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J Virol ; 82(17): 8733-42, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18579610

RESUMO

Hepatitis A virus (HAV) is an enterically transmitted virus that replicates predominantly in hepatocytes within the liver before excretion via bile through feces. Hepatocytes are polarized epithelial cells, and it has been assumed that the virus load in bile results from direct export of HAV via the apical domain of polarized hepatocytes. We have developed a subclone of hepatocyte-derived HepG2 cells (clone N6) that maintains functional characteristics of polarized hepatocytes but displays morphology typical of columnar epithelial cells, rather than the complex morphology that is typical of hepatocytes. N6 cells form microcolonies of polarized cells when grown on glass and confluent monolayers of polarized cells on semipermeable membranes. When N6 microcolonies were exposed to HAV, infection was restricted to peripheral cells of polarized colonies, whereas all cells could be infected in colonies of nonpolarized HepG2 cells (clone C11) or following disruption of tight junctions in N6 colonies with EGTA. This suggests that viral entry occurs predominantly via the basolateral plasma membrane, consistent with uptake of virus from the bloodstream after enteric exposure, as expected. Viral export was also found to be markedly vectorial in N6 but not C11 cells. However, rather than being exported from the apical domain as expected, more than 95% of HAV was exported via the basolateral domain of N6 cells, suggesting that virus is first excreted from infected hepatocytes into the bloodstream rather than to the biliary tree. Enteric excretion of HAV may therefore rely on reuptake and transcytosis of progeny HAV across hepatocytes into the bile. These studies provide the first example of the interactions between viruses and polarized hepatocytes.


Assuntos
Polaridade Celular , Endocitose/fisiologia , Vírus da Hepatite A/fisiologia , Hepatócitos/virologia , Animais , Caliciviridae/crescimento & desenvolvimento , Carcinoma Hepatocelular/patologia , Gatos , Técnicas de Cultura de Células , Linhagem Celular , Linhagem Celular Tumoral , Células Cultivadas , Chlorocebus aethiops , Células Clonais , Meios de Cultura Livres de Soro , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Técnica Indireta de Fluorescência para Anticorpo , Hepatite A/transmissão , Hepatite A/virologia , Vírus da Hepatite A/crescimento & desenvolvimento , Vírus da Hepatite A/isolamento & purificação , Hepatócitos/metabolismo , Hepatócitos/ultraestrutura , Humanos , Rim/citologia , Neoplasias Hepáticas/patologia , Proteínas de Membrana/metabolismo , Proteínas de Membrana/ultraestrutura , Microvilosidades/ultraestrutura , Fosfoproteínas/metabolismo , Fosfoproteínas/ultraestrutura , Junções Íntimas/ultraestrutura , Ensaio de Placa Viral , Proteína da Zônula de Oclusão-1
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