</p> <p>Methods</p> <p>To refine the critical region of 11q loss, the chromosome 11 status of 100 primary neuroblastoma tumours and 29 cell lines was analyzed using a BAC array containing a chromosome 11 tiling path. For the genes mapping within our refined region of loss, meta-analysis on published neuroblastoma mRNA gene expression datasets was performed for candidate gene selection. The DNA methylation status of the resulting candidate gene was determined using re-expression experiments by treatment of neuroblastoma cells with the demethylating agent 5-aza-2'-deoxycytidine and bisulphite sequencing.</p> <p>Results</p> <p>Two small critical regions of loss within 11q23 at chromosomal band 11q23.1-q23.2 (1.79 Mb) and 11q23.2-q23.3 (3.72 Mb) were identified. In a first step towards further selection of candidate neuroblastoma tumour suppressor genes, we performed a meta-analysis on published expression profiles of 692 neuroblastoma tumours. Integration of the resulting candidate gene list with expression data of neuroblastoma progenitor cells pinpointed <it>CADM1 </it>as a compelling candidate gene. Meta-analysis indicated that <it>CADM1 </it>expression has prognostic significance and differential expression for the gene was noted in unfavourable neuroblastoma versus normal neuroblasts. Methylation analysis provided no evidence for a two-hit mechanism in 11q deleted cell lines.</p> <p>Conclusion</p> <p>Our study puts <it>CADM1 </it>forward as a strong candidate neuroblastoma suppressor gene. Further functional studies are warranted to elucidate the role of <it>CADM1 </it>in neuroblastoma development and to investigate the possibility of <it>CADM1 </it>haploinsufficiency in neuroblastoma.</p

Instituto Nacional de Ciência e Tecnologia em Medicina Translacional (INCT-TM): abordagens metodológicas National Science and Technology Institute for Translational Medicine (INCT-TM): advancing the field of translational medicine and mental health

OBJETIVO: Medicina translacional pode ser descrita como a aplicação integrada de ferramentas farmacológicas inovadoras, biomarcadores, métodos e tecnologias clínicas e delineamentos de pesquisa para aumentar o conhecimento a respeito das doenças. A pesquisa translacional oferece uma oportunidade para aplicar os achados de pesquisa básica na clínica cotidiana. O Instituto Nacional de Ciência e Tecnologia - Medicina Translacional foi criado com seis centros (Universidade Federal do Rio Grande do Sul, Universidade de São Paulo-Ribeirão Preto, Universidade Federal do Rio de Janeiro, Pontifícia Universidade Católica do Rio Grande do Sul, Universidade Estadual de Santa Catarina e uma core facility que serve a todos os centros). Os objetivos deste projeto são divididos em quatro dimensões: institucional, pesquisa, formação de recursos humanos e transferência de tecnologia para a comunidade e setor produtivo. MÉTODO: Neste artigo, são apresentadas algumas das estratégias utilizadas para atingir os objetivos do Instituto Nacional de Ciência e Tecnologia - Medicina Translacional: 1) Desenvolvimento de modelos animais para o transtorno bipolar; 2) Estratégias de investigação neurocomportamental e disfunções cognitivas dos transtornos cerebrais; 3) Modelos experimentais de funcionamento cerebral e comportamento, proliferação celular e câncer; 4)Teste de Simulação de Falar em Público e 5) Realidade Virtual para indução de Pânico e Agorafobia. CONCLUSÃO: O Instituto Nacional de Ciência e Tecnologia - Medicina Translacional possui como foco principal o desenvolvimento de metodologias mais úteis para aumentar a aplicabilidade dos conhecimentos da pesquisa básica em medicina.<br>OBJECTIVE: Translational medicine has been described as the integrated application of innovative pharmacology tools, biomarkers, clinical methods, clinical technologies and study designs to improve the understanding of medical disorders. In medicine, translational research offers an opportunity for applying the findings obtained from basic research to every-day clinical applications. The National Science and Technology Institute for Translational Medicine is comprised of six member institutions (Universidade Federal do Rio Grande do Sul, Universidade de São Paulo-Ribeirão Preto, Universidade Federal do Rio de Janeiro, Pontifícia Universidade Católica do Rio Grande do Sul, Universidade Estadual de Santa Catarina and a core facility that serves all centers). The objectives of the project are divided into four areas: Institutional, Research, Human Resources and Technology for the Community and Productive Sector. METHOD: In this manuscript, we describe some of the approaches used to attain the main objectives of the National Science and Technology Institute for Translational Medicine, which include the development of 1) animal models for bipolar disorder; 2) strategies to investigate neurobehavioral function and cognitive dysfunction associated with brain disorders; 3) experimental models of brain function and behavior, neuropsychiatric disorders, cell proliferation, and cancer; 4) Simulated Public Speaking and 5) Virtual reality simulation for inducing panic disorder and agoraphobia. CONCLUSION: The main focus of the National Science and Technology Institute for Translational Medicine is the development of more useful methods that allow for a better application of basic research-based knowledge to the medical field

Collection and morphological characterization of sweet potato landraces in north of Rio de Janeiro state Coleta e caracterização morfológica de variedades locais de batata-doce no norte do Rio de Janeiro

The traditional farmers play an important role in plant genetic resources conservation. Collecting the germplasm maintained by these farmers is a very important action to avoid genetic variability losses. The goals of this work were to collect sweet potato from farms in the north of Rio de Janeiro state; to gather information regarding to the farmers profile, and to characterize the sweet potato landraces collected using morphological descriptors. Fifty three farms were visited in six collection expedition and 46 accessions were collected. During the visits the farmers were interviewed using a query with ten items. Six root traits and eight descriptors for vegetative parts were used for morphological characterization. The data were analyzed based on Cole-Rodgers distance and clustering was done with UPGMA method. Familiar agriculture with subsistence objective was observed and sweet potato was cultivated by 72% of the farmers at least for more than a decade, supporting the observation that this vegetable is traditionally cultivated in small areas in the specific region. The morphological characterization was efficient to detect genetic variability among accessions, revealing that traditional farmers from Campos dos Goytacazes and São João da Barra are responsible for sweet potato genotypes conservation with expressive genetic diversity in their properties. There was no relationship between genetic distance and collecting areas.<br>Os agricultores tradicionais têm um papel fundamental na conservação dos recursos genéticos vegetais e a coleta de germoplasma mantido por esses produtores é muito importante para evitar a perda da variabilidade genética. Os objetivos deste trabalho foram coletar germoplasma de batata-doce em propriedades rurais situadas no norte do estado do Rio de Janeiro; levantar informações quanto ao perfil dos produtores rurais visitados durante as coletas, e caracterizar morfologicamente as variedades locais de batata-doce coletadas. Em seis viagens de coleta foram visitadas 53 propriedades rurais, e coletados 46 acessos. Em todas as visitas foi possível entrevistar o produtor por meio de questionário contendo dez itens. Para a caracterização, foram usados seis caracteres de raiz e oito da parte aérea. A análise dos dados foi efetuada com o uso da distância de Cole-Rodgers e o agrupamento foi gerado com o método UPGMA. Foi constatado predomínio da agricultura familiar, voltada para subsistência, sendo que 72% dos agricultores cultivavam a batata-doce há mais de uma década, o que evidencia a tradição de cultivo desta hortaliça em pequenas áreas na região. A caracterização morfológica foi eficiente para detectar a variabilidade genética entre os acessos, demonstrando que os agricultores tradicionais dos municípios de Campos dos Goytacazes e São João da Barra detêm genótipos de batata-doce com expressiva diversidade genética em suas propriedades. Não foi detectada correlação entre a distância genética e os locais de coleta

Thermal decomposition of selected chlorinated hydrocarbons during gas combustion in fluidized bed

<p>Abstract</p> <p>Background</p> <p>The process of thermal decomposition of dichloromethane (DCM) and chlorobenzene (MCB) during the combustion in an inert, bubbling fluidized bed, supported by LPG as auxiliary fuel, have been studied. The concentration profiles of C₆H₅CI, CH₂Cl₂, CO₂, CO, NO_x, COCl₂, CHCl₃, CH₃Cl, C₂H₂, C₆H₆, CH₄ in the flue gases were specified versus mean bed temperature.</p> <p>Results</p> <p>The role of preheating of gaseous mixture in fluidized bed prior to its ignition inside bubbles was identified as important factor for increase the degree of conversion of DCM and MCB in low bed temperature, in comparison to similar process in the tubular reactor.</p> <p>Conclusions</p> <p>Taking into account possible combustion mechanisms, it was identified that autoignition in bubbles rather than flame propagation between bubbles is needed to achieve complete destruction of DCM and MCB. These condition occurs above 900°C causing the degree of conversion of chlorine compounds of 92-100%.</p