Liver: An organ with predominant innate immunity*

Authors

  • Bin Gao,

    Corresponding author
    1. Section on Liver Biology, Laboratory of Physiologic Studies, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD
    • Section on Liver Biology, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, 5625 Fishers Lane, Room 2S-33, Bethesda, MD 20892
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    • fax: 301-480-0257

  • Won-Il Jeong,

    1. Section on Liver Biology, Laboratory of Physiologic Studies, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD
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  • Zhigang Tian

    1. Institute of Immunology, School of Life Sciences, University of Science and Technology of China, Hefei, China
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  • *

    Potential conflict of interest: Nothing to report.

Abstract

Blood circulating from the intestines to the liver is rich in bacterial products, environmental toxins, and food antigens. To effectively and quickly defend against potentially toxic agents without launching harmful immune responses, the liver relies on its strong innate immune system. This comprises enrichment of innate immune cells (such as macrophages, natural killer, natural killer T, and γδ T cells) and removal of waste molecules and immunologic elimination of microorganisms by liver endothelial cells and Kupffer cells. In addition, the liver also plays an important role in controlling systemic innate immunity through the biosynthesis of numerous soluble pathogen-recognition receptors and complement components. Conclusion: The liver is an organ with predominant innate immunity, playing an important role not only in host defenses against invading microorganisms and tumor transformation but also in liver injury and repair. Recent evidence suggests that innate immunity is also involved in the pathogenesis of liver fibrosis, providing novel therapeutic targets to treat such a liver disorder. (HEPATOLOGY 2007.)

The liver is the largest solid organ in the body with dual inputs for its blood supply. It receives 80% of its blood supply from the gut through the portal vein, which is rich in bacterial products, environment toxins, and food antigens. The remaining 20% is from vascularization by the hepatic artery. Seventy percent of the cell number or 80% of the liver volume is composed of hepatocytes that fulfill the metabolic and detoxifying needs of the body. The remaining cells are made up of nonparenchymal cells, including endothelial cells, stellate cells, Kupffer cells, and lymphocytes. Emerging evidence suggests that the liver is an important part of the body's immune response and is therefore considered an immunologic organ.1 In this review, evidence is presented demonstrating that the liver plays a key role in innate immune defenses against pathogens, which supports the notion that the liver is an organ with predominant innate immunity and acts as an organ barrier or a filter between the digestive tract and the rest of the body (see details below). Moreover, additional evidence suggests that innate immunity is also involved in the pathogenesis of liver fibrosis, which will also be discussed.

Innate Immunity

Innate immunity is an important first line of defense against infection, quickly responding to potential attacks by pathogens. It comprises physical barriers (for example, skin and mucous membranes), chemical barriers (urine, vaginal secretions, and hydrochloric acid in the stomach), humoral factors (complements and interferons [IFNs]), phagocytic cells (neutrophils and macrophages), and lymphocytic cells (natural killer [NK] and natural killer T [NKT] cells). Many of these barriers can kill pathogens nonspecifically. However, recent evidence suggests that innate immunity can also specifically detect infection through pattern-recognition receptors (PRRs) that recognize specific structures, called pathogen-associated molecular patterns (PAMPs), that are expressed by invading pathogens.2 The best defined PAMPs include lipopolysaccharide (LPS) found on gram-negative bacteria and peptidoglycan on gram-positive bacteria. The PRRs can be divided into 3 categories: secreted PRRs, membrane-bound PRRs, and phagocytic PRRs. Secreted PRRs are a group of proteins that kill pathogens through complement activation and opsonization of microbial cells for phagocytosis. Some secreted PRRs also have direct bactericidal effects on bound bacteria. The best examples of secreted PRRs include complements, pentraxins, peptidoglycan-recognition proteins, and lipid transferases, which are mainly produced by hepatocytes and secreted into the blood stream (Table 1). Membrane-bound or intracellular PRRs include the toll-like receptor (TLR) family of proteins,3 the recently identified nucleotide-binding oligomerization domain (NOD)–like receptors, and the retinoic acid-induced gene I (RIG)-like helicases.4 Phagocytic (or endocytic) PRRs, which are expressed on the surface of macrophages, neutrophils, and dendritic cells, can bind directly to pathogens, and this is followed by phagocytosis into lysosomal compartments and elimination. These phagocytic PRRs include scavenger receptors, macrophage mannose receptors, and β-glucan receptors.

Table 1. Biosynthesis of Cs, SPRRs, and APPs of the Innate Immune System by Hepatocytes
SPRRs and APPsMainly Synthesized in HepatocytesFunctions
  1. The references for Table 1 are listed in the supplementary material.

  2. Abbreviations: α1-CPI, α1-cysteine proteinase inhibitor (thiostain); α2M, α2-macroglobulin; AAP, acute phase protein; AAT, antitrypsin; ACT, antichymotrypsin; B, factor B; C1-INH, C1 inhibitor; CRP, C-reactive protein; Cs, complements; H, factor H; I, factor I; LBP, lipopolysaccharide-binding protein; LEAP, liver-expressed antimicrobial peptide; LPS, lipopolysaccharide; MAp19, mannan-binding lectin-associated protein 19; MASP, mannan-binding lectin-associated serine protease; MBL, mannan-binding lectin; MD2, myeloid differentiation factor-2; PGLYP2, peptidoglycan-recognition protein-2; PGRP, peptidoglycan-recognition protein; SAP, serum amyloid P; SPRR, secreted pattern-recognition molecule; TLR, toll-like receptor.

CsClassicalC1r/s, C2, C4, C4bpActivate C classical pathway
 AlternativeC3, BActivate C alternative pathway
 LectinMBL, MASP1, MASP2, MASP3, MAp19Activate C MBL pathway
 TerminalC5, C6, C8, C9Terminal C components
 RegulatorsI, H, C1-INHInhibit C activation
SPRRsPentraxinsCRP, SAPBind microbes and subsequently activate Cs to kill microbes
 Lipid transferasePGRPssCD14LBPPGLYP2Soluble CD14Binds LPS and subsequently transfers LPS to a receptor complex (TLR4/MD2) via a CD14-enhanced mechanismAntibacterial protein via digestion of peptidoglycan on the bacterial wall
   Stimulates or inhibits LPS signaling dependent on its concentration and environment
Other APPsAntimicrobial peptideHepcidin (also LEAP)Antimicrobial peptide by limiting iron availability
 Clotting factorsFibrinogenA central regulator of the inflammatory response
 Proteinase inhibitorsAAT, ACT, α1-CPI, α2MInactivate proteases released by pathogens and dead or dying cells
Abbreviations

α1-CPI, α1-cysteine proteinase inhibitor; α2M, α2-macroglobulin; AAP, acute phase protein; AAT, antitrypsin; ACT, antichymotrypsin; C1-INH, C1 inhibitor; CRP, C-reactive protein; CTC, connective tissue component; HBV, hepatitis B virus; HCV, hepatitis C virus; HSC, hepatic stellate cell; IFN, interferon; Ig, immunoglobulin; IL, interleukin; LBP, lipopolysaccharide-binding protein; LEAP, liver-expressed antimicrobial peptide; LPS, lipopolysaccharide; MAp19, mannan-binding lectin-associated protein 19; MASP, mannan-binding lectin-associated serine protease; MBL, mannan-binding lectin; MHC, major histocompatibility complex; NK, natural killer; NKT, natural killer T; NOD, nucleotide-binding oligomerization domain; NS, nonstructural protein; PAMP, pathogen-associated molecular pattern; PGLYP2, peptidoglycan-recognition protein-2; PGRP, peptidoglycan-recognition protein; PRR, pattern-recognition receptor; RAE-1, retinoic acid early inducible gene 1; SAP, serum amyloid P; SPRR, secreted pattern-recognition molecule; TCR, T cell receptor; TGF-β, transforming growth factor β; TLR, toll-like receptor; TNF-α, tumor necrosis factor α; TRAIL, tumor necrosis factor–related apoptosis-inducing ligand.

Hepatocytes

Biosynthesis of 80% to 90% of Complement Components and Secreted PRRs of the Innate Immune System.

Hepatocytes play a key role in controlling systemic innate immunity via production of secreted PRRs and complement components found in plasma (Table 1). Expression of the genes encoding these proteins is controlled by liver-specific transcription factors, such as hepatocyte nuclear factors, nuclear factor-1, and CCAAT-enhancer-binding protein, which account for their liver-specific expression. During an acute phase or systemic inflammatory response, a variety of proinflammatory cytokines [such as interleukin-6 (IL-6), IL-1, tumor necrosis factor α (TNF-α), and IFN-γ] can stimulate hepatocytes to produce high levels of complements and secreted PRRs. Alterations in the normally stable plasma levels of these innate proteins occur in liver diseases, resulting in increased incidence of microbial infections. Recently, an elegant study showed that transplant patients receiving donor livers with a genetic predisposition to lowered production of secreted PRRs had a higher risk for bacterial infections post transplantation, providing an unequivocal demonstration for the important role of hepatocytes in systemic innate immunity against infection.5

Complements.

The complement system consists of more than 35 plasma or membrane proteins that interact with one another in a cascading fashion to protect against infections. Three different pathways have been identified that activate the complement system. These include the classical pathway (target-bound antibody), the lectin pathway (microbial repetitive polysaccharide structures), and the alternative pathway (recognition of other foreign surface structures). After activation, the complement system generates a wide range of biologic activities such as opsonic, inflammatory, and cytotoxic functions. The liver (primarily hepatocytes) is a major site that biosynthesizes complement components found in plasma (Table 1). These include C1r/s, C2, C4, and Cbp of the classical pathway, C3 and factor B of the alternative pathway, mannan-binding lectin, mannan-binding lectin-associated serine proteases 1-3, and mannan-binding lectin-associated protein 19 of the lectin pathway, and terminal components C5, C6, C8, and C9 of the complement system.6, 7 Although immune cells and endothelial cells also produce these components, their contributions are minor compared to those of hepatocytes.6 Additionally, hepatocytes are also primarily responsible for the biosynthesis of several complement regulator proteins found in plasma, such as factor I, factor H, and the C1 inhibitor.6 In contrast, the membrane-bound complement regulators are expressed ubiquitously in all tissues.6, 7 In addition to being an important part of innate defenses against infection, the complement system also contributes to the pathogenesis of a variety of liver disorders, including liver fibrosis, alcoholic liver disease, liver ischemia/reperfusion, and liver transplantation.7–9 However, the molecular mechanisms underlying the involvement of complements in liver injury and repair remain obscure.

Secreted PRRs.

Hepatocytes are also the major sources for production of secreted PRRs, which have two main functions: complement activation and microbial cell opsonization for phagocytosis. Four major classes of soluble PRRs have been identified according to their domain composition: collectins, pentraxins, lipid transferases, and peptidoglycan-recognition proteins. Many of these proteins are synthesized mainly in hepatocytes and released into the bloodstream, thereby playing an important role in innate immunity against local and systemic microbial infection (see Table 1). In addition, the liver is also a major source of many other acute phase proteins, which play key roles in innate defenses against infection and in reducing tissue damage through inactivation of proteinases released by pathogens and dead or dying cells.

Liver

Expression of Membrane-Bound PRRs of the Innate Immune System.

The liver not only is the major source of secreted PRRs but also expresses membrane-bound PRRs, such as TLRs. TLRs are a family of proteins that recognize PAMPs expressed by microorganisms, but not by eukaryotes. TLRs can also be activated by endogenous signals such as uric acid activation of the NALP3/ASC/Caspase 1 (NALP3/ASC: pyrin domain-containing protein 3/apoptosis-associated speck-like protein containing a caspase activation and recruitment domain) and apoptotic mammalian DNA activation of TLR9.3 There are 13 different TLRs identified so far. Each of them recognizes specific PAMPS and activates specific signaling pathways and antimicrobial responses.3 Liver cells express a variety of TLRs,10 which have been shown to participate in liver injury and repair, and contribute to the pathogenesis of a variety of liver diseases.11, 12 However, the role of TLRs on liver cells in host defenses against invading pathogens is less clear. The TLR4 protein has been detected on all types of liver cells and is likely involved in the uptake and clearance of endotoxins, production of proinflammatory and anti-inflammatory cytokines, and generation of reactive oxidative stress.11, 12 Additionally, hepatocytes express messenger RNAs for all other TLRs,10 the functions of which on hepatocytes remain to be determined. Expression of functional TLR2 has been reported in Kupffer cells, stellate cells, and sinusoidal endothelial cells, and activation of TLR2 leads to production of proinflammatory cytokines.11, 12

Recently, several other cytoplasmic PRRs have been identified, including NOD-like receptors and the RIG-like helicases.4 Among them, RIG-1 serves as a pathogen receptor to regulate cellular permissiveness to hepatitis C virus (HCV) replication13; however, HCV nonstructural protein 3/4A (NS3/4A) blunts RIG-1/mitochondrial antiviral signaling protein (MAVS) signaling, leading to persistent infection.14, 15 Additional details about the effects of HCV infection on PRRs are described in Table 2. In addition, activation of several TLRs has been shown to inhibit hepatitis B virus (HBV) and HCV infection, providing novel strategies to treat hepatitis viral infection.16, 17

Table 2. Effects of HCV Infection on PRRs
HCVEffects of HCV on PRRs
  1. References for Table 2 are listed in the supplementary material.

  2. Abbreviations: HCV, hepatitis C virus; NS, nonstructural protein; PRR, pattern-recognition receptor; TLR, toll-like receptor; MAVS, mitochondrial antiviral signaling protein; MyD88, myeloid differentiation protein-88; MD-2, myeloid differentiation factor-2; TRIF, TIR domain-containing adapter-inducing interferon; RIG-I, retinoic acid-induced gene I.

HCV infection HCV infection increases expression of TLRs 2, 6, 7, 8, 9, and 10 and MD-2 messenger RNA levels in both monocytes and T cells and increases expression of TLR4 in T lymphocytes and TLR5, CD14, and MyD88 expression in monocytes.
HCV proteinsCoreCore protein activates TLR2, involvement of TLR1/6.
 NS3NS3 activates TLR2, involvement of TLR1/6.
 NS4ANS3/4A inhibits TLR3 signaling via cleavage of the TLR3 adaptor protein TRIF.
  NS3/4A blunts RIG-I/MAVS activation.
 NS5BNS5B activates TLR3 signaling

Elimination of Soluble Macromolecules via Sinusoidal Endothelial Cells and Elimination of Insoluble Waste via Kupffer Cells.

The liver is the major site for removing circulating macromolecules and microorganisms from the systemic circulation through the hepatic reticuloendothelial system, which is composed of sinusoidal endothelial cells and Kupffer cells. The latter accounts for 80% to 90% of the total population of fixed tissue macrophages in the body. Sinusoidal endothelial cells are mainly responsible for removal of soluble macromolecular and colloidal waste (smaller than 100 nm) from the circulation by endocytosis through 5 types of endocytosis receptors (Table 3), whereas Kupffer cells are responsible for elimination of insoluble waste by phagocytosis through a variety of receptors (Table 3).

Table 3. Expression of PRRs on Liver Sinusoidal Endothelial Cells and Kupffer Cells
Liver Endothelial Cells*
  • References for Table 3 are listed in the supplementary material.

  • Abbreviations: Cs, complements; CTC, connective tissue component; Ig, immunoglobulin; LPS, lipopolysaccharide; PRR, pattern-recognition receptor; R, receptors.

  • a

    Liver endothelial cells are exclusively responsible for endocytosis of soluble macromolecules and collides that are smaller than 100 nm.

  • Kupffer cells are mainly responsible for phagocytosis of insoluble particles and also contribute to endocytosis of some soluble macromolecules.

Receptors (R) Functions
R for CTCHyaluronan REliminates major matrix polysaccharides and proteoglycans such as hyaluronan and chondroitin sulfate
 Collagen REliminates collagen α chains of several types of collagen
 Fibronectin RInvolvement of cell attachment but not for endocytosis
Scavenger R Removal of physiological and foreign waste macromolecules, including LPS, intracellular macromolecules, modified serum proteins, and microbial proteins
Mannose R Clearance of a large number of molecules with mannosyl residues
Fc-γ R Eliminate IgG-antigen complexes
Kupffer Cells
Receptors (R) Functions
R for CsC5aRStimulates Kupffer cells to produce prostanoid and proinflammatory cytokines
 C3R, CR1, CR3, CR4, CRIgPlay a key role in Kupffer cell clearance of C3-opsonized immune complexes, IgM-opsonized E and β-glucans, and therapeutic β-glucan polysaccharides
Scavenger R: SR-AI, SR-AII Endocytosis of Ac LDL and Mal-BSA; phagocytosis of gram-negative and gram-positive bacteria through recognition of LPS and LTA, respectively; phagocytosis of apoptotic cells and red blood cell–derived vesicles
Mannose R Clearance of a large number of molecules with mannosyl residues
Fc-γ R Eliminate IgG-antigen complexes

One of the Richest Sources for Innate Immune Cells: NK, NKT, and T Cell Receptor γδ (TCRγδ) T Cells.

Liver lymphocytes are abundant in NK, NKT, and TCRγδ T cells (see Table 4).18, 19 NK cells represent a third class of lymphocytes distinct from B and T cells and do not express a clonally distributed antigen receptor that is subject to somatic diversification. Mouse liver lymphocytes contain about 10% NK cells, whereas rat and human liver lymphocytes contain about 30% to 50% NK cells. The functions of NK cells are controlled by a balance of signals from the stimulatory and inhibitory receptors expressed on NK cells. Stimulatory receptors can be activated by stimulatory ligands expressed on infected, transformed, or stressed cells, whereas binding of inhibitory receptors to self class I major histocompatibility complex (MHC) molecules leads to inhibition of NK cell function.20 Hepatic NK cells, originally termed Pit cells in rats, are not only enriched in the liver but also naturally activated as they show higher cytotoxicity against tumor cells than splenic or peripheral blood NK cells in rodents21 and in humans.22 This may be due to an up-regulated expression of tumor necrosis factor–related apoptosis-inducing ligand (TRAIL) or perform/granzymes on liver NK cells compared with peripheral NK cells.

Table 4. The Liver Lymphocytes Are Enriched in Innate Immune Cells
LiverMiceRatsHumans
  1. References for Table 4 are listed in the supplementary material.

  2. Abbreviations: NK, natural killer; NKT, natural killer T; and TCRγδ, T cell receptor γδ; Ac LDL, acetylated low-density lipoprotein; Mal-BSA, maleylated bovine serum albumin; LTA, lipoteichoic acid; Fc-γ, immunoreceptor Fc gamma.

NKMarkerNK1.1+CD3, DX5+NK1P+CD3CD56+CD3
 % liver lymphocytes5%-10%30%-50%30%-50%
NKTMarkerNK1.1+CD3+NK1P+CD3+CD56+CD3+
 % liver lymphocytes30%-40%5%-10%5%-10%
 Marker: classicalCD1dCD1dCD1d
 % liver lymphocytes10%-30%<1%<1%
TCRγδ T cellsMarkerTCRγδ+TCRγδ+TCRγδ+
 % liver lymphocytes3%-5%3%-5%3%-5%

Over the past several years, many studies have shown that hepatic NK cells play an important role in innate immune responses against tumors, viruses, intracellular bacteria, and parasites. The antitumor effects of NK cells in the liver have been well documented in a variety of experimental liver tumor models.23 Clinical studies also suggest that NK cells contribute to innate defenses against primary liver tumors and liver metastases in patients. It has been reported that hepatic NK cell numbers are greatly elevated in patients with hepatic malignancy, accounting for up to 90% of all hepatic lymphocytes. Moreover, the reduced activity of NK cells in patients appears to be associated with the progression of hepatocellular carcinoma. The antitumor action of NK cells in the liver is likely mediated via direct killing of tumor cells and induction of tumor-specific immunity. The antiviral effect of NK cells has been well documented in animal models infected with several viruses, particularly murine cytomegalovirus. However, the role of NK cells in human HBV and HCV infections is less clear because of a lack of suitable small-animal models. Studies using transgenic mice overexpressing HBV genome suggest that NK cells inhibit HBV replication in vivo through production of IFNs and direct killing of infected hepatocytes.24In vitro culture experiments showed that NK cells inhibit HCV replication in human hepatoma cells via an IFN-γ–dependent mechanism. Recently, a retrospective study revealed that individuals with a genetic predisposition to enhanced NK function had greater chances of spontaneously clearing HCV during acute infection,25 suggesting that NK cells play an important role in early antiviral defenses against HCV. In contrast, HCV can escape the antiviral response of NK cells by inhibiting NK cell function, and this results in chronic HCV infection in the majority of patients. Finally, activation of NK cells has also been implicated in liver injury, fibrosis, and repair.26–28 Taken together, these data show that NK cells not only play an important role in innate response against pathogens in the liver but also contribute to the pathogenesis of liver disease.

NKT cells are a subset of lymphocytes that express both αβ TCR (T cell marker) and cell surface receptors characteristic of NK cells (NK1.1 in C57BL/6 mice). Among them, classical NKT cells are controlled developmentally by β2-microglobulin–associated nonpolymorphic CD1d. Classical NKT cells are reactive to lipid antigen (α-galactosylceramide) and produce both type I and type II cytokines. Nonclassical NKT cells are CD1d-independent and produce only type I cytokines.29 NKT cells have been suggested to play important roles in linking innate immunity with adoptive immunity, antiviral defenses, antibacterial defenses, and antitumor defenses in the liver.29, 30 Mouse liver lymphocytes contain about 20% to 30% NKT cells, which are further elevated to 50% to 60% after partial hepatectomy or liver ischemia reperfusion.26 NKT cells appear to be involved in induction of liver injury in these models as well as other models of liver injury induced by concanavalin A, α-galactosylceramide, alcohol, and drugs.31

TCRγδ T cells represent a minority of T cells in lymphoid organs and peripheral blood, but a high percentage of γδ T cells is found in the intraepithelial lymphocytic compartments of skin, intestine, and genitourinary. Interestingly, liver lymphocytes are also enriched in γδ T cells. In normal mouse livers, γδ T cells account for 3% to 5% of total liver lymphocytes but 15% to 25% of total liver T cells, and the liver is one of the richest sources of γδ T cells in the body. The percentage of γδ T cells in the liver is significantly increased in the liver of tumor-bearing mice. Elevation of γδ T cells was also found in the livers of patients with viral hepatitis infection, but not in patients with nonviral hepatitis. However, the role of γδ T cells in the liver has not been paid much attention. Emerging evidence suggests that γδ T cells may play a prominent role in innate defenses against viral and bacterial infection and against tumor formation.32 Thus, elevated γδ T cells in the liver may also play an important role in innate defenses against pathogens and transformed cells.

As shown in Table 4, the distribution of innate immune cells in the liver is different in mice and humans. For instance, mouse liver lymphocytes contain about 30% to 40% NKT cells, whereas human liver lymphocytes contain about 5% to 10% NKT cells. In addition, unlike human NKT cells, murine NKT cells express CD28 molecules constitutively, which are important costimulatory molecules found on T cells.33 These findings suggest that mice may be more sensitive to NKT-mediated liver injury, whereas humans may be resistant to such injury. Indeed, treatment with α-galactosylceramide, an NKT activator, was shown to induce liver injury in mice,34 but results from a phase I clinical study revealed that α-galactosylceramide injection failed to produce signs of liver injury in humans. In contrast, it could be speculated that NK cells play more important roles in human liver diseases than in murine liver disease. For example, because NK cells inhibit liver fibrosis in mice27, 28 and human liver lymphocytes contain more NK cells, it is likely that NK cells may be more effective in inhibiting liver fibrosis in humans than in mice.

A Major Site To Induce T Cell Tolerance.

Contrary to expressing strong innate immunity, the liver is also a major site of induction of T cell tolerance as evidenced by the spontaneous acceptance of liver allografts, the persistence of some liver pathogens (HBV, HCV, and malaria), and the induction of oral tolerance to food antigens. Studies from many laboratories suggest that a variety of cell types, cytokines, and innate immunity components in the liver synergistically or additively work together within the unique environment of the liver to induce T cell tolerance, thereby resulting in hepatic tolerance.35–37

Sterile Inflammatory Response in the Liver

In addition to critical roles in host defenses against infection, the innate immune system can also sense danger signals from damaged hepatocytes during non–infection-related liver injury, resulting in an inflammatory response. This so-called sterile inflammation not only contributes to liver injury but conversely may also be involved in liver repair. For example, acetaminophen hepatotoxicity and ischemia/reperfusion liver injury are associated with sterile neutrophilic inflammation, which contributes to liver injury,38, 39 but on the other hand, sterile neutrophilic inflammation after partial hepatectomy can promote liver regeneration by triggering a local inflammatory response leading to Kupffer cell–dependent release of TNF-α and IL-6, eventually leading to hepatocyte proliferation.40 Moreover, an accumulation of NK and NKT cells has also been observed in several models of liver injury induced by acetaminophen, ischemia reperfusion, and partial hepatectomy, which appears to contribute to liver injury and impaired liver regeneration in these models.26, 31, 41–43 Although it has been well documented that the innate immune system detects infection via recognition of PAMPs expressed by pathogens, the molecular mechanisms underlying the sterile inflammatory response in the liver have just begun to reveal themselves. It was shown recently that IL-1 is an important mediator of sterile neutrophilic inflammation during acetaminophen-induced and ischemia/reperfusion-induced liver injury, but it is less important in microbial stimulus–induced neutrophilic inflammation, providing a novel therapeutic target to treat sterile inflammation without markedly increasing susceptibility to infection.44, 45 More extensive studies to investigate the underlying mechanisms of sterile inflammatory responses in the liver are required that may help to identify novel therapeutic targets to treat liver disease.

Innate Immunity, Stellate Cells, and Liver Fibrosis

Regardless of etiology, all chronic liver diseases lead to liver fibrosis, which is characterized by hepatic stellate cell (HSC) activation and subsequent overproduction and accumulation of collagens in the liver.46, 47 HSCs are generally quiescent in normal healthy livers, but during liver injury, they become activated and differentiate into myofibroblastic cells that are characterized by a loss of vitamin A (retinol) and enhanced collagen expression.46, 47 In the last several decades, multiple cytokines and growth factors have been identified to control HSC activation and liver fibrogenesis. Among them, transforming growth factor β (TGF-β) and platelet-derived growth factor are the most important factors that promote HSC transformation and proliferation.46, 47 Recent evidence suggests that a variety of innate immunity components also play an important role in regulating HSC activation and liver fibrosis.

The complement system is typically activated after liver injury. A recent study demonstrated clearly that C5 and C5aR contribute to the pathogenesis of liver fibrosis because C5 deficiency resulted in lowered liver fibrosis, whereas overexpression of the C5 gene resulted in increased liver fibrosis.8 Consistently, genetic analyses also suggest that human C5 gene variants are associated with liver fibrosis in HCV patients.8 At present, the molecular mechanisms by which the C5 contributes to liver fibrosis are not fully understood and require further studies.

Because a variety of TLRs are expressed on liver cells including HSCs,10–12 TLRs likely play an important role in the pathogenesis of liver fibrosis. Activated HSCs express TLR receptors and respond to stimulation by TLR ligands such as LPS, lipoteichoic acid, and N-acetyl muramyl peptide, and this suggests that LPS and other TLR ligands may be involved in hepatic fibrogenesis via the direct targeting of HSCs.12 TLR9-deficient mice are resistant to liver fibrosis because apoptotic hepatocyte DNA activation of HSCs requires TLR9.48 Activation of TLR3 by polyinosinic:polycytidylic acid inhibits liver fibrosis by activating NK cell killing of activated HSCs and producing IFN-γ.27, 49 Thus, TLRs could be potential therapeutic targets to treat liver fibrosis.

Normal livers and injured livers are enriched in innate immune cells, which have a significant impact on hepatic fibrogenesis. Among them, Kupffer cells and NK cells have been shown to play an important role in regulating liver fibrosis, whereas other innate immune cells such as mast cells, neutrophils, and NKT cells seem to have less effect on experimental liver fibrosis.27, 46, 47 The role of Kupffer cells in liver injury and fibrosis has been extensively investigated and well documented. It is generally believed that Kupffer cells can promote stellate cell activation via the production of cytokines/growth factors (such as TGF-β) and regulate the production of metalloproteinases and their inhibitors. A recent study suggests that macrophages play a distinct and opposing role in liver fibrosis: promoting extracellular matrix accumulation during ongoing liver injury but enhancing matrix degradation during recovery.50 In contrast, NK cells seem to have only an inhibitory effect on liver fibrogenesis via multiple mechanisms.27, 28 First, NK cells directly kill activated HSCs but not quiescent HSCs. This is because activated HSCs express increased levels of NK cell–activating ligand retinoic acid early inducible gene 1 (RAE-1) and TRAIL receptors but express decreased levels of NK cell–inhibitory ligand MHC-1.27, 28, 51 Second, NK cells inhibit liver fibrosis via production of IFN-γ, which induces HSC cell cycle arrest and apoptosis in a signal transducer and activator of transcription-1–dependent manner.49 Moreover, clinical data show that NK cells from HCV patients were able to kill human HSCs and that their activity was negatively correlated with liver fibrosis in HCV patients, suggesting that activation of NK cells may have a beneficial effect by inhibiting liver fibrosis in patients.28, 52

Analogous to activation of the innate immune system by cellular apoptosis, HSCs also respond to hepatocyte apoptosis and subsequently become activated. It has been reported that hepatocyte-specific disruption of Bcl-xL leads to continuous hepatocyte apoptosis and subsequent HSC activation and liver fibrosis,53 providing in vivo evidence that hepatocyte apoptosis activates HSCs. However, the molecular mechanisms by which apoptotic hepatocytes activate HSCs have just begun to be unveiled. Watanabe et al.48 demonstrated recently that apoptotic hepatocyte DNA acts as an important mediator of HSC differentiation via a TLR9-dependent mechanism. Further studies will be required to identify other signals involved in apoptotic hepatocyte DNA activation of HSCs and determine if these signals are shared between innate immune cells and HSCs.

In addition, emerging evidence suggests that activation of HSCs may lead to activation of innate immunity. First, activated HSCs synthesize and secrete a variety of growth factors, cytokines, and chemokines, which can promote leukocyte chemotaxis and adherence and influence leukocyte activation.54 Second, activated HSCs could participate in the innate immune response via expression of TLR4.12 Lastly, activated HSCs express NK cell–activating ligand RAE1, which has been shown to modulate activated macrophages and NK cells via activation of NKG2D receptors.27, 55

Concluding Remarks

In summary, the liver is an organ with strong innate immunity contributing to the antiviral, antibacterial, and antitumor defenses within the liver. In addition, innate immunity also plays an important role in regulating liver injury, fibrosis, and regeneration, which represents novel therapeutic targets with which to treat chronic liver diseases. For example, activation of NK cells could be a new strategy to treat liver fibrosis,56 which will likely have more beneficial effects than numerous target-directed drugs that have been proposed or used experimentally in clinical trials to treat liver fibrosis.46 This is because not only can activation of NK cells kill specifically activated HSCs, thereby ameliorating liver fibrosis, but also they have beneficial effects on inhibiting viral hepatitis infection and liver tumor formation.23–25 Indeed, IFN-α, one of the strongest NK cell activators, has been shown to inhibit liver fibrosis, hepatitis virus infection, and the progression of liver cancer in HCV patients. Treatment with IFN-γ, another NK cell activator, has also been shown to have antifibrotic effects in animal models and some HBV and HCV patients. In the future, it would be very interesting to investigate whether other NK cell activators (such as IL-2, IL-15, and IL-12) have beneficial effects on ameliorating liver fibrosis in animal models and human patients.

Ancillary