Dendritic cells, the liver, and transplantation

Authors

  • Tina L. Sumpter,

    1. Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA
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  • Masanori Abe,

    1. Department of Gastroenterology and Metabology, Ehime University Graduate School of Medicine, To-on, Ehime, Japan
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  • Daisuke Tokita,

    1. Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA
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  • Angus W. Thomson

    Corresponding author
    1. Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA
    2. Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA
    • W1544 Biomedical Science Tower, University of Pittsburgh School of Medicine, 200 Lothrop Street, Pittsburgh, PA 15213
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    • fax: 412-624-1172


  • Potential conflict of interest: Nothing to report.

Abstract

Interstitial liver dendritic cells (DCs) exhibit phenotypic diversity and functional plasticity. They play important roles in both innate and adaptive immunity. Their comparatively low inherent T cell stimulatory ability and the outcome of their interactions with CD4+ and CD8+ T cells, as well as with natural killer (NK) T cells and NK cells within the liver, may contribute to regulation of hepatic inflammatory responses and liver allograft outcome. Liver DCs migrate in the steady state and after liver transplantation to secondary lymphoid tissues, where the outcome of their interaction with antigen-specific T cells determines the balance between tolerance and immunity. Systemic and local environmental factors that are modulated by ischemia-reperfusion injury, liver regeneration, microbial infection, and malignancy influence hepatic DC migration, maturation, and function. Current research in DC biology is providing new insights into the role of these important antigen-presenting cells in the complex events that affect liver transplant outcome. (HEPATOLOGY 2007.)

Although well-recognized as a potential site of inflammatory disease, the liver also displays inherent tolerogenic properties, as evidenced by its role in oral tolerance and the comparative immune privilege of hepatic allografts. “Spontaneous” acceptance of liver transplants, in the absence of immunosuppressive therapy, was demonstrated initially in outbred pigs,1 then across major histocompatibility complex (MHC) barriers in mice2 and in some rat strain combinations.3 In humans, liver transplants confer protection on other organ grafts from the same donor.4 Further, experimental liver transplantation can induce systemic, donor-specific tolerance.5 There are many distinctive immunologic features of the hepatic environment,6 including a constituency of various antigen (Ag)-presenting cells (APCs) and locally-produced factors that can regulate immune responses.

Resident APCs within the liver include both leukocytes (Kupffer cells [KCs] and dendritic cells [DCs]) and parenchymal cells, that is, sinusoid-lining endothelial cells, hepatocytes, and stellate (Ito) cells. These APCs occupy distinct functional niches and may display functional plasticity with ability to instigate immune responses or promote tolerance. Here, we focus on aspects of liver DC biology that relate to their roles in the induction and regulation of innate and adaptive immunity, and in liver transplant outcome. Many factors affect liver DC recruitment, maturation, function, and migration, and likely include anti-inflammatory and immunosuppressive agents (Fig. 1).7

Figure 1.

Factors affecting liver DC recruitment, maturation, function, and migration are discussed in the text. Anti-inflammatory and immunosuppressive drugs likely to affect liver DCs include aspirin, corticosteroids, calcineurin inhibitors, rapamycin, and cell proliferation inhibitors that inhibit DC differentiation, maturation, and function.7

DC Phenotype, Diversity, and Plasticity

DCs are rare, ubiquitously distributed leukocytes, derived from CD34+ hematopoietic stem cells.8 In normal liver, they are restricted largely to the perivenular region, portal space, and beneath the Glisson capsule, with a few cells scattered throughout the parenchyma9, 10 (Fig. 2A). The cytokines fms-like tyrosine kinase 3 ligand (Flt3L) and granulocyte macrophage colony-stimulating factor (GM-CSF) mobilize DCs from bone marrow. They can be used to enrich DCs in the liver (Fig. 3),11 enhancing the feasibility of liver DC studies. DCs can also be propagated from liver nonparenchymal cells12 (Fig. 2B–D).

Figure 2.

(A) Periportal distribution of MHC class II+ DCs in normal mouse liver. Magnification: ×400. Reproduced with permission from Transplantation (J. Woo et al., 1994;58:485). (B-D) Morphologic appearance of myeloid DCs (mDCs) propagated in vitro from precursors in normal mouse liver. (B,C) Transmission and (D,E) scanning electron micrographs. Note the irregular-shaped nuclei, prominent nucleoli, few electron-dense granules, numerous mitochondria, and prominent cytoplasmic veils. Bars = 1 μm. Reproduced with permission from The Journal of Experimental Medicine (L. Lu et al., 1994;179:1827).

Figure 3.

Detection of classic mDCs (CD11c+ CD11b+ CD8α) and CD11c+ CD8α+ DCs by flow cytometric analysis in normal mouse liver, and expansion of their numbers in liver in response to Flt3L administration from days 0–9.

Various cell surface markers have been employed to identify and purify rodent, nonhuman primate, and human DCs (Table 1).13–23 CD11c is a very useful marker of DCs in mice. Other Ags, such as the mannose receptor CD205 (formerly DEC205), can help identify mouse DCs. CD205 and CD103 (T.L.S., unpublished observation) are expressed at higher levels on mouse liver compared with spleen DCs. OX62, an integrin molecule, is often used to identify rat DCs. Rat liver DCs also express the receptor for adenosine triphosphate, P2X.24 Expression of receptors for other products enriched in the liver, such as liver X receptors (LXR), that are involved in lipid and cholesterol metabolism, have not been evaluated on hepatic DCs, despite high intrahepatic expression of LXR and recent evidence of its role in disruption of human monocyte-derived DC–T cell synapse formation.25 In humans, DCs are lineage HLA-DR+. DC-SIGN (DC-specific intercellular adhesion molecule-3 [ICAM-3] grabbing nonintegrin), a c-type lectin receptor, is used as a marker for immature DCs. Recently, the human DC subset-specific markers blood DC Ag-1 (BDCA-1) (CD1c), BDCA-2 (CD303), BDCA-3 (CD141), and BDCA-4 (CD304) have been identified.

Table 1. Phenotype Characteristics of Liver DC Subsets in Mice, Rats, Nonhuman Primates, and Humans
SubsetMouseRatNonhuman Primate (NHP)*HumanReferences
  • *

    Rhesus macaque.

  • Overnight migrated cells from normal liver.

  • In wedge biopsies.

  • §

    Hepatic lymph node, not liver tissue.

Myeloid DCCD11c+CD8αCD11b+MHC II+ ANAEFcR or MHCIhi MHC IIhiCD54+OX62+CD11c+HLA-DR+ fascin+CD45RAMHC IIhi CD11chi CD11bhi CD14+ CD1c+ (BDCA-1+)CD83lo or CD83+ CD1c+Mouse: Morelli et al. 200013; O'Connell et al., 200014; Pillarisetty et al., 200415; Rat: Brenan et al., 199216; NHP: Morelli et al., 200717; Human: Bosma et al., 200618; Goddard et al., 200419
CD8α+ DCCD11c+CD8α+ CD11b-Not identified in ratNot identified in NHPNot identified in humansPillarisetty et al., 200415; O'Connell et al. 200014
Plasmacytoid DCCD11cloB220+Ly6C+CD11b or PDCA-1+MHC-II+ CD45R+CD45RA CD3CD103CD11cloCD45RAhiCD11cCD123hi HLA-DR+Lin BDCA2+ (CD303+)§Mouse: Pillarisetty et al., 200415; Abe et al., 200420; Rat: Yrlid et al., 200621; NHP: Morelli et al., 200717; Human: Tanis et al., 200422
Natural Killer DCCD11c+NK1.1+Not identified in rat liverNot identified in NHPNot identified in humansLai et al., 200723; Pillarisetty et al., 200415

At least 4 DC subsets are found in mouse liver.20 Both myeloid DCs (mDCs) (CD11c+CD8α CD11b+) and CD8α+ DCs (CD11c+CD8α+CD11b) are present in normal liver15 and can be expanded strikingly by Flt3L administration.14 Mouse plasmacytoid DCs (pDCs) (Fig. 4) are CD11cloB220+Ly-6C+CD11b and produce large amounts of type-I interferon (IFN) in response to microbial stimulation.26 The incidence of liver pDCs relative to mDCs and CD8α+ DCs is higher than that in spleen.15 An additional mouse liver DC subset, the natural killer (NK)-DC,27 expresses both the NK cell marker NK1.1 and CD11c. NK-DCs present Ag while exhibiting lytic function28 and produce IFNγ via autocrine interleukin-12 (IL-12) in response to CpG (cytosine-guanine dinucleotide) stimulation.

Figure 4.

Morphological appearance of freshly-isolated, purified pre-plasmacytoid DCs from the liver of a mouse treated for 10 days with the DC poietin Flt3L. Note the round or reniform nuclei, absence of cytoplasmic projections, and a clearly defined perinuclear area that contains a well-developed Golgi and numerous rough endoplasmic reticulum cisternae. Magnification: ×1000.

Human myeloid (monocytoid) DCs are CD11c+, CD11b+ and lineage (lin) BDCA-1+ (approximately 90% of human mDCs) or CD11c+CD123 lin HLA-DR+, whereas human pDCs are BDCA-2+, BDCA-4+, or lin HLA-DR+ CD11c CD123hi or CD4+CD11c. Functionally, human mDCs and pDCs fulfill roles similar to those in mice. There is no known phenotypic counterpart of murine CD8α+ DCs in humans. The NK-DC subset has not been clearly defined in humans, though human NK cells can present Ag29 and human pDCs, like mouse pDCs, express NK cell activation markers. The mDCs have been characterized in human liver biopsies and perfusates.18 Detailed studies of pDCs within the human liver have not been performed, although a relatively small population of pDCs have been identified in human hepatic lymph nodes.22

Abbreviations

Ag, antigen; APC, antigen-presenting cell; DC, dendritic cell; Flt3L, fms-like tyrosine kinase 3 ligand; HBV, hepatitis B virus; HCV, hepatitis C virus; KC, Kupffer cell; LPS, lipopolysaccharide; mDC, myeloid DC; MHC, major histocompatibility complex; NK, natural killer; pDC, plasmacytoid DC; Th, T helper; TLR, Toll-like receptor; Treg, regulatory T cell.

DC Interactions with T Cells

DCs convey Ag, including alloAg (Fig. 5), from peripheral sites, such as the liver, via afferent lymphatics or blood to T cells in secondary lymphoid organs. In normal liver, DCs reside as “immature” APCs14, 15 that express low levels of surface MHC and costimulatory molecules (CD40, CD80, CD86) necessary for T cell activation. These immature DCs are extremely well-equipped for Ag capture, processing, and loading onto MHC class II molecules for export to the cell surface, but are poor stimulators of naïve T cells.

Figure 5.

Migration of mouse liver-derived mDCs to T cell areas of allogeneic secondary lymphoid tissue following their local subcutaneous injection. Spleen sections were stained with monoclonal antibody to donor MHC class II expressed on the migrating DCs. Magnification: ×400. Inset: ×1000. Reproduced with permission from the Journal of Experimental Medicine (L. Lu et al., 1994;179:1831).

DC maturation is essential for the initiation of acquired immune reactivity. Maturation of freshly-isolated liver DCs occurs in overnight culture14 and is induced by microbial products (for example, lipopolysaccharide [LPS] or unmethylated CpG-containing sequences [CpG]), proinflammatory cytokines (tumor necrosis factor- α [TNF-α] and IL-12) and cyclooxygenase metabolites. Maturation is triggered by signaling through TNF receptor (R) family members or ligation of Toll-like receptor (TLR) 1-13 in mice or TLR1-10 in humans.30 Upon maturation, DCs express high levels of MHC, intercellular adhesion molecules, and costimulatory molecules, and synthesize large amounts of bioactive IL-12p70. Up-regulation of cell surface CCR7 chemokine receptor expression promotes DC trafficking to T cell areas of secondary lymphoid tissues in response to CCL19 or CCL21. Interaction between DC-MHC II and the T cell receptor (TCR) on CD4+ T cells leads to proliferation of Ag-specific T helper (Th) cells and their differentiation into Th type-1 or type-2 cells. DCs also play a prominent role in inducing IL-17–producing Th cells31 and regulatory T cells (Tregs).32

Normal, liver-resident DCs are less immunostimulatory than spleen DCs in mice.15, 33, 34 In response to LPS stimulation, they produce less IL-12,33 express lower MHC class II and CD86,34 and remain comparatively poor stimulators of allogeneic T cells.33, 34 Liver mDCs, CD8α+ DCs, and pDCs up-regulate CCR7 in response to maturation and migrate to lymphoid tissue.20 Freshly-isolated mouse liver pDCs exhibit very weak allostimulatory capacity, but secrete more TNFα compared to spleen pDCs.35 Murine liver-derived mDC progenitors are weak stimulators of naïve allogeneic T cells and, following their transfer to allogeneic recipients, prolong survival of donor-derived pancreatic islet allografts.36 On the other hand, these cells have been reported to induce Ag-specific memory T cell proliferation.37 Liver-derived mDC progenitors exhibit high surface expression of CD45, CD11b, CD24, and CD44, and moderate expression of CD11c and CD205.37, 38 Injection of liver-derived mDC progenitors into allogeneic recipients elicits IL-10–producing T cells,39 whereas by contrast, functionally mature liver-derived mDCs induce IFNγ-producing cells.13 Lu et al.40 have described a unique population of liver-derived DC-like cells (CD205hiCD11cB220+CD19) with poor T cell allostimulatory capacity that can abrogate cardiac allograft rejection.

DCs also display Ag processed exogenously on MHC class I (cross-presentation or cross-priming)41 or complete MHC class I-peptide complexes acquired externally from dead cells (“cross-dressing”) to CD8+ T cells.42 DCs use the scavenger receptor to “nibble” Ag, including MHC molecules, from the surface of other live cells.43 This allows DCs to acquire peptide–MHC class I or peptide–MHC class II complexes from allogeneic DCs in vivo and in vitro through direct cell contact.44 Through the “semidirect” pathway of Ag presentation, a single DC can interact with CD4+ and CD8+ T cells simultaneously.45 Activated CD8+ T cells are found in transplanted rodent livers.46 The role of cross-presentation or semidirect Ag presentation relative to these cells is yet to be defined.

The intrinsic differences between liver DCs and other tissue-resident/tissue-derived DCs may reflect distinct features of the liver extracellular milieu. Human monocytes differentiated into DCs when cocultured with rat liver epithelial cells or liver-conditioned media secrete IL-10 but not IL-12p70, and direct Th2 rather than Th1 responses.47 These data also suggest that liver DCs may give rise to greater frequencies of forkhead box P3 (Foxp3)+ Treg cells than other tissue-derived DCs. The relative abundance of pDCs in the liver15 may also support the expansion/induction of Tregs. In vitro, mature human pDCs induce Tregs that are either TGFβ-independent and IL-10–independent suppressors,48 or IL-10–dependent suppressors.49 Interactions between alloAg-presenting pDCs and Tregs in lymph nodes promote the survival of mouse vascularized allografts.50 Induction of liver allograft survival has been attributed to Treg activity and rejection to depletion of Tregs.51 These findings contrast with a recent report suggesting that Tregs are not required for liver allograft acceptance.52

DC Interactions with NK Cells and NK T Cells

The liver is comparatively rich in NK cells and NKT cells in humans and rodents.53 Interactions between these cells and DCs may be important for promotion of immunity or tolerance. DCs present lipid Ag displayed on CD1d molecules to NKT cells promoting their activation. DCs acting with NKT cells may activate NK cells synergistically. Maturation of DCs with α-galactosyl ceramide (α-GalCer) and activation of NKT cells with α-GalCer–pulsed DCs stimulates secretion of IFNγ by the NKT cells and IL-12 by DCs, resulting in synergistic activation of NK cells.54 This orchestrated cell-cell interaction suppresses tumor growth through NK cell activation in a murine liver tumor model.55

On the other hand, NK cells can lyse donor-derived APCs, contributing to allograft tolerance.56 Costimulation pathway blockade (anti-CD154, anti-CD28, and anti-OX40) used to induce tolerance in animal allograft models is commonly thought to disrupt interactions between APCs and T cells. However, costimulation blockade fails to induce transplant tolerance in NK cell–deficient animals,56 highlighting the importance of NK cells in tolerance induction. In vitro, interactions between NK cells and hepatocytes prime liver-derived DCs to induce a type of Treg that relies on programmed death-1 (PD-1) to inhibit CD4+CD25 T cells.57 Such cell-cell interactions may be important for the induction of liver transplant tolerance.

The Liver Microenvironment, Endotoxin Tolerance, and Cross-Tolerance

Because the liver is located downstream of the gut, it is exposed constantly to endotoxin. Human liver transplantation is associated with increased levels of circulating endotoxin.58 We have shown34 that compared with spleen DCs, freshly-isolated mouse liver DCs express comparatively low levels of TLR4 messenger RNA (mRNA) and are less able to activate allogeneic T cells, or polarize naïve T cells toward Th1 responses in response to LPS. However, adoptive transfer of LPS-activated allogeneic liver DCs induces Th2 skewing.39 Moreover, IL-12 production by liver mDCs is lower than that of spleen mDCs following TLR9 or TLR7 ligation in vitro33 (Fig. 6) or in vivo. Allogeneic T cells stimulated by similarly-activated liver mDCs secrete lower levels of IFN-γ than T cells stimulated with similarly-activated spleen mDCs. Such impaired TLR-mediated responses of liver DCs reflect a phenomenon known as “endotoxin tolerance,” that is, transient hyporesponsiveness to LPS (endotoxin) that may extend to other TLR ligands (“cross tolerance”). Endotoxin tolerance in liver DCs could play a significant role in liver allograft acceptance. Mice deficient in the TLR signaling molecule MyD88,59 or both MyD88 and TRIF (Toll/IL-1R domain-containing adaptor inducing IFNβ),60 accept skin grafts more readily than wild-type controls.

Figure 6.

(A) Production of IFNγ by allogeneic T cells stimulated with CpG-activated liver mDCs from normal mice is lower than that of T cells stimulated with spleen mDCs. *P < 0.01. (B) IL-12 production by liver mDC stimulated with various concentrations of the TLR9 ligand CpG or the TLR7 ligand imiquimod is significantly lower than that of spleen mDCs. Reproduced with permission of the European Journal of Immunology (M. Abe et al., 2006;36:2487).

The lower levels of TLR4 expressed by mouse liver DCs relative to splenic DCs may also influence the pathway of alloAg presentation in the liver. Phagosomes containing Ag and TLR determine the efficiency of Ag presentation and the outcome of the ensuing immune response.61 TLR ligand-peptide conjugates improve Ag presentation by DCs.62 TLR4 ligation inhibits cross-presentation.63 With low TLR4 expression, DCs in the liver may present Ag less efficiently, but cross-present Ag more readily. How this may affect CD8+ T cell populations in the liver and transplant outcome is yet to be determined.

The balance between potential immunostimulatory and tolerogenic DC function in the liver may be an important predictor of transplant outcome. Augmentation of DCs in donor livers by Flt3L administration before transplant promotes rejection11 and has provided a valuable tool for understanding the role of DCs in graft acceptance, as well as rejection. Flt3L boosts the production of large numbers of liver DC progenitors64 and increases liver mDCs, CD8α+ DCs,13 and pDCs.35 It has been reported that mDCs11 and pDCs35 from these livers exhibit increased activation markers, such as MHC II, CD80, and CD86 and enhanced allogeneic T cell stimulatory capacity compared with hepatic DCs from untreated mice. In spite of this, the T cell allostimulatory capacity of freshly-isolated or LPS-activated bulk CD11c+ DCs from the livers of Flt3L-treated mice remains inferior to that of splenic mDCs from the same animals.33, 34

Rejection of Flt3L-treated mouse donor livers (as opposed to “spontaneous” acceptance of normal livers) may result from enhanced Th1-polarizing ability of the liver DCs following Flt3L mobilization.13 Antagonism of IL-12 in recipients of these livers abrogates rejection.65 Rejection in Flt3L-treated mice may also reflect a breach of endotoxin tolerance. Whereas hepatic DCs from Flt3L-treated animals exhibit characteristics of endotoxin tolerance, such as diminished IL-12 secretion after stimulation with LPS or CpG,33 Flt3L treatment after the development of systemic endotoxin tolerance reverses the tolerant state.66 These studies suggest that conditioning of DCs in the liver, rather than their mobilization to this site, is necessary for tolerance. Understanding the contribution of LPS resistance to the outcome of liver transplantation may provide instructive insights for promoting acceptance not just of the liver, but of other organ grafts.

DC Migration to and from the Normal and Transplanted Liver

In rats, DCs expressing CC-chemokine receptor 1 (CCR1) and CCR5 are recruited to the liver in response to macrophage inflammatory protein-1α secreted by KCs.67, 68 KCs bind to N-acetylgalactosamine on DC precursors or Ag-loaded DCs to recruit DCs from the peripheral blood into the liver.69

Unlike other liver APCs, DCs are highly motile. They migrate from rat livers at a rate of approximately 105 DCs/hour.70 Resident DCs mature during migration from the portal vein toward the central vein. These DCs then traverse the sinusoidal lumen to the hepatic lymph via the space of Disse.71 During inflammation, DCs in the space of Disse form close contacts with lymphocytes; similar interactions are observed in portal tracts.72 DCs within portal tract-associated lymphoid tissue (PALT) appear to activate T cells.73 During their maturation, hepatic DCs up-regulate CCR7 and respond to CCL21, promoting their homing to secondary lymphoid tissue.67 Following uptake of exogenous Ag, liver DCs travel via the lymph to the celiac lymph nodes,74 wherein they prime T cells. Intriguingly, in rats, hepatic pDCs are not found in the intestinal or hepatic lymph,21 suggesting that pDCs use alternative migratory routes to reach T cells, or that liver pDCs may activate T cells primarily within the PALT.

Following the severing of hepatic lymph vessels, liver DCs that presumably migrate via the blood are found in the spleen and celiac lymph nodes.75 Similar findings have been reported following allogeneic liver transplantation. Migration of donor MHC II+ cells from transplanted rat livers to host spleens is greater than migration of these cells to the spleen following transplantation of more readily rejected heart allografts.76 In rats tolerized by donor-specific transfusion (DST) prior to liver transplantation, donor hepatic DCs (OX-62+ cells) migrate to the splenic red pulp, whereas donor DCs in control rats (no DST) are found only in the splenic white pulp.77

DC chimerism and ex vivo propagation of donor-derived DCs from recipient bone marrow has been described after mouse liver transplantation.78 In a fully mismatched rat spontaneous liver allograft acceptance model (PVG→DA), donor-derived DCs are found in recipient celiac lymph nodes79 shortly after transplantation. These nodes express elevated IL-2 mRNA and the lymphocytes display increased CD25 compared to those in nodes draining skin grafts.79 In the secondary lymphoid organs, donor leukocyte migration is associated with increased T cell apoptosis in animals receiving spontaneously accepted liver grafts compared with more readily rejected kidney grafts.80 Thus, donor-derived DCs that present Ag via the direct pathway of allorecognition may play important roles in the regulation of early liver transplant outcome. Less insight is available regarding the role of these APCs and host-derived DCs in the context of late alloimmune responses and graft acceptance, for example, whether allospecific Tregs capable of maintaining tolerance may be induced/expanded via the indirect pathway.

DCs and Liver Dysfunction

In patients with hepatocellular carcinoma (HCC), monocyte-derived DCs exhibit impaired T cell allostimulatory capacity and IL-12 production.81 These DCs remain resistant to maturation in the presence of TNFα. In HCC, the frequency of circulating mDCs and pDCs is reduced, and human leukocyte antigen (HLA)-DR and costimulatory molecule expression on both subsets is decreased.82 These findings are associated with increased serum IL-10 levels and with tumor progression, suggesting that the tumor environment may affect DC function in patients with HCC. Chen et al.83 have shown that CD83+ (mature) DCs in liver biopsies are decreased significantly in patients with HCC and that HCC tumor nodules lack CD83+ DCs. These deficiencies may aid tumor immune escape.

Chronic alcohol consumption and infection with hepatitis C virus (HCV) or hepatitis B virus (HBV) are associated with progressive liver disease, including liver cirrhosis and HCC. In mice, chronic alcohol consumption impairs the maturation and function of liver DCs.84, 85 Several groups have studied the functions of DCs in patients with chronic hepatitis due to HCV. Circulating mDCs and pDCs are reduced substantially in these patients86–88 and accumulate in the liver.87 Changes in DC distribution and migration may be associated with interactions between the viral E2 protein and DC CD81.87 Impairment of IL-12 production and of allogeneic T cell stimulatory capacity by mDCs,92 together with reduced IFN-γ86, 88, 90 and increased IL-1088 production by T cells, have also been associated with HCV. IFNα production by circulating pDCs is also impaired in HCV infection.91 Very recently, Lai et al.23 have reported that mDCs from HCV-infected liver express higher MHC II, CD86, and CD123, are more efficient stimulators of allogeneic T cells, and secrete less IL-10 than do mDCs from noninfected inflamed liver. Similar perturbations have been reported for mDCs or pDCs following HBV infection.92 Defects in DC–T cell allostimulatory capacity have been restored after antiviral treatment in HCV-positive patients who have not undergone transplantation.90 These findings indicate that impaired DC function may contribute to viral persistence and disease chronicity. Higher proportions of all subsets of intrahepatic DCs have been associated with decreased viral load in HBV infection.93

Decreases in number of circulating DCs early after liver transplantation may favor HCV recurrence.94 Reinfection with HBV or HCV occurs frequently in liver transplant patients. HCV-infected patients undergoing treatment for acute allograft rejection are at higher risk for HCV-related disease, such as cirrhosis.95 This is associated with diminished populations of circulating DCs after transplantation.94 These data suggest that suppression of peripheral DC responses may be a means of immune evasion for both HCV and HBV. Overriding the natural “tolerogenic” state of hepatic DCs may be important for controlling viral infection. Additionally, antiviral therapies, such as HBV-immunoglobulins, that improve viral clearance, whereas inhibiting DC–T cell interactions are correlated with decreased incidence of acute rejection following liver transplantation.96

DCs and Liver Ischemia-Reperfusion Injury

Ischemia-reperfusion injury (IRI) occurs during organ procurement. During IRI, DCs surround the portal vein, enter the liver parenchyma, and exhibit increased surface CD80, CD86, and CD1d expression; increased IL-10 and TGF-β RNA expression; and decreased IL-12p40 message.97 This suggests that liver DCs may inhibit IRI through suppressive cytokines. Another report,98 however, suggests that increasing the number of DCs in the liver can exacerbate IRI mediated by the endogenous danger signal high motility group box 1 (HMGB1).98

Preservation injury of the liver increases circulating LPS in rodents and enhances the activity of transcription factors (nuclear factor κB [NFκB] and AP-1) activated by TLR signaling.99 Consistent with increased transcriptional activity of NFκB, IRI is associated with increased IL-6 and TNFα mRNA expression in the liver, which is dependent on TLR4 but not TLR2.100 An elegant study using chimeric mice with inactive TLR4 on hepatocytes, but wild-type TLR4 on bone marrow-derived cells, including macrophages and DCs, showed that TLR4 activity on leukocytes was necessary for IRI to occur.101 HMGB1 is also elevated during IRI, and neutralizing HMGB1 diminishes tissue damage.102 HMGB1 increases TLR4 expression on DCs98 and interacts with the receptor for advanced glycation end-products (RAGE) and TLR9103 to activate DCs. RAGE activation also enhances IRI-mediated liver injury in mice.104 These data suggest that during IRI, liver DCs may adopt a proinflammatory role to perpetuate tissue damage.

DCs and Liver Regeneration

Unlike other solid organs, the liver can regenerate rapidly following injury. This allows the use of smaller size liver grafts than native liver and partial liver transplantation from live donors. Alterations in the extracellular environment during liver regeneration may alter the size and function of the resident DC population. Castellaneta et al.105 reported increased murine liver DC populations without surface phenotypic changes after partial hepatectomy. The DCs up-regulated IL-10, down-regulated IFNγ mRNA, and induced Th2 cytokine production by naïve allogeneic T cells. Additionally, estrogen receptors were expressed on the hepatic DCs during liver regeneration, corresponding to increased serum estradiol, regardless of the animal's gender. Increasing liver DC numbers by Flt3L administration accelerated liver regeneration. These data suggest that liver DCs may play a role in local immune regulation to support liver regeneration. In a mouse model of massive hepatectomy, liver DCs up-regulated expression of RAGE.106 Blocking of RAGE with a soluble decoy receptor promoted liver regeneration and increased survival. MyD88-mediated signaling initiated by TLRs is necessary for liver regeneration.107, 108 Therefore, given the association between RAGE and TLR9 activation, the RAGE decoy receptor may alter the balance between DC-mediated immunity and tolerance.

Immune Monitoring and DCs in Liver Transplant Recipients

Given the role of DCs in regulation of alloimmunity, monitoring of DC subsets and their activation status or cytokine production may provide a useful tool for predicting allograft outcome.109 Following liver transplantation, circulating mDCs and pDCs decrease initially, then rebound to pretransplant values.110 Analysis of peripheral blood DC subsets in pediatric liver transplant recipients revealed elevations in pDCs relative to mDCs in clinically tolerant patients compared with those on maintenance immunosuppression, independent of the type or extent of immunosuppressive therapy.111 The altered cytokine levels identified by Gras et al.112 in pediatric liver transplantation may reflect such alterations in peripheral DC subsets.

Conclusion

Liver DC subsets appear to play critical roles in regulation of innate and adaptive immunity. Their responses to local environmental factors, interactions with other leukocyte populations in the liver, and the outcome of their interactions with T cells in the liver and secondary lymphoid tissue likely influence liver transplant outcome. This outcome may also be affected by the role of DCs in resistance to microbial infection. Much remains to be learned about the functional biology of these important liver APCs, especially in humans.

Acknowledgements

We thank lab group members and collaborators past and present for their invaluable contributions to these studies, Miriam Freeman for excellent administrative support and Rich James for generating Fig. 1.

Ancillary