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J Proteome Res ; 9(5): 2658-68, 2010 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-20373825

RESUMO

The liver is the major site of xenobiotic metabolism and detoxification. Primary cultures of hepatocytes are a vital tool in the development of new therapeutic agents but their utility is hindered by the rapid loss of phenotype. Hepatocytes cultured in a sandwich of extracellular matrix protein maintain better hepatic function compared with cells cultured as a monolayer but a wide-ranging proteomics study of the differences in cultures has never been performed. We characterize the changing phenotype of rat hepatocytes in primary culture using iTRAQ proteomics and systems biology network analysis of the identified, significantly regulated, proteins. A total of 754 unique proteins were identified from 4 independent experiments. Of these, 413 proteins were common to at least 3 experiments and 328 proteins were identified in all experiments. Both culture systems displayed altered expression of many common proteins. Network analysis showed that the primary functions of these proteins were in metabolic pathways, immune responses and cytoskeleton remodelling. Monolayer cultures uniquely regulate proteins mapping to pathways of oxidative stress and cell migration, whereas sandwich culture affected translation regulation and apoptosis pathways. These experiments provide a detailed proteomics data set to direct further work into maintaining hepatic phenotype using cultured primary hepatocytes and stem cell derived hepatocyte-like cells.


Assuntos
Desdiferenciação Celular/fisiologia , Hepatócitos/citologia , Hepatócitos/metabolismo , Mapeamento de Peptídeos/métodos , Proteômica/métodos , Animais , Técnicas de Cultura de Células , Células Cultivadas , Análise por Conglomerados , Colágeno/metabolismo , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Combinação de Medicamentos , Regulação da Expressão Gênica , Histocitoquímica , Marcação por Isótopo/métodos , Laminina/metabolismo , Masculino , Proteínas/genética , Proteínas/metabolismo , Proteoglicanas/metabolismo , Ratos , Ratos Wistar , Transdução de Sinais , Biologia de Sistemas
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