RESUMO
BACKGROUND/AIMS: The direct implication of low-density lipoprotein receptor (LDLR) in hepatitis C virus (HCV) infection of human hepatocyte has not been demonstrated. Normal primary human hepatocytes infected by serum HCV were used to document this point. METHODS: Expression and activity of LDLR were assessed by RT-PCR and LDL entry, in the absence or presence of squalestatin or 25-hydroxycholesterol that up- or down-regulates LDLR expression, respectively. Infection was performed in the absence or presence of LDL, HDL, recombinant soluble LDLR peptides encompassing full-length (r-shLDLR4-292) or truncated (r-shLDLR4-166) LDL-binding domain, monoclonal antibodies against r-shLDLR4-292, squalestatin or 25-hydroxycholesterol. Intracellular amounts of replicative and genomic HCV RNA strands used as end point of infection were assessed by RT-PCR. RESULTS: r-shLDLR4-292, antibodies against r-shLDLR4-292 and LDL inhibited viral RNA accumulation, irrespective of genotype, viral load or liver donor. Inhibition was greatest when r-shLDLR4-292 was present at the time of inoculation and gradually decreased as the delay between inoculation and r-shLDLR4-292 treatment increased. In hepatocytes pre-treated with squalestatin or 25-hydroxycholesterol before infection, viral RNA accumulation increased or decreased in parallel with LDLR mRNA expression and LDL entry. CONCLUSIONS: LDLR is involved at an early stage in infection of normal human hepatocytes by serum-derived HCV virions.
Assuntos
Hepacivirus/patogenicidade , Hepatite C/fisiopatologia , Hepatócitos/virologia , Receptores de LDL/fisiologia , Adolescente , Adulto , Idoso , Anticorpos/fisiologia , Anticolesterolemiantes/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Antígenos CD18/fisiologia , Células Cultivadas , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Hepacivirus/genética , Hepacivirus/fisiologia , Hepatite C/patologia , Hepatócitos/patologia , Humanos , Hidroxicolesteróis/farmacologia , Lipoproteínas HDL/fisiologia , Lipoproteínas LDL/fisiologia , Masculino , Pessoa de Meia-Idade , RNA Viral/genética , RNA Viral/metabolismo , Receptores de LDL/genética , Receptores de LDL/imunologia , Receptores Depuradores Classe B/fisiologia , Ácidos Tricarboxílicos/farmacologia , Carga Viral , VírionRESUMO
Different diseases of the CNS are associated with blood-brain barrier (BBB) damage and mononuclear cell infiltration. In order to study genes that may play a role in endothelial cell regulation in inflammatory CNS diseases, we performed differential gene expression (DGE) analysis using a mouse brain endothelial cell line. We found that interferon-gamma (IFNgamma)-induced monokine (MIG), a chemokine that plays a role in T lymphocyte and monocyte chemoattraction, is highly expressed in the presence of inflammatory cytokines. We show that MIG, produced by brain endothelial cells in vitro, is biologically active in attractingT lymphocytes and that it is possible to interfere with this mechanism of action using anti-MIG antibodies. We suggest that blocking MIG may be beneficial in CNS inflammation. We detected constitutive expression of the MIG receptor, CXCR3, on the surface of the endothelial cells and therefore hypothesize that it plays a role in maintaining the cytokine gradient at the region of CNS inflammation.
