Advertisement

Liver antigen-presenting cells

  • Ian Nicholas Crispe
    Correspondence
    Address: Malaria Program, Seattle BioMed, 307 North Westlake Avenue, Suite 500, Seattle, WA 98109, USA. Tel.: +1 206 256 7163; fax: +1 206 256 7229.
    Affiliations
    Malaria Program, Seattle BioMed, Seattle, WA, USA
    Department of Immunology, University of Washington, Seattle WA, USA
    Search for articles by this author
Open AccessPublished:October 29, 2010DOI:https://doi.org/10.1016/j.jhep.2010.10.005
      The liver is an organ in which several major pathogens evade immune clearance and achieve chronicity. How do they do it? Recent research has documented multiple mechanisms by which immune responses in the liver are biased towards tolerance. In this review, the induction of local, intrahepatic tolerance is explored from the perspective of antigen presentation. Experiments support the role not only of liver dendritic cell subsets but also of diverse subsets of unconventional antigen-presenting cells in inducing immune suppression. The literature on this topic is controversial and sometimes contradictory, making it difficult to formulate a unified model of antigen handling and T cell priming in the liver. Here I offer a critical review of the state of the art in understanding antigen presentation in the liver.

      Abbreviations:

      IFN (Interferon), MHC (Major Histocompatibility Complex), LSEC (liver sinusoidal endothelial cells), APC (antigen-presenting cells), DC (dendritic cells), FoxP3 (Forkhead transcription factor-P3), LPS (lipopolysaccharide endotoxin), NK (natural killer), ICAM-1 (Intercellular Cell Adhesion Molecule-1), VCAM-1 (Vascular Cell Adhesion Molecule-1), CXCR6 (C-X-C chemokine receptor-6), TLR (Toll-like receptor), mDC (myeloid DC), pDC (plasmacytoid DC), BDCA (blood dendritic cell antigen), B220 (B cell isoform of CD45 of mass 220kd), LDL (low-density lipoproteins), HUVEC (human umbilical vein endothelial cells), Tie-2 (endothelium-specific receptor tyrosine kinase type-2)

      Keywords

      Introduction

      The liver is the site of several infections of major importance, against which the immune system normally delivers either an ineffective or a pathogenic response. In the case of Hepatitis B and Hepatitis C, immune responses occur but they are frequently ineffective. With the lack of virus elimination, chronic immune responses cause cumulative tissue damage and eventual fibrosis, leading to disruption of the liver’s hemodynamics, and the loss of liver function. In contrast, malaria parasites migrate through the liver and undergo an essential part of their maturation there, yet there is no evidence of an endogenous immune response. While much of the understanding of human liver immunology is based on the study of immune responses to viral hepatitis, the world’s most prevalent serious infection, malaria, is also a liver pathogen. Most species of murine, and all human malaria parasites undergo an obligatory developmental stage in the liver. The sporozoite, introduced by the bite of an infected mosquito, interacts sequentially with the liver sinusoidal endothelial cells [
      • Pradel G.
      • Garapaty S.
      • Frevert U.
      Proteoglycans mediate malaria sporozoite targeting to the liver.
      ] and Kupffer cells [
      • Pradel G.
      • Frevert U.
      Malaria sporozoites actively enter and pass through rat Kupffer cells prior to hepatocyte invasion.
      ]. The Kupffer cells appear to be an essential “gateway” through which sporozoites penetrate the endothelial barrier and enter the hepatocytes [
      • Baer K.
      • Roosevelt M.
      • Clarkson Jr., A.B.
      • van Rooijen N.
      • Schnieder T.
      • Frevert U.
      Kupffer cells are obligatory for Plasmodium yoelii sporozoite infection of the liver.
      ]. Once in the hepatocytes, the parasites develop rapidly over several days, after which the host cell dies and merozoites are released, which in turn parasitize red blood cells. The liver stage is an attractive vaccine target, and genetically modified murine malaria parasites create sterilizing immunity that appears to intercept the infection at the liver stage [
      • Mueller A.K.
      • Labaied M.
      • Kappe S.H.
      • Matuschewski K.
      Genetically modified Plasmodium parasites as a protective experimental malaria vaccine.
      ]. The mechanism of action of the vaccine is not understood, but it appears to depend on Interferon (IFN)-γ, and on CD8+ T cells [
      • Jobe O.
      • Lumsden J.
      • Mueller A.K.
      • Williams J.
      • Silva-Rivera H.
      • Kappe S.H.
      • et al.
      Genetically attenuated Plasmodium berghei liver stages induce sterile protracted protection that is mediated by major histocompatibility complex Class I-dependent interferon-γ-producing CD8+ T cells.
      ].
      In many mammalian species, the transplantation of the liver across a Major Histocompatibility Complex (MHC) difference does not result in rejection [
      • Calne R.Y.
      Immunological tolerance – the liver effect.
      ,
      • Calne R.Y.
      • White H.J.
      • Yoffa D.E.
      • Binns R.M.
      • Maginn R.R.
      • Herbertson R.M.
      • et al.
      Prolonged survival of liver transplants in the pig.
      ]. This stands in contrast to the consequences of transplanting kidneys, skin, pancreas or other organs, where rejection is the usual outcome. In addition, the transplanted liver is able to confer tolerance on another solid organ transplant from the same donor, arguing that the liver can induce systemic tolerance [
      • Calne R.Y.
      • Sells R.A.
      • Pena J.R.
      • Davis D.R.
      • Millard P.R.
      • Herbertson B.M.
      • et al.
      Induction of immunological tolerance by porcine liver allografts.
      ]. This effect is not fully understood, but it has been attributed to: the effects of liver-derived APC dispersed throughout the host, also known as microchimerism [
      • Qian S.
      • Demetris A.J.
      • Murase N.
      • Rao A.S.
      • Fung J.J.
      • Starzl T.E.
      Murine liver allograft transplantation: tolerance and donor cell chimerism.
      ]; the effects of Kupffer cells or liver sinusoidal endothelial cells (LSEC) as antigen-presenting cells (APC), promoting tolerance [
      • Knolle P.A.
      • Gerken G.
      Local control of the immune response in the liver.
      ,
      • Sato K.
      • Yabuki K.
      • Haba T.
      • Maekawa T.
      Role of Kupffer cells in the induction of tolerance after liver transplantation.
      ]; the distinctive properties of liver-resident dendritic cells (DC) [
      • Mazariegos G.V.
      • Zahorchak A.F.
      • Reyes J.
      • Chapman H.
      • Zeevi A.
      • Thomson A.W.
      Dendritic cell subset ratio in tolerant, weaning and non-tolerant liver recipients is not affected by extent of immunosuppression.
      ]; and the induction of allospecific regulatory T cells of the CD4+, CD25+, Forkhead transcription factor-P3 (FoxP3)+ type [
      • Li W.
      • Kuhr C.S.
      • Zheng X.X.
      • Carper K.
      • Thomson A.W.
      • Reyes J.D.
      • et al.
      New insights into mechanisms of spontaneous liver transplant tolerance: the role of Foxp3-expressing CD25+CD4+ regulatory T cells.
      ]. Despite the well-documented liver allograft tolerance in many animal models, human liver transplants are undertaken with the use of immunosuppressive drugs. In the context of a pathogen that re-infects the liver allograft, such as Hepatitis C Virus, this situation leads to rapid progression of the infection [
      • McCaughan G.W.
      • Zekry A.
      Impact of immunosuppression on immunopathogenesis of liver damage in hepatitis C virus-infected recipients following liver transplantation.
      ].
      Figure thumbnail fx1

      The hepatic vasculature and leukocyte trafficking

      The liver is a major focus of metabolic activity, where the products of digestion are processed, plasma proteins synthesized, and dangerous foreign chemicals detoxified. To serve these functions, the liver receives its blood supply from two sources: around 20% of the blood is arterial, delivered via the hepatic artery which branches off from the celiac axis; while the other 80% originates in the intestine. This portal venous blood carries to the liver a mixture of antigens from food and bacterial products from the intestinal bacteria. In particular, the portal blood carries lipopolysaccharide endotoxin (LPS) at concentrations of up to 1 ng/ml [
      • Freudenberg M.A.
      • Freudenberg N.
      • Galanos C.
      Time course of cellular distribution of endotoxin in liver, lungs and kidneys of rats.
      ,
      • Lumsden A.B.
      • Henderson J.M.
      • Kutner M.H.
      Endotoxin levels measured by a chromogenic assay in portal, hepatic and peripheral venous blood in patients with cirrhosis.
      ]. Thus, in the liver, both antigen-specific lymphocyte receptors and pattern recognition receptors are exposed to their ligands.
      The liver contains a diverse population of both adaptive and innate immune cells. T cells are abundant, with a bias towards CD8+ T cells, and activated T cells predominate [
      • Crispe I.N.
      • Mehal W.Z.
      Strange brew: T cells in the liver.
      ,
      • Tu Z.
      • Bozorgzadeh A.
      • Crispe I.N.
      • Orloff M.S.
      The activation state of human intrahepatic lymphocytes.
      ]. Natural killer (NK) cells are abundant, and these cells similarly express activation markers [
      • Tu Z.
      • Bozorgzadeh A.
      • Crispe I.N.
      • Orloff M.S.
      The activation state of human intrahepatic lymphocytes.
      ]. NK-T cells are more frequent in the liver than in the blood in humans, and more frequent than in the lymphoid organs in mice; this holds true whether these cells are defined expansively as NK1.1+ T cells in the mouse, or CD56+ T cells in the human, or CD1d-reactive cells, or narrowly defined as T cells that bind tetramers of a glycolipid, α-galactosyl ceramide, associated with a CD1d molecule [
      • Baron J.L.
      • Gardiner L.
      • Nishimura S.
      • Shinkai K.
      • Locksley R.
      • Ganem D.
      Activation of a nonclassical NKT cell subset in a transgenic mouse model of hepatitis B virus infection.
      ,
      • Norris S.
      • Collins C.
      • Doherty D.G.
      • Smith F.
      • McEntee G.
      • Traynor O.
      • et al.
      Resident human hepatic lymphocytes are phenotypically different from circulating lymphocytes.
      ]. Lymphocytes with exactly these features can be eluted from the hepatic vasculature of a human liver lobe prior to transplant [
      • Tu Z.
      • Bozorgzadeh A.
      • Crispe I.N.
      • Orloff M.S.
      The activation state of human intrahepatic lymphocytes.
      ], suggesting that they are found in the lumen of the blood vessels, and immunohistology similarly reveals individual T cells through the normal human liver parenchyma, as well as in portal tracts [
      • Smith F.
      • Golden-Mason L.
      • Deignan T.
      • Norris S.
      • Nolan N.
      • Traynor O.
      • et al.
      Localization of T and B lymphocytes in histologically normal adult human donor liver.
      ]. For most of these cell populations, it is not possible to say how far these cells are long-term hepatic residents, and how far they are preferentially slowed down in the liver during their recirculation by adhesion molecules on the hepatic endothelium. The liver has the capacity to preferentially sequester activated CD8+ T cells from the circulation [
      • Mehal W.Z.
      • Juedes A.E.
      • Crispe I.N.
      Selective retention of activated CD8+ T cells by the normal liver.
      ], and this effect depends in part on Intercellular Cell Adhesion Molecule-1 (ICAM-1) and Vascular Cell Adhesion Molecule-1 (VCAM-1) expressed on the hepatic vasculature [
      • John B.
      • Crispe I.N.
      Passive and active mechanisms trap activated CD8+ T cells in the liver.
      ]. However, in the specific case of NK-T cells, in vivo microscopy was used to identify these cells in the living liver, exploiting their expression of CXCR6 and a knock-in strategy to render them fluorescent. These cells were observed patrolling the hepatic sinusoids, both with and against the direction of blood flow [
      • Geissmann F.
      • Cameron T.O.
      • Sidobre S.
      • Manlongat N.
      • Kronenberg M.
      • Briskin M.J.
      • et al.
      Intravascular immune surveillance by CXCR6+ NKT cells patrolling liver sinusoids.
      ]. Activation of these cells causes them to stop patrolling, consistent with them having a defensive function [
      • Velazquez P.
      • Cameron T.O.
      • Kinjo Y.
      • Nagarajan N.
      • Kronenberg M.
      • Dustin M.L.
      Cutting edge: activation by innate cytokines or microbial antigens can cause arrest of natural killer T cell patrolling of liver sinusoids.
      ]. These cells, at least, were not passively drifting through the liver, and are likely to be long-term residents.
      Blood percolates through the liver in thin-walled vessels termed sinusoids, the endothelium of which is penetrated by small holes (fenestrations) grouped in clusters (sieve plates). The fenestrations are large enough to permit contact between lymphocytes in the blood space, and the underlying hepatocytes (Fig. 1). Electron micrographs show contact between T cell microvilli and their counterparts on the hepatocytes [
      • Warren A.
      • Le Couteur D.G.
      • Fraser R.
      • Bowen D.G.
      • McCaughan G.W.
      • Bertolino P.
      T lymphocytes interact with hepatocytes through fenestrations in murine liver sinusoidal endothelial cells.
      ], though the physiological significance of such interaction is not clear. Certainly, these contacts are not sufficient to allow the formation of an immunological synapse, but in the living sinusoid they may act as initiators of more intimate contact. This progression has not yet been observed directly. The flow of blood is slow, due to the large cross-sectional area of the sinusoidal bed, and this is likely to facilitate interactions with both intra-sinusoidal and peri-sinusoidal cells. Some electron micrographs reveal gaps in the endothelial layer, but it is likely that in the living sinusoid, the liver’s resident macrophage population, Kupffer cells, occupies these gaps. We know this partly because elimination of the Kupffer cells using toxic liposomes results in gaps in the endothelial barrier, through which malaria sporozoites gain easy access to hepatocytes, bypassing their usual route through Kupffer cells [
      • Baer K.
      • Roosevelt M.
      • Clarkson Jr., A.B.
      • van Rooijen N.
      • Schnieder T.
      • Frevert U.
      Kupffer cells are obligatory for Plasmodium yoelii sporozoite infection of the liver.
      ].
      Figure thumbnail gr1
      Fig. 1Immunological players in the hepatic sinusoid. The liver sinusoidal endothelial cells are penetrated by holes (fenestrations), through which a CD8+ T cell can make direct contact with an underlying hepatocyte. Between the endothelial cells and the hepatocytes is the Space of Disse, in which reside hepatic stellate cells (Ito cells), a specialized pericyte with immunological properties. These cells respond to TLR ligands and synthesize chemokines, and they may also act as antigen-presenting cells, particularly for CD1d-restricted NK-T cells.

      Resident and transient myeloid cells

      The liver’s large macrophage population, also known as Kupffer cells, is unusual in that a large fraction of these cells are radio-resistant and difficult to extract from tissue, even after collagenase digestion. These sessile Kupffer cells are nevertheless phagocytic, but they do not migrate to inflammatory foci in the liver, which are formed by incoming, blood-derived cells [
      • Klein I.
      • Cornejo J.C.
      • Polakos N.K.
      • John B.
      • Wuensch S.A.
      • Topham D.J.
      • et al.
      Kupffer cell heterogeneity: functional properties of bone marrow derived and sessile hepatic macrophages.
      ]. In these features, the sessile Kupffer cells resemble microglia in the brain [
      • Kennedy D.W.
      • Abkowitz J.L.
      Kinetics of central nervous system microglial and macrophage engraftment: analysis using a transgenic bone marrow transplantation model.
      ]. Kupffer cells have some credentials as APC, but the balance of evidence suggests that they commonly promote T cell tolerance. Thus, Kupffer cells stimulated with LPS secrete the immunosuppressive cytokine, IL-10 [
      • Knolle P.
      • Schlaak J.
      • Uhrig A.
      • Kempf P.
      • Meyer zum Buschenfelde K.H.
      • Gerken G.
      Human Kupffer cells secrete IL-10 in response to lipopolysaccharide (LPS) challenge.
      ], and secrete the immunosuppressive prostaglandin E2 under metabolic stress [
      • Callery M.P.
      • Mangino M.J.
      • Flye M.W.
      Arginine-specific suppression of mixed lymphocyte culture reactivity by Kupffer cells – a basis of portal venous tolerance.
      ]. Kupffer cells express MHC class I and MHC class II, as well as co-stimulatory molecules at low density [
      • You Q.
      • Cheng L.
      • Kedl R.M.
      • Ju C.
      Mechanism of T cell tolerance induction by murine hepatic Kupffer cells.
      ], and can induce T cell activation. However, in mixed cultures they suppressed T cell activation induced by DC, and in this study, prostaglandins were key to the immunosuppressive effect, rather than IL-10. In contrast, human Kupffer cells activated through Toll-like receptor-2 (TLR2) and TLR4 ligation, synthesized IL-10, and in this case the IL-10 suppressed IL-18-dependent NK cell activation [
      • Tu Z.
      • Bozorgzadeh A.
      • Pierce R.H.
      • Kurtis J.
      • Crispe I.N.
      • Orloff M.S.
      TLR-dependent cross talk between human Kupffer cells and NK cells.
      ]. The capacity of Kupffer cells to act as APC can clearly be modulated by innate signals, since both reactive oxygen species and TLR3 ligation increased the expression of MHC class II and promoted APC function [
      • Maemura K.
      • Zheng Q.
      • Wada T.
      • Ozaki M.
      • Takao S.
      • Aikou T.
      • et al.
      Reactive oxygen species are essential mediators in antigen presentation by Kupffer cells.
      ,
      • You Q.
      • Cheng L.
      • Kedl R.M.
      • Ju C.
      Mechanism of T cell tolerance induction by murine hepatic Kupffer cells.
      ]. This is particularly provocative, since TLR3 may sometimes make the critical difference between liver tolerance and immunity [
      • Lang K.S.
      • Georgiev P.
      • Recher M.
      • Navarini A.A.
      • Bergthaler A.
      • Heikenwalder M.
      • et al.
      Immunoprivileged status of the liver is controlled by Toll-like receptor 3 signaling.
      ]. In the case of the spirochaete bacterium Borellia burgdorferi, phagocytosis of the bacterium by Kupffer cells led to CXCR3 chemokine secretion and recruitment of iNKT cells, which contributed to containment of the infection [
      • Lee W.Y.
      • Moriarty T.J.
      • Wong C.H.
      • Zhou H.
      • Strieter R.M.
      • van Rooijen N.
      • et al.
      An intravascular immune response to Borrelia burgdorferi involves Kupffer cells and iNKT cells.
      ]. Kupffer cells can therefore switch their immunological role in two senses: from inactivators to activators of NK cells, and from tolerance-inducing APC to immunogenic APC. In both cases, their dominant action appears to depend on innate immune signals, particularly on LPS (Fig. 2).
      Figure thumbnail gr2
      Fig. 2CD4+ T cells that are primed in the hepatic sinusoids receive mixed messages. Pointed arrows indicate activating signals, while flat-ended arrows indicate suppressive signals. Both Kupffer cells and liver sinusoidal endothelial cells (LSEC) take up antigen from the blood and present it to CD4+ T cells together with co-stimulatory molecules. However, both cell types also respond to lipopolysaccharide endotoxin (LPS) via their TLR4 receptors; in both cases this leads to secretion of IL-10. The LSEC also secrete TFG-β1 and express PD-L1, while Kupffer cells secrete PGE2. Thus, the CD4+ T cell is subject to a barrage of conflicting signals.
      The liver contains multiple populations of DC, including classical myeloid DC (mDC) and plasmacytoid DC (pDC), as well as other populations that are more elusive (Fig. 3). In the mouse, mDC are defined as CD11b+, CD11c+, and lacking in both CD8-α and B220. Compared to cells of similar phenotype isolated from the spleen, mouse liver mDC are less potent stimulators of T cell activation, possibly because of partial tolerance induced by constitutive exposure to endotoxin [
      • De Creus A.
      • Abe M.
      • Lau A.H.
      • Hackstein H.
      • Raimondi G.
      • Thomson A.W.
      Low TLR4 expression by liver dendritic cells correlates with reduced capacity to activate allogeneic T cells in response to endotoxin.
      ]. In human liver, a similar population of mDC expresses CD11b, CD11c, and blood DC antigen-1 (BDCA-1) and exhibits similar functions. Direct comparison of human liver-derived mDC with skin-derived mDC revealed that the liver cells secreted more IL-10; in addition, this promoted less proliferation, but more IL-10 secretion by T cells with which they interacted [
      • Goddard S.
      • Youster J.
      • Morgan E.
      • Adams D.H.
      Interleukin-10 secretion differentiates dendritic cells from human liver and skin.
      ]. This supports the model that human’s hepatic mDC predispose T cells towards tolerance. Such a bias may be imposed locally, since liver stromal cells were able to bias the differentiation of hematopoietic progenitor cells towards DC with a regulatory function, based in part on the stromal cells’ secretion of M-CSF [
      • Xia S.
      • Guo Z.
      • Xu X.
      • Yi H.
      • Wang Q.
      • Cao X.
      Hepatic microenvironment programs hematopoietic progenitor differentiation into regulatory dendritic cells, maintaining liver tolerance.
      ].
      Figure thumbnail gr3
      Fig. 3Complex subsets of murine liver DC. The figure shows that liver resident myeloid DC (mDC) are relatively weak APC, as are liver resident plasmacytoid DC (pDC). However, liver pDC are immature cells, which can be induced to differentiate into more powerful APC by TLR signals. Lymphoid-related (CD8α+) DC are rare cells with strong APC effects. The population termed NKDC is capable of killing tumor cells and can present their antigens to T cells. Hepatic pDC secrete IFN-α, while liver NKDC secrete IFN-γ.
      Myeloid DC are involved in trafficking through the liver, and one study shows that they migrate from the hepatic parenchyma to the portal tracts, which also contain T cells and are therefore a potential site of T cell priming [
      • Kudo S.
      • Matsuno K.
      • Ezaki T.
      • Ogawa M.
      A novel migration pathway for rat dendritic cells from the blood: hepatic sinusoids-lymph translocation.
      ]. Such an interaction could result in the formation of portal lymphoid aggregates, as occurs for example in Propionibacterium acnes-induced liver inflammation [
      • Yoneyama H.
      • Matsuno K.
      • Zhang Y.
      • Murai M.
      • Itakura M.
      • Ishikawa S.
      • et al.
      Regulation by chemokines of circulating dendritic cell precursors, and the formation of portal tract-associated lymphoid tissue, in a granulomatous liver disease.
      ], and could also occur in viral hepatitis. One issue for future investigation is the mechanism through which mDCs and the immune responses they initiate may be compartmentalized either to the parenchyma or to the portal tracts.
      Murine liver pDC are B220+ and express lower levels of CD11c than mDC. As with pDC from other sources, they secrete type 1 IFN but directly-isolated cells are not highly effective APC in T cell activation [
      • Tokita D.
      • Sumpter T.L.
      • Raimondi G.
      • Zahorchak A.F.
      • Wang Z.
      • Nakao A.
      • et al.
      Poor allostimulatory function of liver plasmacytoid DC is associated with pro-apoptotic activity, dependent on regulatory T cells.
      ]. However, both growth factors and TLR ligation can cause these cells to mature into effective APC that can stimulate T cells [
      • Kingham T.P.
      • Chaudhry U.I.
      • Plitas G.
      • Katz S.C.
      • Raab J.
      • DeMatteo R.P.
      Murine liver plasmacytoid dendritic cells become potent immunostimulatory cells after Flt-3 ligand expansion.
      ]. Therefore, their weak APC action ex vivo may reflect their differentiation state, rather than an intrinsic property of this subset. In human liver, pDC lack CD11c but express the marker BDCA-2. These cells are relatively abundant in liver, compared to other tissues [
      • Pillarisetty V.G.
      • Shah A.B.
      • Miller G.
      • Bleier J.I.
      • DeMatteo R.P.
      Liver dendritic cells are less immunogenic than spleen dendritic cells because of differences in subtype composition.
      ], and in vitro they activate regulatory T cells [
      • Moseman E.A.
      • Liang X.
      • Dawson A.J.
      • Panoskaltsis-Mortari A.
      • Krieg A.M.
      • Liu Y.J.
      • et al.
      Human plasmacytoid dendritic cells activated by CpG oligodeoxynucleotides induce the generation of CD4+CD25+ regulatory T cells.
      ], and may thus contribute to organ-specific immune tolerance.
      In the mouse, liver DC also contain a population that expresses the classic DC marker CD11c, along with CD8-α [
      • Pillarisetty V.G.
      • Shah A.B.
      • Miller G.
      • Bleier J.I.
      • DeMatteo R.P.
      Liver dendritic cells are less immunogenic than spleen dendritic cells because of differences in subtype composition.
      ]. These cells have been termed “lymphoid-related DC”; unlike pDC, these cells were strong simulators of T cell proliferation [
      • O’Connell P.J.
      • Morelli A.E.
      • Logar A.J.
      • Thomson A.W.
      Phenotypic and functional characterization of mouse hepatic CD8 alpha+ lymphoid-related dendritic cells.
      ]. However, no such cells have been identified in other mammalian species, raising questions about their significance in human disease. Still more enigmatic is a subset of mouse liver cells termed “NK-DC”, based on their expression of both NK cell markers (NK-1.1) and DC markers (CD11c). These cells are cytotoxic, but also act as APC in vitro [
      • Pillarisetty V.G.
      • Katz S.C.
      • Bleier J.I.
      • Shah A.B.
      • Dematteo R.P.
      Natural killer dendritic cells have both antigen presenting and lytic function and in response to CpG produce IFN-γ via autocrine IL-12.
      ]. Currently it is unclear whether the strongest affinity of these cells is with NK cells, or with other groups of DC. However, there is precedent for the close linkage of T lymphocyte, NK and DC maturation pathways in the thymic mDC, since all these arise from the earliest intrathymic progenitor cells [
      • Ardavin C.
      • Wu L.
      • Li C.L.
      • Shortman K.
      Thymic dendritic cells and T cells develop simultaneously in the thymus from a common precursor population.
      ,
      • Shen H.Q.
      • Lu M.
      • Ikawa T.
      • Masuda K.
      • Ohmura K.
      • Minato N.
      • et al.
      T/NK bipotent progenitors in the thymus retain the potential to generate dendritic cells.
      ]. Therefore, we could view these cells as NK cells that have acquired some DC-like properties, or as DC that retain some NK cell attributes from their precursors, or as aberrant cells that failed to commit to one or other of the lineages. Like the CD8α+ DC, these cells do not correspond to any subset in other mammals, so their relevance to human disease is unclear.

      Liver sinusoidal endothelial cells as APC

      A strong case has been made that LSEC are important APC that induce T cell tolerance, based on the isolation of these cells by centrifugal elutriation and evaluation of their purity based on their uptake of acetylated low-density lipoproteins (LDL). Thus, these cells isolated from primed mice will present ovalbumin given systemically, and cause CD8+ T cell tolerance [
      • Limmer A.
      • Ohl J.
      • Kurts C.
      • Ljunggren H.G.
      • Reiss Y.
      • Groettrup M.
      • et al.
      Efficient presentation of exogenous antigen by liver endothelial cells to CD8+ T cells results in antigen-specific T-cell tolerance.
      ]. Similarly, ovalbumin given orally induces T cell tolerance, which was transferable using isolated LSEC from the antigen-fed animals [
      • Limmer A.
      • Ohl J.
      • Wingender G.
      • Berg M.
      • Jungerkes F.
      • Schumak B.
      • et al.
      Cross-presentation of oral antigens by liver sinusoidal endothelial cells leads to CD8 T cell tolerance.
      ]. In such experiments, apoptotic tumor cells will also serve as an antigen donor for the induction of CD8+ T cell tolerance [
      • Berg M.
      • Wingender G.
      • Djandji D.
      • Hegenbarth S.
      • Momburg F.
      • Hammerling G.
      • et al.
      Cross-presentation of antigens from apoptotic tumor cells by liver sinusoidal endothelial cells leads to tumor-specific CD8+ T cell tolerance.
      ]. In an allostimulation model in vitro, cultured liver cells were APC poor, for both CD4+ and CD8+ T cells, but the depletion of LSEC from the stimulator cells revealed APC activity in other (albeit undefined) liver cells [
      • Onoe T.
      • Ohdan H.
      • Tokita D.
      • Shishida M.
      • Tanaka Y.
      • Hara H.
      • et al.
      Liver sinusoidal endothelial cells tolerize T cells across MHC barriers in mice.
      ].
      Several explanations have been advanced for the tolerance-inducing properties of LSEC. Two immunosuppressive products of Kupffer cells, PGE2 and IL-10, both down-regulate the APC function of LSEC. IL-10, in particular, down-regulated the expression of MHC class II, CD80 and CD86, compromising the antigen-specific and co-stimulatory signals [
      • Knolle P.A.
      • Uhrig A.
      • Hegenbarth S.
      • Loser E.
      • Schmitt E.
      • Gerken G.
      • et al.
      IL-10 down-regulates T cell activation by antigen-presenting liver sinusoidal endothelial cells through decreased antigen uptake via the mannose receptor and lowered surface expression of accessory molecules.
      ]. During interaction with antigen-specific T cells, LSEC up-regulated B7-H1 (PD-L1), which interacts with the PD-1 receptor on activated T cells, and leads to T cell inactivation [
      • Diehl L.
      • Schurich A.
      • Grochtmann R.
      • Hegenbarth S.
      • Chen L.
      • Knolle P.A.
      Tolerogenic maturation of liver sinusoidal endothelial cells promotes B7-homolog 1-dependent CD8+ T cell tolerance.
      ]. Recently, LSEC were shown to have an immunoregulatory effect mediated via cross-talk, due to a contact-dependent action on DC involving the loss of IL-12, CD80, and CD86 [
      • Schildberg F.A.
      • Hegenbarth S.I.
      • Schumak B.
      • Scholz K.
      • Limmer A.
      • Knolle P.A.
      Liver sinusoidal endothelial cells veto CD8 T cell activation by antigen-presenting dendritic cells.
      ]. A consistent model emerges in which LSEC scavenge circulating proteins from the circulation, present them with a battery of immunosuppressive signals, and thereby help to maintain self-tolerance (Fig. 2). In support of this concept, antigen presented by non-hematopoietic cells that expressed Kb under the control of the Tie-2 promoter, putative LSECs, were able to cause rapid localization of resting CD8+ T cells to the liver [
      • von Oppen N.
      • Schurich A.
      • Hegenbarth S.
      • Stabenow D.
      • Tolba R.
      • Weiskirchen R.
      • et al.
      Systemic antigen cross-presented by liver sinusoidal endothelial cells induces liver-specific CD8 T-cell retention and tolerization.
      ]. T cells primed on LSEC did not express the classic CD25+, FoxP3-high regulatory T cell phenotype, but were nevertheless able to suppress liver injury in experimental hepatitis in mice [
      • Kruse N.
      • Neumann K.
      • Schrage A.
      • Derkow K.
      • Schott E.
      • Erben U.
      • et al.
      Priming of CD4+ T cells by liver sinusoidal endothelial cells induces CD25low forkhead box protein 3-regulatory T cells suppressing autoimmune hepatitis.
      ]. The model of LSEC-based tolerance can easily be extended to serve the need to maintain immunological silence to harmless antigenic material in the diet.
      The claim that LSEC express MHC antigens and co-stimulatory molecules was challenged in a study that isolated these cells using density gradient centrifugation, followed by immunomagnetic beads to deplete essentially all CD45+ cells. The cell population isolated was evidently endothelial, since it expressed CD31, von Willebrand factor and Fcγ receptors, but not CD11c. These cells did not, however, express MHC class II or CD86, and did not appear to engage CD4+ or CD8+ T cells, based on measurements of proliferation [
      • Katz S.C.
      • Pillarisetty V.G.
      • Bleier J.I.
      • Shah A.B.
      • DeMatteo R.P.
      Liver sinusoidal endothelial cells are insufficient to activate T cells.
      ]. This study further showed that the uptake of acetylated LDL is not a unique feature of LSEC, but was also shared by DC, although the LSEC were faster at accumulating fluorescent acetylated LDL in vitro. So, are these cells really so different from the LSEC that were capable of cross-presentation of ovalbumin? Both cell populations were CD11c negative [
      • Katz S.C.
      • Pillarisetty V.G.
      • Bleier J.I.
      • Shah A.B.
      • DeMatteo R.P.
      Liver sinusoidal endothelial cells are insufficient to activate T cells.
      ,
      • Limmer A.
      • Ohl J.
      • Wingender G.
      • Berg M.
      • Jungerkes F.
      • Schumak B.
      • et al.
      Cross-presentation of oral antigens by liver sinusoidal endothelial cells leads to CD8 T cell tolerance.
      ] and both ultimately promoted an ineffective immune response, although in the case of the cells isolated using magnetic beads, there was no initial T cell proliferation. One key difference between the two kinds of cell isolates is the expression of CD86; LSEC isolated by magnetic bead depletion were CD86 negative, while LSEC isolated by elutriation were clearly CD86 positive, comparable to Kupffer cells [
      • Lohse A.W.
      • Knolle P.A.
      • Bilo K.
      • Uhrig A.
      • Waldmann C.
      • Ibe M.
      • et al.
      Antigen-presenting function and B7 expression of murine sinusoidal endothelial cells and Kupffer cells.
      ]. If the cells were in fact consonant populations, the magnetic bead-isolated LSEC might correspond most closely to LSEC that had been exposed to IL-10 during isolation; as already noted, such an exposure causes them to down-regulate both MHC class II and co-stimulatory molecules [
      • Knolle P.A.
      • Uhrig A.
      • Hegenbarth S.
      • Loser E.
      • Schmitt E.
      • Gerken G.
      • et al.
      IL-10 down-regulates T cell activation by antigen-presenting liver sinusoidal endothelial cells through decreased antigen uptake via the mannose receptor and lowered surface expression of accessory molecules.
      ]. In the present state of knowledge, it seems more likely that the isolation using magnetic beads on a column could have activated IL-10 secretion from Kupffer cells and modified the phenotype and function of the LSEC, rather than the extensive literature on LSEC as APC is entirely due to DC contamination.
      If we accept that LSEC are authentic APC, these studies raise the issue of their uniqueness among endothelia. There is, in fact, considerable evidence both for and against antigen presentation by other vascular endothelial cells. For example, highly-purified human umbilical vein endothelial cells (HUVEC), containing fewer than 0.01% leukocytes, were competent to cause primary activation in purified primary CD4+ and CD8+ T cells [
      • Page C.S.
      • Holloway N.
      • Smith H.
      • Yacoub M.
      • Rose M.L.
      Alloproliferative responses of purified CD4+ and CD8+ T cells to endothelial cells in the absence of contaminating accessory cells.
      ]. This study is noteworthy for the care with which cell purity was assessed; in particular, the T cells were unresponsive to anti-CD3 and PHA, indicating very effective depletion of contaminating APC. However, the finding is controversial, since a subsequent study found that HUVEC do not express CD80 and even under the influence of IFN-γ were able to induce only tolerance in primary CD4+ T cells [
      • Marelli-Berg F.M.
      • Hargreaves R.E.
      • Carmichael P.
      • Dorling A.
      • Lombardi G.
      • Lechler R.I.
      Major histocompatibility complex class II-expressing endothelial cells induce allospecific nonresponsiveness in naive T cells.
      ]. The reason for these apparently contradictory results is unclear. An analysis of the significance of MHC class II expression on vascular endothelium versus bone marrow-derived APC found that the presence or absence of MHC class II on the endothelium was not a major influence on the speed of cardiac allograft rejection by CD4+ T cells [
      • Kreisel D.
      • Krasinskas A.M.
      • Krupnick A.S.
      • Gelman A.E.
      • Balsara K.R.
      • Popma S.H.
      • et al.
      Vascular endothelium does not activate CD4+ direct allorecognition in graft rejection.
      ]. Several transgenic studies have used the endothelium-specific receptor tyrosine kinase type-2 (Tie-2) promoter to drive antigen expression in vascular endothelium. One study expressed β-galactosidase as antigen and led to an antibody response, but it was possible neither to raise an endogenous T cell response nor to induce vascular inflammation, even after priming with a recombinant vaccinia vector [
      • Rothermel A.L.
      • Wang Y.
      • Schechner J.
      • Mook-Kanamori B.
      • Aird W.C.
      • Pober J.S.
      • et al.
      Endothelial cells present antigens in vivo.
      ]. Similarly, Tie-2-β-galactosidase transgenic mice were infused with antigen-specific TCR transgenic CD8+ T cells, but these were neither activated nor tolerated [
      • Bolinger B.
      • Krebs P.
      • Tian Y.
      • Engeler D.
      • Scandella E.
      • Miller S.
      • et al.
      Immunologic ignorance of vascular endothelial cells expressing minor histocompatibility antigen.
      ]. The latter study is noteworthy because the livers of Tie-2-β-galactosidase transgenic mice were transplanted into naïve recipients; in such transplant experiments, the antigen should have been expressed primarily on hepatic vasculature and on LSEC. However, there was no interaction with CD8+ T cells, in direct contradiction to the expectations derived from studies with ex vivo isolated LSEC.
      These data could be reconciled in several ways. First, the LSEC could express APC activity ex vivo, but not in vivo. However, this would mean that the capacity of LSEC to cross-present oral and circulating antigens in a way that induces T cell tolerance does not explain either oral or IV induction of systemic tolerance, hardly a satisfying outcome! Secondly, it could mean that the in vitro activities of LSEC cultures are in fact due to low-level contamination by bone marrow-derived APC, such as Kupffer cells or DC. It can always be argued that any primary cell preparation is “not pure enough”; however, it does not make sense to write off the APC function of LSEC in view of the expression of MHC class I, MHC class II, and co-stimulatory molecules by these cells, if carefully isolated [
      • Knolle P.A.
      • Gerken G.
      Local control of the immune response in the liver.
      ,
      • Lalor P.F.
      • Lai W.K.
      • Curbishley S.M.
      • Shetty S.
      • Adams D.H.
      Human hepatic sinusoidal endothelial cells can be distinguished by expression of phenotypic markers related to their specialised functions in vivo.
      ]. Finally, many of the apparent discrepancies could be resolved if LSEC present exogenous antigens, but not their own intrinsic antigens, due to unusual features of their MHC class I and class II processing pathways. There is evidence that LSEC can take up antigen using many different cell surface receptors [
      • Elvevold K.H.
      • Nedredal G.I.
      • Revhaug A.
      • Smedsrod B.
      Scavenger properties of cultivated pig liver endothelial cells.
      ]. One very interesting feature of these cells is the disparity in the efficiency at antigen uptake, versus cross-presentation of a soluble antigen. A direct comparison with DCs suggested that the LSECs were much more efficient at uptake, but equivalent in terms of cross-presentation, suggesting differences in the cell biology of antigen handling in LSECs [
      • Schurich A.
      • Bottcher J.P.
      • Burgdorf S.
      • Penzler P.
      • Hegenbarth S.
      • Kern M.
      • et al.
      Distinct kinetics and dynamics of cross-presentation in liver sinusoidal endothelial cells compared to dendritic cells.
      ]. This observation argues strongly against the idea that the potent APC functions of LSECs can be explained by low-level DC contamination.

      Hepatic stellate cells as APC

      The sinusoids of the liver are surrounded by the Space of Disse, in which resides a distinctive population of vascular pericytes termed hepatic stellate cells, or Ito cells. These cells are understood to regulate blood flow through the sinusoids and have a key role in liver fibrosis, since they undergo trans-differentiation into myofibroblasts in response to diverse inflammatory and toxic insults, and in this role they secrete inhibitors of tissue matrix metalloproteinases, deposit collagen, and create dense fibrous tissue [
      • Friedman S.L.
      Hepatic fibrosis – overview.
      ]. The stellate cells recently surfaced as potential APC. Since the studies supporting the new function are few, it is challenging to synthesize the evidence for APC function in hepatic stellate cells.
      In a mouse model of liver fibrogenesis induced by injection of Carbon tetrachloride, immunohistochemistry revealed the co-location of lymphocytes and stellate cells, and the isolated stellate cells expressed both MHC class II and CD11c, consistent with an APC function [
      • Muhanna N.
      • Horani A.
      • Doron S.
      • Safadi R.
      Lymphocyte-hepatic stellate cell proximity suggests a direct interaction.
      ]. However, this approximation is also consistent with the concept that stellate cells are important in secreting chemokines that can promote lymphocyte localization, including CXCL9 and CXCL10 [
      • Holt A.P.
      • Haughton E.L.
      • Lalor P.F.
      • Filer A.
      • Buckley C.D.
      • Adams D.H.
      Liver myofibroblasts regulate infiltration and positioning of lymphocytes in human liver.
      ].
      The key paper supporting a role for stellate cells as APC is based on cells isolated from the mouse liver by collagenase-pronase digestion, followed by density gradient centrifugation. These cells were placed in culture and used in experiments to test APC function. The cells expressed CD1d, a low level of MHC class II, and half of them expressed a low level of CD11c; the flow cytometric profile in Fig. 1 of this paper reveals also a trace of CD11c-high cells [
      • Winau F.
      • Hegasy G.
      • Weiskirchen R.
      • Weber S.
      • Cassan C.
      • Sieling P.A.
      • et al.
      Ito cells are liver-resident antigen-presenting cells for activating T cell responses.
      ]. The cells were clearly shown to activate NK-T cells in the presence of α-galactosyl ceramide and to activate T cells also. The key issue here is the potential co-purification of trace numbers of CD11c-high DCs along with the stellate cells. Photomicrographs showed mainly fibroblastic cells, and individual cells stained for the presence of glial fibrillary acidic protein, consistent with stellate cells. However, while the staining for low levels of CD11c on many of the cells is consistent with other reports, the small minority of CD11c-high cells raises the possibility that co-purified DC might contribute to the APC effect. To be fair, this caveat could apply to many other studies based on liver APC purification, including our own [
      • Wuensch S.A.
      • Pierce R.H.
      • Crispe I.N.
      Local intrahepatic CD8+ T cell activation by a non-self-antigen results in full functional differentiation.
      ].
      Figure thumbnail fx2

      Cholangiocytes as APC?

      The epithelial cells lining the bile ducts, cholangiocytes, express many molecules often linked to APC function, and these cells are a focus of inflammation in biliary cirrhosis and sclerosing cholangitis, raising the possibility that they might initiate an immune response. Human cholangiocytes express a variety of TLRs, and at least TLR-2 and TLR-4 are functional, signaling via MyD88 and causing NF-κB activation [
      • Chen X.M.
      • O’Hara S.P.
      • Nelson J.B.
      • Splinter P.L.
      • Small A.J.
      • Tietz P.S.
      • et al.
      Multiple TLRs are expressed in human cholangiocytes and mediate host epithelial defense responses to Cryptosporidium parvum via activation of NF-κB.
      ]. Cholangiocytes can secrete chemokines such as CXCL16 that promote adhesion of T cells [
      • Heydtmann M.
      • Lalor P.F.
      • Eksteen J.A.
      • Hubscher S.G.
      • Briskin M.
      • Adams D.H.
      CXC chemokine ligand 16 promotes integrin-mediated adhesion of liver-infiltrating lymphocytes to cholangiocytes and hepatocytes within the inflamed human liver.
      ]. Freshly isolated murine cholangiocytes expressed MHC class I, class II, and CD40, and after a period of culture also expressed CD80 and CD86. However, this same study did not find APC activity in the cholangiocytes [
      • Barnes B.H.
      • Tucker R.M.
      • Wehrmann F.
      • Mack D.G.
      • Ueno Y.
      • Mack C.L.
      Cholangiocytes as immune modulators in rotavirus-induced murine biliary atresia.
      ]. This may be linked to the fact that T cells express CD154, while ligation of its counter-receptor CD40 on cholangiocytes results in their apoptosis [
      • Alabraba E.B.
      • Lai V.
      • Boon L.
      • Wigmore S.J.
      • Adams D.H.
      • Afford S.C.
      Coculture of human liver macrophages and cholangiocytes leads to CD40-dependent apoptosis and cytokine secretion.
      ]. In the current consensus, it seems likely that cholangiocytes can be the targets of immune responses, but are unique among major liver cell populations in that no evidence supports the possibility that they activate naïve T cells.

      Hepatocytes as APC

      The doctrine that antigen presentation is the concern of a specialized subset of cells has given rise to the core idea that whereas T cells respond to antigen on such specialized APC by full activation, clonal expansion and differentiation, the outcome of antigen encounter of most tissue cells is either the lack of a response or the induction of tolerance. The mainstream doctrine requires some qualification with respect to MHC class I-restricted cytotoxic T cells; these cells must deliver their effector functions by recognizing antigenic tissue cells. From this perspective, we can approach the issue of how hepatocytes act as APC, and what are the outcomes for T cell function.
      Several studies document the effects of isolated, cultured hepatocytes as APC. In a fully allogeneic liver transplant model, the early apoptosis of graft-infiltrating T cells was investigated by exposing isolated, allogeneic hepatocytes to T cells; the outcome was T cell activation, followed by apoptosis [
      • Qian S.
      • Wang Z.
      • Lee Y.
      • Chiang Y.
      • Bonham C.
      • Fung J.
      • et al.
      Hepatocyte-induced apoptosis of activated T cells, a mechanism of liver transplant tolerance, is related to the expression of ICAM-1 and hepatic lectin.
      ]. Similarly, the abortive activation of TCR transgenic CD8+ T cells in mice expressing the Kb alloantigen in hepatocytes led to in vitro experiments, and again the outcome was T cell activation, followed by apoptosis [
      • Bertolino P.
      • Trescol-Biemont M.C.
      • Rabourdin-Combe C.
      Hepatocytes induce functional activation of naive CD8+ T lymphocytes but fail to promote survival.
      ]. The first feature of interest in these latter experiments is that the naïve, as well as primed, T cells were activated. How can we explain this? In the case of the fully allogenic experiments, it could be hypothesized that the alloreactive T cells were in fact previously-activated T cells that were cross-reactive on alloantigens. In the case of the experiments using the Kb transgenic hepatocytes, it could be argued that the responding T cells were in fact expressing dual T cell receptors, due to a lack of allelic exclusion of the TCR α chain [
      • Heath W.R.
      • Miller J.F.
      Expression of two alpha chains on the surface of T cells in T cell receptor transgenic mice.
      ]. Both of these interpretations essentially deny that the hepatocytes could activate naive CD8+ T cells. The alternative interpretation, taken by the authors, is that such activation is possible, but that the activation leads to early apoptosis, resulting in functional tolerance.
      As with the experiments discussed above in relation to Ito cell APC function, the key question here is: what was in the culture? Hepatocytes are isolated by a standard protocol involving perfusion of the liver with collagenase, followed by the making of a single-cell suspension which is then plated, typically in a specialized medium that favors hepatocyte growth, rather than the media used for lymphocyte culture. This cell isolation protocol was developed to generate cultures of hepatocytes for metabolic studies, and the contribution of LSEC, Ito cells, Kupffer cells, and possibly other leukocytes is difficult to assess. Certainly, these cell isolates present the microscopic appearance of a monoculture of large cells. However, it is customary neither to validate the hepatocyte culture by testing the cultures for the uptake of acetylated LDL, to identify contaminating LSEC, nor to stain for CD45, to detect leukocytes, nor to perform quantitative qRT-PCR to estimate the expression of genes characteristic of other cell types, such as F4/80 in Kupffer cells and α-smooth muscle actin in activated Ito cells. Therefore, the possibility remains that, in a culture of hepatocytes, the active APC were one of the other cell types we have already discussed.
      Experiments in vivo have evaluated the APC functions of hepatocytes, using alloantigens driven by hepatocyte-specific promoters such as albumin and metallothionine. These experiments support the broad consensus that hepatocytes cause primary T cell activation and that this activation is abortive and leads to premature T cell apoptosis [
      • Arnold B.
      Parenchymal cells in immune and tolerance induction.
      ,
      • Bertolino P.
      • McCaughan G.W.
      • Bowen D.G.
      Role of primary intrahepatic T-cell activation in the ‘liver tolerance effect’.
      ,
      • Schonrich G.
      • Momburg F.
      • Malissen M.
      • Schmitt-Verhulst A.M.
      • Malissen B.
      • Hammerling G.J.
      • et al.
      Distinct mechanisms of extrathymic T cell tolerance due to differential expression of self antigen.
      ]. However, such transgenic experiments are complicated by the possibility that the transgenic antigen could be expressed in the thymic medulla [
      • Smith K.M.
      • Olson D.C.
      • Hirose R.
      • Hanahan D.
      Pancreatic gene expression in rare cells of thymic medulla: evidence for functional contribution to T cell tolerance.
      ], potentially promoting the differentiation of regulatory T cells [
      • Jordan M.S.
      • Boesteanu A.
      • Reed A.J.
      • Petrone A.L.
      • Holenbeck A.E.
      • Lerman M.A.
      • et al.
      Thymic selection of CD4+CD25+ regulatory T cells induced by an agonist self-peptide.
      ]. Taking an alternative approach, investigators used an AAV vector to specifically deliver antigen to hepatocytes; this resulted in T cell activation, leading to effector function [
      • Wuensch S.A.
      • Pierce R.H.
      • Crispe I.N.
      Local intrahepatic CD8+ T cell activation by a non-self-antigen results in full functional differentiation.
      ]. However, this experiment used a very high number of antigen-specific CD8+ T cells, which may have been substituting for missing CD4+ T cell help [
      • Wuensch S.A.
      • Spahn J.
      • Crispe I.N.
      Direct, help-independent priming of CD8+ T cells by adeno-associated virus-transduced hepatocytes.
      ]. All of these experiments, taken together, fit with the idea that hepatocytes are effective primary APC for CD8+ T cells, but that the outcome of such T cell activation occurs in a context where CD4+ T cell help may be limiting, and regulatory T cells may be present; both of these factors favor an abortive immune response.

      Conclusions

      In the preceding discussion, I have argued that, despite some inconsistencies in the literature, immune responses in the liver follow distinctive rules, and this may be explained in part by features of the liver’s APC. It is not possible to say which cell type is the most important liver APC, but many cell populations can make a strong claim. A critical analysis of the current literature broadly supports the following tenets:
      • 1.
        The liver’s DCs are distinctive and create a bias towards tolerance both because of their intrinsic properties and because of a different representation of DC subsets.
      • 2.
        Kupffer cells act as APC but favor immunosuppression, partly due to secretion of IL-10 and partly to secretion of PGE2.
      • 3.
        Liver endothelial cells capture and present protein antigens, but the outcome is often immunosuppression due to their secretion of IL-10, TGF-β1, and their expression of PD-L1. Strangely, evidence suggests that they do not present their own cell-intrinsic antigens.
      • 4.
        Hepatic stellate cells are a potent source of chemokines, and activate CD1d-reactive T cells, but whether they promote immunity or immune tolerance in CD4+ and CD8+ T cells still remains to be clarified.
      • 5.
        There is no clear evidence that cholangiocytes are antigen-presenting cells.
      • 6.
        Hepatocytes can cause primary CD8+ T cell activation, which may result in effector cells but does not result in a sustained response.
      This summary of the state-of-the-art provides many explanations for the phenomenon of liver transplantation tolerance, and for the failure of immunity to clear some liver pathogens. As a real-world problem, liver allograft tolerance is of limited significance, because of the effectiveness of immunosuppressive drugs. The issue now is to determine which of these mechanisms actually contribute to defective anti-pathogen immunity and which are innocent bystanders.

      Financial support

      The development of these ideas over the years has been supported principally by the NIAID and the NIDDK (Grants AI063353 , AI064463 , AI072049 , and DK075274 ).

      Conflict of interest

      The authors who have taken part in this study declare that they do not have anything to disclose regarding conflict of interest with respect to this manuscript.

      Acknowledgements

      I want to thank Drs. Lin Huang, Wajahat Mehal, Katia Klugewitz, Ingo Klein, Sherry Wuensch, Percy Knolle, Patrick Bertolino, Angus Thomson, David Adams, Cliona O’Farrelly, Mark Orloff, and Robert Pierce for many invaluable discussions on liver antigen presentation.

      References

        • Alabraba E.B.
        • Lai V.
        • Boon L.
        • Wigmore S.J.
        • Adams D.H.
        • Afford S.C.
        Coculture of human liver macrophages and cholangiocytes leads to CD40-dependent apoptosis and cytokine secretion.
        Hepatology. 2008; 47: 552-562
        • Ardavin C.
        • Wu L.
        • Li C.L.
        • Shortman K.
        Thymic dendritic cells and T cells develop simultaneously in the thymus from a common precursor population.
        Nature. 1993; 362: 761-763
        • Arnold B.
        Parenchymal cells in immune and tolerance induction.
        Immunol Lett. 2003; 89: 225-228
        • Baer K.
        • Roosevelt M.
        • Clarkson Jr., A.B.
        • van Rooijen N.
        • Schnieder T.
        • Frevert U.
        Kupffer cells are obligatory for Plasmodium yoelii sporozoite infection of the liver.
        Cell Microbiol. 2007; 9: 397-412
        • Barnes B.H.
        • Tucker R.M.
        • Wehrmann F.
        • Mack D.G.
        • Ueno Y.
        • Mack C.L.
        Cholangiocytes as immune modulators in rotavirus-induced murine biliary atresia.
        Liver Int. 2009; 29: 1253-1261
        • Baron J.L.
        • Gardiner L.
        • Nishimura S.
        • Shinkai K.
        • Locksley R.
        • Ganem D.
        Activation of a nonclassical NKT cell subset in a transgenic mouse model of hepatitis B virus infection.
        Immunity. 2002; 16: 583-594
        • Berg M.
        • Wingender G.
        • Djandji D.
        • Hegenbarth S.
        • Momburg F.
        • Hammerling G.
        • et al.
        Cross-presentation of antigens from apoptotic tumor cells by liver sinusoidal endothelial cells leads to tumor-specific CD8+ T cell tolerance.
        Eur J Immunol. 2006; 36: 2960-2970
        • Bertolino P.
        • McCaughan G.W.
        • Bowen D.G.
        Role of primary intrahepatic T-cell activation in the ‘liver tolerance effect’.
        Immunol Cell Biol. 2002; 80: 84-92
        • Bertolino P.
        • Trescol-Biemont M.C.
        • Rabourdin-Combe C.
        Hepatocytes induce functional activation of naive CD8+ T lymphocytes but fail to promote survival.
        Eur J Immunol. 1998; 28: 221-236
        • Bolinger B.
        • Krebs P.
        • Tian Y.
        • Engeler D.
        • Scandella E.
        • Miller S.
        • et al.
        Immunologic ignorance of vascular endothelial cells expressing minor histocompatibility antigen.
        Blood. 2008; 111: 4588-4595
        • Bomble M.
        • Tacke F.
        • Rink L.
        • Kovalenko E.
        • Weiskirchen R.
        Analysis of antigen-presenting functionality of cultured rat hepatic stellate cells and transdifferentiated myofibroblasts.
        Biochem Biophys Res Commun. 2010; 396: 342-347
        • Callery M.P.
        • Mangino M.J.
        • Flye M.W.
        Arginine-specific suppression of mixed lymphocyte culture reactivity by Kupffer cells – a basis of portal venous tolerance.
        Transplantation. 1991; 51: 1076-1080
        • Calne R.Y.
        Immunological tolerance – the liver effect.
        Immunol Rev. 2000; 174: 280-282
        • Calne R.Y.
        • Sells R.A.
        • Pena J.R.
        • Davis D.R.
        • Millard P.R.
        • Herbertson B.M.
        • et al.
        Induction of immunological tolerance by porcine liver allografts.
        Nature. 1969; 223: 472-476
        • Calne R.Y.
        • White H.J.
        • Yoffa D.E.
        • Binns R.M.
        • Maginn R.R.
        • Herbertson R.M.
        • et al.
        Prolonged survival of liver transplants in the pig.
        Br Med J. 1967; 4: 645-648
        • Chen X.M.
        • O’Hara S.P.
        • Nelson J.B.
        • Splinter P.L.
        • Small A.J.
        • Tietz P.S.
        • et al.
        Multiple TLRs are expressed in human cholangiocytes and mediate host epithelial defense responses to Cryptosporidium parvum via activation of NF-κB.
        J Immunol. 2005; 175: 7447-7456
        • Crispe I.N.
        • Mehal W.Z.
        Strange brew: T cells in the liver.
        Immunol Today. 1996; 17: 522-525
        • De Creus A.
        • Abe M.
        • Lau A.H.
        • Hackstein H.
        • Raimondi G.
        • Thomson A.W.
        Low TLR4 expression by liver dendritic cells correlates with reduced capacity to activate allogeneic T cells in response to endotoxin.
        J Immunol. 2005; 174: 2037-2045
        • Diehl L.
        • Schurich A.
        • Grochtmann R.
        • Hegenbarth S.
        • Chen L.
        • Knolle P.A.
        Tolerogenic maturation of liver sinusoidal endothelial cells promotes B7-homolog 1-dependent CD8+ T cell tolerance.
        Hepatology. 2008; 47: 296-305
        • Eksteen B.
        • Mora J.R.
        • Haughton E.L.
        • Henderson N.C.
        • Lee-Turner L.
        • Villablanca E.J.
        • et al.
        Gut homing receptors on CD8 T cells are retinoic acid dependent and not maintained by liver dendritic or stellate cells.
        Gastroenterology. 2009; 137: 320-329
        • Elvevold K.H.
        • Nedredal G.I.
        • Revhaug A.
        • Smedsrod B.
        Scavenger properties of cultivated pig liver endothelial cells.
        Comp Hepatol. 2004; 3: 4
        • Freudenberg M.A.
        • Freudenberg N.
        • Galanos C.
        Time course of cellular distribution of endotoxin in liver, lungs and kidneys of rats.
        Br J Exp Pathol. 1982; 63: 56-65
        • Friedman S.L.
        Hepatic fibrosis – overview.
        Toxicology. 2008; 254: 120-129
        • Geissmann F.
        • Cameron T.O.
        • Sidobre S.
        • Manlongat N.
        • Kronenberg M.
        • Briskin M.J.
        • et al.
        Intravascular immune surveillance by CXCR6+ NKT cells patrolling liver sinusoids.
        PLoS Biol. 2005; 3: e113
        • Goddard S.
        • Youster J.
        • Morgan E.
        • Adams D.H.
        Interleukin-10 secretion differentiates dendritic cells from human liver and skin.
        Am J Pathol. 2004; 164: 511-519
        • Heath W.R.
        • Miller J.F.
        Expression of two alpha chains on the surface of T cells in T cell receptor transgenic mice.
        J Exp Med. 1993; 178: 1807-1811
        • Heydtmann M.
        • Lalor P.F.
        • Eksteen J.A.
        • Hubscher S.G.
        • Briskin M.
        • Adams D.H.
        CXC chemokine ligand 16 promotes integrin-mediated adhesion of liver-infiltrating lymphocytes to cholangiocytes and hepatocytes within the inflamed human liver.
        J Immunol. 2005; 174: 1055-1062
        • Holt A.P.
        • Haughton E.L.
        • Lalor P.F.
        • Filer A.
        • Buckley C.D.
        • Adams D.H.
        Liver myofibroblasts regulate infiltration and positioning of lymphocytes in human liver.
        Gastroenterology. 2009; 136: 705-714
        • Jiang G.
        • Yang H.R.
        • Wang L.
        • Wildey G.M.
        • Fung J.
        • Qian S.
        • et al.
        Hepatic stellate cells preferentially expand allogeneic CD4+ CD25+ FoxP3+ regulatory T cells in an IL-2-dependent manner.
        Transplantation. 2008; 86: 1492-1502
        • Jobe O.
        • Lumsden J.
        • Mueller A.K.
        • Williams J.
        • Silva-Rivera H.
        • Kappe S.H.
        • et al.
        Genetically attenuated Plasmodium berghei liver stages induce sterile protracted protection that is mediated by major histocompatibility complex Class I-dependent interferon-γ-producing CD8+ T cells.
        J Infect Dis. 2007; 196: 599-607
        • John B.
        • Crispe I.N.
        Passive and active mechanisms trap activated CD8+ T cells in the liver.
        J Immunol. 2004; 172: 5222-5229
        • Jordan M.S.
        • Boesteanu A.
        • Reed A.J.
        • Petrone A.L.
        • Holenbeck A.E.
        • Lerman M.A.
        • et al.
        Thymic selection of CD4+CD25+ regulatory T cells induced by an agonist self-peptide.
        Nat Immunol. 2001; 2: 301-306
        • Katz S.C.
        • Pillarisetty V.G.
        • Bleier J.I.
        • Shah A.B.
        • DeMatteo R.P.
        Liver sinusoidal endothelial cells are insufficient to activate T cells.
        J Immunol. 2004; 173: 230-235
        • Kennedy D.W.
        • Abkowitz J.L.
        Kinetics of central nervous system microglial and macrophage engraftment: analysis using a transgenic bone marrow transplantation model.
        Blood. 1997; 90: 986-993
        • Kingham T.P.
        • Chaudhry U.I.
        • Plitas G.
        • Katz S.C.
        • Raab J.
        • DeMatteo R.P.
        Murine liver plasmacytoid dendritic cells become potent immunostimulatory cells after Flt-3 ligand expansion.
        Hepatology. 2007; 45: 445-454
        • Klein I.
        • Cornejo J.C.
        • Polakos N.K.
        • John B.
        • Wuensch S.A.
        • Topham D.J.
        • et al.
        Kupffer cell heterogeneity: functional properties of bone marrow derived and sessile hepatic macrophages.
        Blood. 2007; 110: 4077-4085
        • Knolle P.
        • Schlaak J.
        • Uhrig A.
        • Kempf P.
        • Meyer zum Buschenfelde K.H.
        • Gerken G.
        Human Kupffer cells secrete IL-10 in response to lipopolysaccharide (LPS) challenge.
        J Hepatol. 1995; 22: 226-229
        • Knolle P.A.
        • Gerken G.
        Local control of the immune response in the liver.
        Immunol Rev. 2000; 174: 21-34
        • Knolle P.A.
        • Uhrig A.
        • Hegenbarth S.
        • Loser E.
        • Schmitt E.
        • Gerken G.
        • et al.
        IL-10 down-regulates T cell activation by antigen-presenting liver sinusoidal endothelial cells through decreased antigen uptake via the mannose receptor and lowered surface expression of accessory molecules.
        Clin Exp Immunol. 1998; 114: 427-433
        • Kreisel D.
        • Krasinskas A.M.
        • Krupnick A.S.
        • Gelman A.E.
        • Balsara K.R.
        • Popma S.H.
        • et al.
        Vascular endothelium does not activate CD4+ direct allorecognition in graft rejection.
        J Immunol. 2004; 173: 3027-3034
        • Kruse N.
        • Neumann K.
        • Schrage A.
        • Derkow K.
        • Schott E.
        • Erben U.
        • et al.
        Priming of CD4+ T cells by liver sinusoidal endothelial cells induces CD25low forkhead box protein 3-regulatory T cells suppressing autoimmune hepatitis.
        Hepatology. 2009; 50: 1904-1913
        • Kudo S.
        • Matsuno K.
        • Ezaki T.
        • Ogawa M.
        A novel migration pathway for rat dendritic cells from the blood: hepatic sinusoids-lymph translocation.
        J Exp Med. 1997; 185: 777-784
        • Lalor P.F.
        • Lai W.K.
        • Curbishley S.M.
        • Shetty S.
        • Adams D.H.
        Human hepatic sinusoidal endothelial cells can be distinguished by expression of phenotypic markers related to their specialised functions in vivo.
        World J Gastroenterol. 2006; 12: 5429-5439
        • Lang K.S.
        • Georgiev P.
        • Recher M.
        • Navarini A.A.
        • Bergthaler A.
        • Heikenwalder M.
        • et al.
        Immunoprivileged status of the liver is controlled by Toll-like receptor 3 signaling.
        J Clin Invest. 2006; 116: 2456-2463
        • Lee W.Y.
        • Moriarty T.J.
        • Wong C.H.
        • Zhou H.
        • Strieter R.M.
        • van Rooijen N.
        • et al.
        An intravascular immune response to Borrelia burgdorferi involves Kupffer cells and iNKT cells.
        Nat Immunol. 2010; 11: 295-302
        • Li W.
        • Kuhr C.S.
        • Zheng X.X.
        • Carper K.
        • Thomson A.W.
        • Reyes J.D.
        • et al.
        New insights into mechanisms of spontaneous liver transplant tolerance: the role of Foxp3-expressing CD25+CD4+ regulatory T cells.
        Am J Transplant. 2008; 8: 1639-1651
        • Limmer A.
        • Ohl J.
        • Kurts C.
        • Ljunggren H.G.
        • Reiss Y.
        • Groettrup M.
        • et al.
        Efficient presentation of exogenous antigen by liver endothelial cells to CD8+ T cells results in antigen-specific T-cell tolerance.
        Nat Med. 2000; 6: 1348-1354
        • Limmer A.
        • Ohl J.
        • Wingender G.
        • Berg M.
        • Jungerkes F.
        • Schumak B.
        • et al.
        Cross-presentation of oral antigens by liver sinusoidal endothelial cells leads to CD8 T cell tolerance.
        Eur J Immunol. 2005; 35: 2970-2981
        • Lohse A.W.
        • Knolle P.A.
        • Bilo K.
        • Uhrig A.
        • Waldmann C.
        • Ibe M.
        • et al.
        Antigen-presenting function and B7 expression of murine sinusoidal endothelial cells and Kupffer cells.
        Gastroenterology. 1996; 110: 1175-1181
        • Lumsden A.B.
        • Henderson J.M.
        • Kutner M.H.
        Endotoxin levels measured by a chromogenic assay in portal, hepatic and peripheral venous blood in patients with cirrhosis.
        Hepatology. 1988; 8: 232-236
        • Maemura K.
        • Zheng Q.
        • Wada T.
        • Ozaki M.
        • Takao S.
        • Aikou T.
        • et al.
        Reactive oxygen species are essential mediators in antigen presentation by Kupffer cells.
        Immunol Cell Biol. 2005; 83: 336-343
        • Marelli-Berg F.M.
        • Hargreaves R.E.
        • Carmichael P.
        • Dorling A.
        • Lombardi G.
        • Lechler R.I.
        Major histocompatibility complex class II-expressing endothelial cells induce allospecific nonresponsiveness in naive T cells.
        J Exp Med. 1996; 183: 1603-1612
        • Mazariegos G.V.
        • Zahorchak A.F.
        • Reyes J.
        • Chapman H.
        • Zeevi A.
        • Thomson A.W.
        Dendritic cell subset ratio in tolerant, weaning and non-tolerant liver recipients is not affected by extent of immunosuppression.
        Am J Transplant. 2005; 5: 314-322
        • McCaughan G.W.
        • Zekry A.
        Impact of immunosuppression on immunopathogenesis of liver damage in hepatitis C virus-infected recipients following liver transplantation.
        Liver Transpl. 2003; 9: S21-S27
        • Mehal W.Z.
        • Juedes A.E.
        • Crispe I.N.
        Selective retention of activated CD8+ T cells by the normal liver.
        J Immunol. 1999; 163: 3202-3210
        • Moseman E.A.
        • Liang X.
        • Dawson A.J.
        • Panoskaltsis-Mortari A.
        • Krieg A.M.
        • Liu Y.J.
        • et al.
        Human plasmacytoid dendritic cells activated by CpG oligodeoxynucleotides induce the generation of CD4+CD25+ regulatory T cells.
        J Immunol. 2004; 173: 4433-4442
        • Mueller A.K.
        • Labaied M.
        • Kappe S.H.
        • Matuschewski K.
        Genetically modified Plasmodium parasites as a protective experimental malaria vaccine.
        Nature. 2005; 433: 164-167
        • Muhanna N.
        • Horani A.
        • Doron S.
        • Safadi R.
        Lymphocyte-hepatic stellate cell proximity suggests a direct interaction.
        Clin Exp Immunol. 2007; 148: 338-347
        • Norris S.
        • Collins C.
        • Doherty D.G.
        • Smith F.
        • McEntee G.
        • Traynor O.
        • et al.
        Resident human hepatic lymphocytes are phenotypically different from circulating lymphocytes.
        J Hepatol. 1998; 28: 84-90
        • O’Connell P.J.
        • Morelli A.E.
        • Logar A.J.
        • Thomson A.W.
        Phenotypic and functional characterization of mouse hepatic CD8 alpha+ lymphoid-related dendritic cells.
        J Immunol. 2000; 165: 795-803
        • Onoe T.
        • Ohdan H.
        • Tokita D.
        • Shishida M.
        • Tanaka Y.
        • Hara H.
        • et al.
        Liver sinusoidal endothelial cells tolerize T cells across MHC barriers in mice.
        J Immunol. 2005; 175: 139-146
        • Page C.S.
        • Holloway N.
        • Smith H.
        • Yacoub M.
        • Rose M.L.
        Alloproliferative responses of purified CD4+ and CD8+ T cells to endothelial cells in the absence of contaminating accessory cells.
        Transplantation. 1994; 57: 1628-1637
        • Pillarisetty V.G.
        • Katz S.C.
        • Bleier J.I.
        • Shah A.B.
        • Dematteo R.P.
        Natural killer dendritic cells have both antigen presenting and lytic function and in response to CpG produce IFN-γ via autocrine IL-12.
        J Immunol. 2005; 174: 2612-2618
        • Pillarisetty V.G.
        • Shah A.B.
        • Miller G.
        • Bleier J.I.
        • DeMatteo R.P.
        Liver dendritic cells are less immunogenic than spleen dendritic cells because of differences in subtype composition.
        J Immunol. 2004; 172: 1009-1017
        • Pradel G.
        • Frevert U.
        Malaria sporozoites actively enter and pass through rat Kupffer cells prior to hepatocyte invasion.
        Hepatology. 2001; 33: 1154-1165
        • Pradel G.
        • Garapaty S.
        • Frevert U.
        Proteoglycans mediate malaria sporozoite targeting to the liver.
        Mol Microbiol. 2002; 45: 637-651
        • Qian S.
        • Demetris A.J.
        • Murase N.
        • Rao A.S.
        • Fung J.J.
        • Starzl T.E.
        Murine liver allograft transplantation: tolerance and donor cell chimerism.
        Hepatology. 1994; 19: 916-924
        • Qian S.
        • Wang Z.
        • Lee Y.
        • Chiang Y.
        • Bonham C.
        • Fung J.
        • et al.
        Hepatocyte-induced apoptosis of activated T cells, a mechanism of liver transplant tolerance, is related to the expression of ICAM-1 and hepatic lectin.
        Transplant Proc. 2001; 33: 226
        • Rothermel A.L.
        • Wang Y.
        • Schechner J.
        • Mook-Kanamori B.
        • Aird W.C.
        • Pober J.S.
        • et al.
        Endothelial cells present antigens in vivo.
        BMC Immunol. 2004; 5: 5
        • Sato K.
        • Yabuki K.
        • Haba T.
        • Maekawa T.
        Role of Kupffer cells in the induction of tolerance after liver transplantation.
        J Surg Res. 1996; 63: 433-438
        • Schildberg F.A.
        • Hegenbarth S.I.
        • Schumak B.
        • Scholz K.
        • Limmer A.
        • Knolle P.A.
        Liver sinusoidal endothelial cells veto CD8 T cell activation by antigen-presenting dendritic cells.
        Eur J Immunol. 2008; 38: 957-967
        • Schonrich G.
        • Momburg F.
        • Malissen M.
        • Schmitt-Verhulst A.M.
        • Malissen B.
        • Hammerling G.J.
        • et al.
        Distinct mechanisms of extrathymic T cell tolerance due to differential expression of self antigen.
        Int Immunol. 1992; 4: 581-590
        • Schurich A.
        • Bottcher J.P.
        • Burgdorf S.
        • Penzler P.
        • Hegenbarth S.
        • Kern M.
        • et al.
        Distinct kinetics and dynamics of cross-presentation in liver sinusoidal endothelial cells compared to dendritic cells.
        Hepatology. 2009; 50: 909-919
        • Shen H.Q.
        • Lu M.
        • Ikawa T.
        • Masuda K.
        • Ohmura K.
        • Minato N.
        • et al.
        T/NK bipotent progenitors in the thymus retain the potential to generate dendritic cells.
        J Immunol. 2003; 171: 3401-3406
        • Smith F.
        • Golden-Mason L.
        • Deignan T.
        • Norris S.
        • Nolan N.
        • Traynor O.
        • et al.
        Localization of T and B lymphocytes in histologically normal adult human donor liver.
        Hepatogastroenterology. 2003; 50: 1311-1315
        • Smith K.M.
        • Olson D.C.
        • Hirose R.
        • Hanahan D.
        Pancreatic gene expression in rare cells of thymic medulla: evidence for functional contribution to T cell tolerance.
        Int Immunol. 1997; 9: 1355-1365
        • Tokita D.
        • Sumpter T.L.
        • Raimondi G.
        • Zahorchak A.F.
        • Wang Z.
        • Nakao A.
        • et al.
        Poor allostimulatory function of liver plasmacytoid DC is associated with pro-apoptotic activity, dependent on regulatory T cells.
        J Hepatol. 2008; 49: 1008-1018
        • Tu Z.
        • Bozorgzadeh A.
        • Crispe I.N.
        • Orloff M.S.
        The activation state of human intrahepatic lymphocytes.
        Clin Exp Immunol. 2007; 149: 186-193
        • Tu Z.
        • Bozorgzadeh A.
        • Pierce R.H.
        • Kurtis J.
        • Crispe I.N.
        • Orloff M.S.
        TLR-dependent cross talk between human Kupffer cells and NK cells.
        J Exp Med. 2008; 205: 233-244
        • Velazquez P.
        • Cameron T.O.
        • Kinjo Y.
        • Nagarajan N.
        • Kronenberg M.
        • Dustin M.L.
        Cutting edge: activation by innate cytokines or microbial antigens can cause arrest of natural killer T cell patrolling of liver sinusoids.
        J Immunol. 2008; 180: 2024-2028
        • von Oppen N.
        • Schurich A.
        • Hegenbarth S.
        • Stabenow D.
        • Tolba R.
        • Weiskirchen R.
        • et al.
        Systemic antigen cross-presented by liver sinusoidal endothelial cells induces liver-specific CD8 T-cell retention and tolerization.
        Hepatology. 2009; 49: 1664-1672
        • Warren A.
        • Le Couteur D.G.
        • Fraser R.
        • Bowen D.G.
        • McCaughan G.W.
        • Bertolino P.
        T lymphocytes interact with hepatocytes through fenestrations in murine liver sinusoidal endothelial cells.
        Hepatology. 2006; 44: 1182-1190
        • Winau F.
        • Hegasy G.
        • Weiskirchen R.
        • Weber S.
        • Cassan C.
        • Sieling P.A.
        • et al.
        Ito cells are liver-resident antigen-presenting cells for activating T cell responses.
        Immunity. 2007; 26: 117-129
        • Wuensch S.A.
        • Pierce R.H.
        • Crispe I.N.
        Local intrahepatic CD8+ T cell activation by a non-self-antigen results in full functional differentiation.
        J Immunol. 2006; 177: 1689-1697
        • Wuensch S.A.
        • Spahn J.
        • Crispe I.N.
        Direct, help-independent priming of CD8+ T cells by adeno-associated virus-transduced hepatocytes.
        Hepatology. 2010; 52: 1068-1077
        • Xia S.
        • Guo Z.
        • Xu X.
        • Yi H.
        • Wang Q.
        • Cao X.
        Hepatic microenvironment programs hematopoietic progenitor differentiation into regulatory dendritic cells, maintaining liver tolerance.
        Blood. 2008; 112: 3175-3185
        • Yoneyama H.
        • Matsuno K.
        • Zhang Y.
        • Murai M.
        • Itakura M.
        • Ishikawa S.
        • et al.
        Regulation by chemokines of circulating dendritic cell precursors, and the formation of portal tract-associated lymphoid tissue, in a granulomatous liver disease.
        J Exp Med. 2001; 193: 35-49
        • You Q.
        • Cheng L.
        • Kedl R.M.
        • Ju C.
        Mechanism of T cell tolerance induction by murine hepatic Kupffer cells.
        Hepatology. 2008; 48: 978-990