RESUMO
UNLABELLED: Both hepatitis B and C viruses frequently establish chronic infection, raising the question whether T cells are poorly primed in the liver. To determine the role of different cell types in the activation of CD8+ T cells against hepatocellular antigens, we used an Adeno-associated virus to deliver ovalbumin to hepatocytes. In contrast to CD8+ T cells, CD4+ T cells were not activated. The CD8+ T cells were activated even in the absence of endogenous CD4+ T cells; however, in the liver, these cells were high in the programmed death-1 protein and low in CD127. Chimera experiments revealed that these CD8+ T cells were activated on a solid tissue cell. CONCLUSION: Priming of CD8+ T cells directly on nonhematopoietic cells, in the absence of CD4+ T cell help, results in suboptimal T cell activation. This could explain the impaired function of CD8+ T cells seen in chronic liver infection.
Assuntos
Adenoviridae/fisiologia , Linfócitos T CD8-Positivos/patologia , Hepatócitos/patologia , Hepatócitos/virologia , Fígado/patologia , Animais , Células Apresentadoras de Antígenos/patologia , Antígenos de Diferenciação/metabolismo , Linfócitos T CD4-Positivos/patologia , Células Dendríticas/patologia , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Receptor de Morte Celular Programada 1RESUMO
We aim to define the role of Kupffer cells in intrahepatic antigen presentation, using the selective delivery of antigen to Kupffer cells rather than other populations of liver antigen-presenting cells. To achieve this we developed a novel antigen delivery system that can target antigens to macrophages, based on a galactosylated low-density lipoprotein nanoscale platform. Antigen was delivered via the galactose particle receptor (GPr), internalized, degraded and presented to T cells. The conjugation of fluoresceinated ovalbumin (FLUO-OVA) and lactobionic acid with LDL resulted in a substantially increased uptake of FLUO-OVA by murine macrophage-like ANA1 cells in preference to NIH3T3 cells, and by primary peritoneal macrophages in preference to primary hepatic stellate cells. Such preferential uptake led to enhanced proliferation of OVA specific T cells, showing that the galactosylated LDL nanoscale platform is a successful antigen carrier, targeting antigen to macrophages but not to all categories of antigen presenting cells. This system will allow targeted delivery of antigen to macrophages in the liver and elsewhere, addressing the question of the role of Kupffer cells in liver immunology. It may also be an effective way of delivering drugs or vaccines directly at macrophages.
Assuntos
Antígenos/administração & dosagem , Lipoproteínas LDL/química , Nanopartículas/química , Animais , Apresentação de Antígeno , Linhagem Celular , Dissacarídeos , Sistemas de Liberação de Medicamentos , Endocitose , Feminino , Produtos Finais de Glicação Avançada , Células Estreladas do Fígado/imunologia , Células Estreladas do Fígado/metabolismo , Técnicas In Vitro , Células de Kupffer/imunologia , Células de Kupffer/metabolismo , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Biológicos , Células NIH 3T3 , Nanopartículas/toxicidade , Nanopartículas/ultraestrutura , Nanotecnologia , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , Receptores de Superfície Celular/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Kupffer cells form a large intravascular macrophage bed in the liver sinusoids. The differentiation history and diversity of Kupffer cells is disputed; some studies argue that they are derived from blood monocytes, whereas others support a local origin from intrahepatic precursor cells. In the present study, we used both flow cytometry and immunohistochemistry to distinguish 2 subsets of Kupffer cells that were revealed in the context both of bone marrow transplantation and of orthotopic liver transplantation. One subset was radiosensitive and rapidly replaced from hematogenous precursors, whereas the other was relatively radioresistant and long-lived. Both were phagocytic but only the former population was recruited into inflammatory foci in response to CD8(+) T-cell activation. We propose the name "sessile" for the radioresistant Kupffer cells that do not participate in immunoinflammatory reactions. However, we found no evidence that these sessile Kupffer cells arise from immature intrahepatic precursors. Our conclusions resolve a long-standing controversy and explain how different experimental approaches may reveal one or both of these subsets.
Assuntos
Diferenciação Celular , Células-Tronco Hematopoéticas/citologia , Células de Kupffer/citologia , Fígado/citologia , Macrófagos/citologia , Monócitos/citologia , Animais , Transplante de Medula Óssea , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/imunologia , Diferenciação Celular/efeitos da radiação , Células-Tronco Hematopoéticas/imunologia , Inflamação/imunologia , Células de Kupffer/imunologia , Fígado/imunologia , Transplante de Fígado , Ativação Linfocitária/imunologia , Ativação Linfocitária/efeitos da radiação , Ativação de Macrófagos/efeitos da radiação , Macrófagos/imunologia , Camundongos , Monócitos/imunologia , Irradiação Corporal TotalRESUMO
The response of T cells to liver Ags sometimes results in immune tolerance. This has been proposed to result from local, intrahepatic priming, while the expression of the same Ag in liver-draining lymph nodes is believed to result in effective immunity. We tested this model, using an exogenous model Ag expressed only in hepatocytes, due to infection with an adeno-associated virus vector. T cell activation was exclusively intrahepatic, yet in contrast to the predictions of the current model, this resulted in clonal expansion, IFN-gamma synthesis, and cytotoxic effector function. Local activation of naive CD8(+) T cells can therefore cause full CD8(+) T cell activation, and hepatocellular presentation cannot be used to explain the failure of CTL effector function against some liver pathogens such as hepatitis C.
Assuntos
Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/imunologia , Proteínas de Fluorescência Verde/imunologia , Hepatócitos/citologia , Hepatócitos/imunologia , Ativação Linfocitária/imunologia , Ovalbumina/imunologia , Transferência Adotiva , Animais , Biomarcadores/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/transplante , Linhagem Celular , Proliferação de Células , Técnicas de Cocultura , Citotoxicidade Imunológica/genética , Dependovirus/imunologia , Vetores Genéticos , Receptores de Hialuronatos/biossíntese , Selectina L/biossíntese , Ativação Linfocitária/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Linfócitos T Citotóxicos/imunologiaRESUMO
The synthesis of the common and well-documented Siaalpha 2,6 to Galbeta 1,4GlcNAc structure (Sia6LacNAc) is principally mediated by the sialyltransferase ST6Gal I, which is particularly highly expressed in liver, lactating mammary gland, intestinal epithelia of newborn animals, and B cells. Multiple independent promoters govern the expression of Siat1, the ST6Gal I gene. In liver, elevation of hepatic and serum ST6Gal is part of the acute phase reaction, the hepatic response to systemic trauma, and is governed by the inducible, liver-specific promoter-regulatory region, P1. A constitutive and nontissue-specific promoter, P3, mediates low-level, basal hepatic Siat1 transcription. We generated a mouse specifically unable to use the transcriptional initiation site uniquely used in P1-mediated ST6Gal I expression. These animals, Siat1deltaP1, are viable and display reduced ST6Gal I mRNA in liver with concomitantly reduced sialyltransferase activities in liver and in serum. Siat1deltaP1 animals are unable to elevate hepatic Siat1 mRNA as part of the inflammatory response induced by turpentine. Surprisingly, serum glycoprotein components exhibit normal extent of sialylation, with no noticeable difference in binding to SNA, the alpha2,6-sialyl-specific lectin. Siat1deltaP1 animals also exhibit an outwardly normal B cell response. On intraperitoneal challenge with the pathogen Salmonella typhimurium, a significantly greater accumulation of neutrophils within the peritoneal space was observed in Siat1deltaP1 animals compared to wild-type mice. Siat1deltaP1 mice also exhibit a greater bacterial burden in liver and spleen, accompanied by more pronounced spleno-/hepatomegaly and greater leukocyte infiltration into affected organs than their wild-type counterparts.