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Identification of Suitable Reference Genes for Quantitative Gene Expression Analysis in Innervated and Denervated Adipose Tissue from Cafeteria Diet-Fed Rats.
Marques-Oliveira, Gleuber Henrique; Silva, Thaís Marques; Valadares, Helder Magno Silva; Raposo, Helena Fonseca; Carolino, Ruither de Oliveira Gomes; Garófalo, Maria Antonieta Rissato; Anselmo-Franci, Janete Aparecida; do Carmo Kettelhut, Isis; de Oliveira, Helena Coutinho Franco; Chaves, Valéria Ernestânia.
Afiliación
  • Marques-Oliveira GH; Laboratory of Physiology, Federal University of São João del-Rei, Avenue Sebastião Gonçalves Coelho, 400, 35.501-296, Divinópolis, Minas Gerais, Brazil.
  • Silva TM; Laboratory of Physiology, Federal University of São João del-Rei, Avenue Sebastião Gonçalves Coelho, 400, 35.501-296, Divinópolis, Minas Gerais, Brazil.
  • Valadares HMS; Laboratory of Molecular Genetic, Federal University of São João del-Rei, Avenue Sebastião Gonçalves Coelho, 400, 35.501-296, Divinópolis, Minas Gerais, Brazil.
  • Raposo HF; Department of Structural and Functional Biology, Institute of Biology, State University of Campinas, Monteiro Lobato Street, 255, 13.083-862, Campinas, São Paulo, Brazil.
  • Carolino ROG; Department of Morphology, Physiology and Basic Pathology, Ribeirão Preto Dentristy School, University of São Paulo, Avenue of Café s/n, 14.040-904, Ribeirão Preto, São Paulo, Brazil.
  • Garófalo MAR; Department of Physiology, Ribeirão Preto Medical School, University of São Paulo, Avenue of Café s/n, 14.040-904, Ribeirão Preto, São Paulo, Brazil.
  • Anselmo-Franci JA; Department of Morphology, Physiology and Basic Pathology, Ribeirão Preto Dentristy School, University of São Paulo, Avenue of Café s/n, 14.040-904, Ribeirão Preto, São Paulo, Brazil.
  • do Carmo Kettelhut I; Departments of Biochemistry-Immunology, Ribeirão Preto Medical School, University of São Paulo, Avenueof Café s/n, 14.040-904, Ribeirão Preto, São Paulo, Brazil.
  • de Oliveira HCF; Department of Structural and Functional Biology, Institute of Biology, State University of Campinas, Monteiro Lobato Street, 255, 13.083-862, Campinas, São Paulo, Brazil.
  • Chaves VE; Laboratory of Physiology, Federal University of São João del-Rei, Avenue Sebastião Gonçalves Coelho, 400, 35.501-296, Divinópolis, Minas Gerais, Brazil.
Lipids ; 54(4): 231-244, 2019 04.
Article en En | MEDLINE | ID: mdl-31025715
Our previous studies show that cafeteria diet increases body adiposity, plasma insulin levels, and sympathetic activity to brown adipose tissue (BAT) and white adipose tissue (WAT) of Wistar rats, leading to rapid and progressive changes in the metabolic profile. The identification of suitable reference genes that are not affected by the experimental conditions is a critical step in accurate normalization of the reverse transcription quantitative real-time PCR (qRT-PCR), a commonly used assay to elucidate changes in the gene expression profile. In the present study, the effects of the cafeteria diet and sympathetic innervation on the gene expression of adrenoceptor beta 3 (Adrb3) from BAT and WAT were assessed using one of the most stable and one of the least stable genes as normalizers. Rats were fed the cafeteria diet and on the 17th day, interscapular BAT or retroperitoneal WAT was denervated and, 7 days after surgery, the contralateral innervated tissue was used as control. Ten reference genes were evaluated (18S, B2m, Actb, CypA, Gapdh, Hprt1, Rpl32, Tbp, Ubc, and Ywhaz) and ranked according to their stability using the following algorithms: geNorm, NormFinder, BestKeeper, and comparative delta threshold cycle (ΔC t ) method. According to the algorithms employed, the normalization of Adrb3 expression by the least stable genes produced opposite results compared with the most stable genes and literature data. In cafeteria and control diet-fed rats, the three most stable genes were Hprt1, Tbp, and Rpl32 for interscapular BAT and Tbp, B2m, and Hprt1 for retroperitoneal WAT, while the least stable genes were 18S, Actb, and Gapdh for both tissues.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Tejido Adiposo Pardo / Dieta / Tejido Adiposo Blanco Tipo de estudio: Diagnostic_studies Límite: Animals Idioma: En Revista: Lipids Año: 2019 Tipo del documento: Article País de afiliación: Brasil Pais de publicación: Estados Unidos
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Tejido Adiposo Pardo / Dieta / Tejido Adiposo Blanco Tipo de estudio: Diagnostic_studies Límite: Animals Idioma: En Revista: Lipids Año: 2019 Tipo del documento: Article País de afiliación: Brasil Pais de publicación: Estados Unidos