Toll-Like Receptor 4 and Hepatitis B Infection: Molecular Mechanisms and Pathogenesis

Abstract

Hepatitis B virus (HBV) infection mainly causes liver disease, including inflammation, cirrhosis, and hepatocellular carcinoma (HCC). It has been documented that prolonged hepatitis B–infected patients are unable to clear HBV from hepatocytes completely. Previous investigations have suggested that various genetic and immunologic parameters may be responsible for the induction of prolonged infection forms. Toll-like receptors (TLRs), as members of pathogen recognition receptors (PRRs), play critical roles in the recognition of viruses and the induction of appropriate immune responses. Thus, TLRs may be considered as essential sensors for the recognition of HBV and the induction of immune responses against this virus. It has been documented that TLR4 plays key roles in the detection of several microbial pathogen-associated molecular pattern molecules, including bacterial lipopolysacharide (LPS), as well as endogenous ligands (damage-associated molecular pattern molecules) and subsequently activates pro-inflammatory transcription factors in either MYD88 or TRIF dependent pathways. Previous investigations have proposed that TLR4 might be involved in appropriate immune responses against HBV. Therefore, the aim of this review is to present the recent data regarding the important roles of TLR4 in HBV recognition and regulation of immune responses against this virus, and also its roles in the pathogenesis of cirrhosis and HCC as complications of prolonged hepatitis B infections.

Introduction

In humans, hepatitis B virus (HBV) is the most dangerous and prevalent infectious agent of the liver, which induces hepatitis B (7,8,32). It has been documented that long-term HBV-infected patients with chronic, asymptomatic, and occult HBV infection are unable to eradicate HBV from either hepatocytes or sera completely (5,12). Previous investigations have suggested that prolonged HBV infections are the main causes of liver cirrhosis and hepatocellular carcinoma (HCC) (40,41). Although the main mechanisms responsible for the development of prolonged forms of HBV infection are yet to be clarified, it has been hypothesized that genetic and immunologic differences between subjects who successfully clear HBV infections (clearance group) and patients who suffer from prolonged forms of hepatitis B may be the main reasons for the persistent infection (1,5,7). Toll-like receptors (TLRs) belong to the family of pathogen recognition receptors (PRR). These molecules play crucial roles in pathogen-associated molecular pattern (PAMP) and damage-associated molecular pattern (DAMP) recognition, and subsequently induce immune responses against viral infections, including hepatitis B (32).

TLR/PAMP or DAMP interactions result in various immune cell functions ranging from production of inflammatory cytokines (25), migration to the infected sites (42), and phagocytosis (24), to activation of the NADPH oxidase pathway (9). Accordingly, TLR4 activation leads to upregulation of major histocompatibility complex (MHC), inflammatory cytokines, and homing molecules via the recognition of extracellular viral PAMPs in either MYD88 or TRIF dependent pathways (27). Thus, defected expression of TLR4 may lead to impaired immune responses against HBV infection. Because patients with prolonged forms of HBV are unable to clear HBV from hepatocytes completely (4,6,31), it seems that the immune system of these patients is attenuated in some parameters. Based on the crucial roles played by TLR4 in the viral PAMPs recognition and induction of immune responses, it may be hypothesized that alterations in TLR4 expression levels may lead to HBV survival in the infected liver of long-term HBV-infected patients. Therefore, the aim of this review article is to consider recent information regarding the relationship between TLR4 and HBV infection, as well as its crucial roles in the development of prolonged hepatitis B infections. This review also presents recent information regarding the plausible mechanisms leading to alteration in expression of TLR4 and its signaling molecules in these patients.

TLR4 Signaling Pathway

TLR4, which is also known as CD284 and age-related macular degeneration10 (ARMD10), has been determined as a receptor for several microbial PAMPs. The gene of TLR4 is located on 9q33.1 and, like other TLRs, is highly conserved (45,64). The TLR4 molecule contains three domain complexes, including extracellular leucine-rich repeats (LRRs), hydrophobic transmembrane, and cytoplasmic toll/interleukin-1 receptor (TIR) domains (Fig. 1). Classically, TLR4 recognizes the microbial lipids in homodimer format, and thus activates various intracellular signaling pathways. It has been established that TLR4 interacts with various ligands, which leads to activation of intracellular signaling pathways (63). TLR4 is unique among the TLRs in its ability to combine TRIF and MYD88 adapters, which leads to transcription from pro-inflammatory cytokines such as IL-12, IL-6, TNF-α, and type I interferons (63). In the MYD88-dependent pathway, dimerization of TLR4 with myeloid differentiation 2 (TLR4-MD2) at the cell surface results in recruitment of two adaptor proteins—TIRAP (MAL) and MYD88—which results in activation of NFκB as a pro-inflammatory transcription factor and consequently production of pro-inflammatory cytokines (29). Additio-nally, TLR4 is internalized into endosomes and recruits TRAM and TRIF, which activates another transcription factor, IRF3, and induces transcription from type I interferons (30). Lipopolysacharide (LPS), as the most important TLR4 ligand, is recognized by the TLR4/CD14/MD2 complex. Following LPS secretion, LPS binding protein (LBP) binds LPS to CD14 (a glycosylphosphatidylinositol-anchored protein). CD14 transfers the LPS to MD2. MD2 is a soluble protein that is associated with the extracellular domain of TLR4 in a noncovalent format. LPS/MD-2 binding leads to a conformational change in MD-2 and results in binding of TLR4-MD-2 complex to a second TLR4 receptor. Hence, TLR4 homodimerization is assembled in intracellular signaling, which is discussed in the next section. In addition to LPS, several molecules are also recognized by TLR4 such as high-mobility group box-1, hyaluronan, heat shock protein 60, free fatty acids, allergenic nickel, and the adjuvant monophosphoryl lipid A (MPLA) (2,21). Interestingly, the proposed endogenous TLR4 ligands are not only capable of activating TLR4 directly, but could also bind and transport LPS, which increases sensitivity of cells to LPS. Therefore, it appears that many endogenous TLR4 ligands may be described as PAMP binding/sensitizing molecules (20).

FIG. 1.

The intracellular signaling of toll-like receptor 4 (TLR4) and inhibitory effects of HBV. The figure shows how TLR4/ligands interactions in cytoplasmic and endosomes membranes lead to intracellular signaling via either myeloid differentiation primary response (MYD88) or TIR-domain-containing adapter-inducing interferon-β (TRIF), respectively. The figure also shows that HBsAg indirectly suppresses TLR4 signaling via promotion of monocytes to produce IL-10. HBsAg also directly inhibits mitogen-activated protein kinase (MAPK) and activator protein 1 (AP-1) molecules. Hepatitis e antigen (HBeAg) suppresses nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation. HBV, hepatitis B virus; HBsAg, hepatitis B surface antigen; PRR, pathogen recognition receptors; RIP1, receptor-interacting protein 1; IRAK1, interleukin-1 receptor associated kinase-1; TRAM, TRIF-related adaptor molecule; TIRAP, TIR domain-containing adaptor protein; TRAF6, TNF receptor associated factor; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; TAK1, transforming growth factor b-activated kinase 1; TBK1, TANK-binding kinase 1; IKK, IκB kinase; IKKi, IκB kinase inhibitor.

It has been documented that TLR4/ligands interactions lead to the recruitment of TIR-containing adaptor molecules, including TIRAP and MYD88, which participate in stimulation of TLR4-MYD88–dependent signaling pathway, and TRAM and TRIF, which play important roles in the induction of TLR4-TRIF–dependent signaling pathway (11). In MYD88-dependent signaling pathway, TLR4/ligands interaction at the plasma membrane results in binding of TIRAP, which allows MYD88 to be recruited as an adaptor protein. Recruited MYD88 leads to phosphorylation of several intracellular signaling molecules, which results in activation of NF-κB, AP-1, and IRF5 transcription in response to several pro-inflammatory cytokines (Fig. 1) (63). Interestingly, upon internalization of TLR4 and interaction with its ligands in the endosomes, TRIF-dependent signaling pathway is activated, resulting in TRAM binding to TIR domain of TLR4 (22). TRIF is an adaptor protein that is recruited to the binding of TRAM and activates (phosphorylates) TRAF3 and RIP1 molecules (22). Phosphorylation of TRAF3 leads to activation of IRF3 and translocation into the nucleus to produce transcripts in response to type I interferons, while phosphorylation of RIP1 leads to activation of other transcription factors, including AP-1 and NF-κB (Fig. 1) (37).

TLR4 and Hepatitis B

According to the fact that expression levels of TLR4 are altered in HBV-infected patients, it appears that TLR4 plays essential roles in hepatitis B infection. For instance, Wei et al. revealed that expression levels of TLR4 were increased in the hepatocytes of chronic HBV-infected patients (57). Xing et al. demonstrated that TLR4 was upregulated on the monocytes of patients with liver cirrhosis and acute on chronic liver failure, which is described as acute decompensation of cirrhosis, liver failures and increased short-term mortality, when compared to healthy controls (62). Another study reported that expression levels of TLR4 were upregulated on monocytes of HBV-infected patients when compared with healthy controls (65). Therefore, it may be concluded that TLR4 activations not only participate in induction of immune responses but also participate in the pathogenesis of hepatitis B–related complications, including cirrhosis. Additionally, previous investigations revealed that patients with prolonged forms of HBV infection are unable to elicit appropriate immune responses against HBV. Hence, the infection persists (4,46). According to the critical roles of TLR4 in the induction of immune responses, it seems that the expression of TLR4 may be impaired in patients infected with prolonged forms of HBV. Chen et al. reported that mRNA levels of TLR4 were significantly decreased in PBMCs from chronic HBV-infected patients in a selected Chinese population (15). Another study from China revealed that the expression levels of TLR4 on peripheral monocytes, hepatocytes, and Kupffer cells did not differ between untreated HBeAg positive in comparison to HBeAg negative chronic HBV-infected patients (53). Heiberg et al. identified that chronic HBV-infected patients displayed lower IFN-α secretion in comparison to healthy controls, following stimulation with specific ligands for TLR4 (23). Therefore, it appears that TLR4 is downregulated in long-term HBV-infected patients. Some evidence shows that TLR4/ligands interaction may be a main route to activation of immune responses against HBV. For example, it has been identified that TLR4 is expressed on the hepatocytes, sinusoidal endothelial cells, biliary epithelial cells, and hepatic dendritic cells, which is capable of responses to LPS (47,51). Additionally, Wu et al. revealed that the supernatants from TLR4-stimulated Kupffer cells from C57/BL6 wild-type mice can suppress HBV replication (60). Zhou et al. reported that upregulation of TLR4 in proximal tubular epithelial cells led to inhibition of expression of HBV antigen, as well as HBV-DNA replication (67). Moreover, another study revealed that TLR4 suppresses HBV replication via upregulation of inducible nitric oxide synthase (iNOS) (13,14). Isogawa et al. also demonstrated that administration of ligands specific for TLR4 results in inhibition of HBV replication (26). Furthermore, the pivotal roles played by TLR4 in promotion of homing of HBV specific CD8+ T-lymphocyte to the infected liver were reported by John et al. (28). It has also been evidenced that LPS level in serum has a positive relation with the severity of clinical presentation in HBV-infected patients (43). Therefore, it appears that TLR4 plays important roles in the induction of appropriate immune responses against HBV and upregulation of endogenous LPS during hepatitis B, confirming this hypothesis. Due to the aforementioned studies, it appears that TLR4 and its signaling pathways can be considered as important mechanisms to fight HBV. Additionally, according to the aforementioned studies, it seems that TLR4 is downregulated in prolonged forms of hepatitis B. Accordingly, it can be hypothesized that three main mechanisms may be responsible for downregulation of TLR4 in prolonged HBV-infected patients. First, HBV and its related antigens may interfere with TLR4 expression directly. A study by Wu et al. confirms the direct effects of HBsAg on the expression of TLR4. They show that HBsAg is capable of suppressing NF-κB, as a TLR4 downstream signaling molecule, leading to virus escape from immune surveillance (61). Wang et al. revealed that HBsAg could inhibit the JNK-MAPK pathway (55). Lang et al. reported that HBeAg suppressed TIR-mediated activation of NF-κB via disruption of homotypic TIR/TIR interactions (33). Wilson et al. also reported that HBeAg blocked NF-κB activation in hepatocytes (58). Zhang et al. showed that HBcAg downregulated TLR4 expression on the CD14 positive monocytes in HBV-infected patients in comparison to healthy controls (65). Another study identified that HBsAg stimulated monocytes to release IL-10, which resulted in activation of JAK/STAT3 pathway (49) and consequently suppression of IRAK1, IRAK2, TRAF6, and MAPK pathways (Fig. 1). Second, the polymorphisms within the gene of TLR4 may be another reason for downregulation of this molecule in long-term HBV infection. Accordingly, Cussigh et al. reported that the polymorphism within +299 region of TLR4 was significantly associated with chronic HBV infection (19). A study on the Taiwanese population revealed that TLR4 rs4986790 (p.Asp299Gly) polymorphism was significantly associated with HBsAg seroclearance/seroconversion in chronic HBV-infected patients (59). Zhou et al. showed that the polymorphism within 3′-untranslated position of the TLR4 gene was significantly associated with protection from HBV recurrence after liver transplantation in the Chinese population (66). Epigenetic factors are the third plausible candidates for regulation of TLR4 expression in prolonged HBV-infected patients. In parallel with the hypothesis, several investigations demonstrated that miRNAs, as important epigenetic factors, play key roles in TLR4 expression, as well as its molecular signaling (18,36,44). Interestingly, our unpublished data showed that expression of miRNA-1, 21, 125, and 155 were significantly increased in Iranian HBV chronically infected patients. Moreover, previous studies revealed that immune tolerance to HBV antigens is plausible in long-term hepatitis B. Hence, it seems that alteration in TLR4 expression and its intracellular signaling molecules is a probable mechanism to induce immune tolerance to HBV antigens. Interestingly, one study revealed that lipopolysaccharide/TLR4 interactions could lead to activation of T regulatory lymphocytes, which play significant roles in induction of immune tolerance to hepatitis B antigens (65). It appears that additional studies using TLR4 agonists as adjuvants can improve our knowledge about the roles of TLR4 and its related signaling pathways in modulation of immune responses against HBV.

TLR4 and Hepatitis B Liver Complications

In contrast to the roles played by TLR4 in induction of appropriate immune responses against HBV, it appears that there is a different scenario regarding the roles of TLR4 in the pathogenesis of hepatitis B–related complications such as cirrhosis and HCC. For example, Cheng et al. revealed that endotoxin is upregulated during active phases of hepatitis B (16). They also showed that mRNA levels of TLR4 are significantly increased during endotoxin stimulation (16), which leads to production of pro-inflammatory cytokines, including TNF-α, IL-1, and IL-6. The phenomenon results in the aggravation of the hepatitis B. Soares et al. reported that the expression of TLR4 on hepatocytes was increased in hepatitis B and its related complications, including cirrhosis and HCC (51). It has also been identified that hepatic stellate cells (HSCs), the important cells involved in induction of liver cirrhosis, express TLR4 and respond to TLR4 ligands vigorously (3). Wang et al. showed that transfection of HK-2 cells, an immortalized proximal tubule epithelial cell line, with HBx gene results in upregulation of TLR4 in vitro (56). Lian et al. revealed that the expression of TLR4 was significantly upregulated in patients with liver cirrhosis when compared with noncirrhotic chronic HBV-infected patients (34). The main responsible mechanisms that lead to cirrhosis and HCC via TLR4 pathway have yet to be completely clarified, but the authors of the current review article suggest that several directly and indirectly plausible mechanisms could be responsible for induction of cirrhosis and HCC as follows. First, as a probable direct mechanism, it has been evidenced that MYD88-dependent intracellular signaling results in activation of several pro-inflammatory transcription factors, including NF-κB and AP-1, which are the main reasons for induction of HCC (35,38). Based on the fact that TLR4/ligands interactions lead to immune cells activation via MYD88-dependent pathway, it may be hypothesized that TLR4 may promote tumoregenesis pathways in the HBV-infected hepatocytes. Second, as plausible indirect mechanisms, studies have revealed that angiotensin-II is an important molecule that participates in induction of liver fibrosis (48). TLR4 has a pivotal cross-talk with angiotensin-II (50). Hence, it may induce cirrhosis via upregulation of angiotensin-II. It has also been identified that inflammation plays pivotal roles in the induction of cirrhosis and HCC (39,52). TLR4 is a main receptor for PAMPs and DAMPs, which stimulate immune cells to produce inflammation in the infected tissues. Thus, it appears that the induction of inflammation via TLR4/ligands interactions may be considered as another mechanism to induce cirrhosis and HCC. Furthermore, researchers have demonstrated that high-mobility group box 1 (HMGB1) stimulates proliferation, migration, and profibrotic effects of HSCs (54). Wang et al. reported that TLR4/ligands interaction leads to upregulation of HMGB1 (54). Transforming growth factor-β1 (TGF-β1) also plays important roles in the pathogenesis of fibrosis (17). It has been documented that TLR4 promotes the fibrotic effects of TGF-β1 on the fibroblasts (10). It has been documented that T regulator lymphocytes plays significant roles in induction of fibrosis by production of TGF-β1 (17). Lian et al. reported that TLR4 expression has a positive correlation with the frequency of T regulatory lymphocytes in HBV-infected patients with liver cirrhosis (34). Therefore, it appears that TLR4 may participate in the pathogenesis of liver complications, and future therapeutic strategy should be directed to the utilization of TLR4 antagonists for the treatment and prevention of hepatitis B–related liver cirrhosis and HCC.

Concluding Remarks

According to the information presented by the aforementioned investigations, it may be concluded that TLR4 plays important roles in the stimulation of appropriate immune responses against HBV. Thus, the virus targets this receptor and also its intracellular signaling molecules to suppress immune responses in long-term hepatitis B infection. Additionally, based on the presented studies, it seems that TLR4 and its intracellular molecular signaling potentially participate in the pathogenesis of hepatitis B–related complications, including liver cirrhosis and HCC. Accordingly, it appears that future therapies should focus on treatments of noncirrhotic/HCC and cirrhotic/HCC hepatitis B–infected patients with agonists and antagonists of TLR4, respectively, to promote immune responses for eradication of HBV in noncirrhotic/HCC and regulate inflammation and pro-inflammatory transcription factors in cirrhotic/HCC hepatitis B–infected patients.

Footnotes

Acknowledgments

This project was supported by the Rafsanjan University of Medical Sciences.

Author Disclosure Statement

No competing financial interests exist.

References

Ahmadabadi

, Hassanshahi

, Arababadi

, et al. The IL-10 promoter polymorphism at position -592 is correlated with susceptibility to occult HBV infection. Inflammation, 2012; 35:818–821.

Alderson

, McGowan

, Baldridge

, and Probst

. TLR4 agonists as immunomodulatory agents. J Endotoxin Res, 2006; 12:313–319.

Aoyama

, Paik

Y-H

, and Seki

. Toll-like receptor signaling and liver fibrosis. Gastroenterol Res Pract, 2010; 2010. doi:10.1155/2010/192543.

Arababadi

, Ahmadabadi

, and Kennedy

. Current information on the immunological status of occult hepatitis B infection. Transfusion, 2012; 52:1819–1826.

Arababadi

, Pourfathollah

, Jafarzadeh

, et al. Association of exon 9 but not intron 8 VDR polymorphisms with occult HBV infection in south-eastern Iranian patients. J Gastroenterol Hepatol, 2009; 25:90–93.

Arababadi

, Pourfathollah

, Jafarzadeh

, and Hassanshahi

. Serum levels of Interleukin (IL)-10 and IL-17A in occult HBV infected south-east Iranian patients. Hepat Mon, 2010; 10:31–35.

Assar

, Arababadi

, Ahmadabadi

, et al. Occult hepatitis B virus (HBV) infection: a global challenge for medicine. Clin Lab, 2012; 58:1225–1230.

Ayoobi

, Hassanshahi

, Zainodini

, et al. Reduced expression of TRIF in chronic HBV infected Iranian patients. Clin Res Hepatol Gastroenterol, 2013; 37:491–495.

Bae

, Lee

, Choi

, et al. Macrophages generate reactive oxygen species in response to minimally oxidized low-density lipoprotein: toll-like receptor 4- and spleen tyrosine kinase-dependent activation of NADPH oxidase 2. Circ Res, 2009; 104:210–218, 221p following 218.

10.

Bhattacharyya

, Kelley

, Melichian

, et al. Toll-like receptor 4 signaling augments transforming growth factor-β responses: a novel mechanism for maintaining and amplifying fibrosis in scleroderma. Am J Pathol, 2013; 182:192–205.

11.

Cario

, Gerken

, and Podolsky

. Toll-like receptor 2 controls mucosal inflammation by regulating epithelial barrier function. Gastroenterology, 2007; 132:1359–1374.

12.

Chan

, and Jia

. Chronic hepatitis B in Asia—new insights from the past decade. J Gastroenterol Hepatol, 2011; 26 Suppl 1:131–137.

13.

Chang

W-W

, Su

I-J

, Lai

M-D

, et al. Toll-like receptor 4 plays an anti-HBV role in a murine model of acute hepatitis B virus expression. World J Gastroenterol, 2005; 11:6631.

14.

Chang

, Su

, Lai

, et al. The role of inducible nitric oxide synthase in a murine acute hepatitis B virus (HBV) infection model induced by hydrodynamics-based in vivo transfection of HBV-DNA. J Hepatol, 2003; 39:834–842.

15.

Chen

, Cheng

, Xu

, et al. Expression profiles and function of Toll-like receptors 2 and 4 in peripheral blood mononuclear cells of chronic hepatitis B patients. Clin Immunol, 2008; 128:400–408.

16.

Cheng

, and Zhang

. [The significance of CD14 and TLR4 expressions in severe hepatitis B induced by endotoxin]. Zhonghua Gan Zang Bing Za Zhi, 2010; 18:428.

17.

Chizzolini

, Brembilla

, Montanari

, and Truchetet

M-E

. Fibrosis and immune dysregulation in systemic sclerosis. Autoimmun Rev, 2011; 10:276–281.

18.

Curtale

, Mirolo

, Renzi

, et al. Negative regulation of Toll-like receptor 4 signaling by IL-10–dependent microRNA-146b. Proc Nat Acad Sci, 2013; 110:11499–11504.

19.

Cussigh

, Fabris

, Fattovich

, et al. Toll like receptor 4 D299G associates with disease progression in Caucasian patients with chronic HBV infection: relationship with gender. J Clin Immunol, 2013; 33:313–316.

20.

Erridge

. Endogenous ligands of TLR2 and TLR4: agonists or assistants?. J Leukoc Biol., 2010; 87:989–999.

21.

Evans

, Cluff

, Johnson

, et al. Enhancement of antigen-specific immunity via the TLR4 ligands MPL™ adjuvant and Ribi. 529. Expert Rev Vaccines, 2003; 2:219–229.

22.

Fitzgerald

, Rowe

, Barnes

, et al. LPS-TLR4 signaling to IRF-3/7 and NF-κB involves the toll adapters TRAM and TRIF. J Exp Med, 2003; 198:1043–1055.

23.

Heiberg

, Winther

, Paludan

, and Hogh

. Pattern recognition receptor responses in children with chronic hepatitis B virus infection. J Clin Virol, 2012; 54:229–234.

24.

Hirayama

, Tamaki

, Takakubo

, et al. Toll-like receptors and their adaptors are regulated in macrophages after phagocytosis of lipopolysaccharide-coated titanium particles. J Orthop Res, 2011; 29:984–992.

25.

Imani Fooladi

, Mousavi

, Seghatoleslami

, et al. Toll-like receptors: role of inflammation and commensal bacteria. Inflamm Allergy Drug Targets, 2011; 10:198–207.

26.

Isogawa

, Robek

, Furuichi

, and Chisari

. Toll-like receptor signaling inhibits hepatitis B virus replication in vivo. J Virol, 2005; 79:7269–7272.

27.

Jia

, Xie

, Lin

, et al. Common variants of the TLR9 gene influence the clinical course of HBV infection. Mol Med Report, 2009; 2:277–281.

28.

John

, and Crispe

. TLR-4 regulates CD8+ T cell trapping in the liver. J Immunol, 2005; 175:1643–1650.

29.

Kagan

, and Medzhitov

. Phosphoinositide-mediated adaptor recruitment controls Toll-like receptor signaling. Cell, 2006; 125:943–955.

30.

Kawai

, and Akira

. Toll-like receptors and their crosstalk with other innate receptors in infection and immunity. Immunity, 2011; 34:637–650.

31.

Khorramdelazad

, Hassanshahi

, Ahmadabadi

, and Arababadi

. High serum levels of TGF-β in Iranians with chronic HBV infection. Hepat Mon, 2012; 12:e7581.

32.

Khvalevsky

, Rivkin

, Rachmilewitz

, et al. TLR3 signaling in a hepatoma cell line is skewed towards apoptosis. J Cell Biochem, 2007; 100:1301–1312.

33.

Lang

, Lo

, Skinner

, et al. The hepatitis B e antigen (HBeAg) targets and suppresses activation of the toll-like receptor signaling pathway. J Hepatol, 2011; 55:762–769.

34.

Lian

, Wang

, Zhang

, et al. Correlation of circulating TLR2/4 expression with CD3+/4+/8+ T cells and Treg cells in HBV-related liver cirrhosis. Viral Immunol, 2009; 22:301–308.

35.

Liang

, Chen

, Wang

, et al. Myeloid differentiation factor 88 promotes growth and metastasis of human hepatocellular carcinoma. Clin Cancer Res, 2013; 19:2905–2916.

36.

Lippai

, Bala

, Csak

, et al. Chronic alcohol-induced microRNA-155 contributes to neuroinflammation in a TLR4-dependent manner in mice. PloS One, 2013; 8:e70945.

37.

Y-C

, Yeh

W-C

, Ohashi

. LPS/TLR4 signal transduction pathway. Cytokine, 2008; 42:145–151.

38.

Maeda

. NF-κB, JNK, and TLR signaling pathways in hepatocarcinogenesis. Gastroenterol Res Pract, 2010; 2010:367694.

39.

Matsuzaki

, Murata

, Yoshida

, et al. Chronic inflammation associated with hepatitis C virus infection perturbs hepatic transforming growth factor β signaling, promoting cirrhosis and hepatocellular carcinoma. Hepatology, 2007; 46:48–57.

40.

Mendy

, Welzel

, Lesi

, et al. Hepatitis B viral load and risk for liver cirrhosis and hepatocellular carcinoma in The Gambia, West Africa. J Viral Hepat, 2009; 27:27.

41.

Michielsen

, and Ho

. Viral hepatitis B and hepatocellular carcinoma. Acta Gastroenterol Belg, 2011; 74:4–8.

42.

Nguyen-Pham

, Lim

, Nguyen

, et al. Type I and II interferons enhance dendritic cell maturation and migration capacity by regulating CD38 and CD74 that have synergistic effects with TLR agonists. Cell Mol Immunol, 2011; 8:341–347.

43.

Pan

, Gu

, Zhang

, et al. Dynamic changes of lipopolysaccharide levels in different phases of acute on chronic hepatitis B liver failure. PloS One, 2012; 7:e49460.

44.

Philippe

, Alsaleh

, Pichot

, et al. MiR-20a regulates ASK1 expression and TLR4-dependent cytokine release in rheumatoid fibroblast-like synoviocytes. Ann Rheum Dis, 2013; 72:1071–1079.

45.

Qureshi

, Larivière

, Leveque

, et al. Endotoxin-tolerant mice have mutations in Toll-like receptor 4 (Tlr4). J Exp Med, 1999; 189:615–625.

46.

Sajadi

SMA

, Mirzaei

, Hassanshahi

, et al. Decreased expressions of TLR9 and its signaling molecules in chronic HBV infected patients. Arch Path Lab Med., 2013; 137:1674–1679.

47.

Schwabe

, Seki

, and Brenner

. Toll-like receptor signaling in the liver. Gastroenterology, 2006; 130:1886–1900.

48.

Shahid

, Fatima

, and Mahboob

. Angiotensin converting enzyme (ACE) gene expression in experimentally induced liver cirrhosis in rats. Pak J Pharm Sci, 2013; 26:853–857.

49.

Shi

, Ren

, Hu

, et al. HBsAg inhibits IFN-alpha production in plasmacytoid dendritic cells through TNF-alpha and IL-10 induction in monocytes. PLoS One., 2012; 7:e44900.

50.

Shirai

, Yoshiji

, Noguchi

, et al. Cross talk between toll‐like receptor‐4 signaling and angiotensin‐II in liver fibrosis development in the rat model of non‐alcoholic steatohepatitis. J Gastroenterol Hepatol, 2013; 28:723–730.

51.

Soares

J-B

, Pimentel-Nunes

, Afonso

, et al. Increased hepatic expression of TLR2 and TLR4 in the hepatic inflammation-fibrosis-carcinoma sequence. Innate Immun, 2012; 18:700–708.

52.

Tarao

, Ohkawa

, Miyagi

, et al. Inflammation in background cirrhosis evokes malignant progression in HCC development from HCV-associated liver cirrhosis. Scand J Gastroenterol, 2013; 48:729–735.

53.

Visvanathan

, Skinner

, Thompson

, et al. Regulation of toll‐like receptor‐2 expression in chronic hepatitis B by the precore protein. Hepatology, 2007; 45:102–110.

54.

Wang

F-P

, Li

, et al. High mobility group box-1 promotes the proliferation and migration of hepatic stellate cells via TLR4-dependent signal pathways of PI3K/Akt and JNK. PloS One, 2013; 8:e64373.

55.

Wang

, Chen

, Hu

, et al. Hepatitis B virus surface antigen selectively inhibits TLR2 ligand-induced IL-12 production in monocytes/macrophages by interfering with JNK activation. J Immunol, 2013; 190:5142–5151.

56.

Wang

, Zhou

, Zhu

, and Yuan

W-J

. Effects of hepatitis B virus X gene on apoptosis and expression of immune molecules of human proximal tubular epithelial cells. Arch Virol, 2013; 158:2479–2485.

57.

Wei

, Guo

, Liu

, et al. The significance of toll-like receptor 4 (TLR4) expression in patients with chronic hepatitis B. Clin Invest Med, 2008; 31:E123–130.

58.

Wilson

, Warner

, Ryan

, et al. The hepatitis B e antigen suppresses IL-1beta-mediated NF-kappaB activation in hepatocytes. J Viral Hepat, 2011; 18:e499–507.

59.

J-F

, Chen

C-H

, Ni

Y-H

, et al. Toll-like receptor and hepatitis B virus clearance in chronic infected patients: a long-term prospective cohort study in Taiwan. J Infect Dis, 2012; 206:662–668.

60.

, Lu

, Meng

, et al. Toll‐like receptor‐mediated control of HBV replication by nonparenchymal liver cells in mice. Hepatology, 2007; 46:1769–1778.

61.

, Meng

, Jiang

, et al. Hepatitis B virus suppresses toll‐like receptor–mediated innate immune responses in murine parenchymal and nonparenchymal liver cells. Hepatology, 2009; 49:1132–1140.

62.

Xing

, Li

, Cao

, and Huang

. Altered immune function of monocytes in different stages of patients with acute on chronic liver failure. Clin Exp Immunol, 2007; 147:184–188.

63.

Yamamoto

, Sato

, Hemmi

, et al. Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway. Science, 2003; 301:640–643.

64.

Zare-Bidaki

, Hakimi

, Abdollahi

, et al. TLR4 in toxoplasmosis; friends or foe?. Microb Pathog, 2014; 69–70:28–32.

65.

Zhang

, Lian

, Huang

, et al. Overexpression of toll-like receptor 2/4 on monocytes modulates the activities of CD4(+)CD25(+) regulatory T cells in chronic hepatitis B virus infection. Virology, 2010; 397:34–42.

66.

Zhou

, Wei

, Xing

, et al. Polymorphism in 3′‐untranslated region of toll‐like receptor 4 gene is associated with protection from hepatitis B virus recurrence after liver transplantation. Transpl Infect Dis, 2011; 13:250–258.

67.

Zhou

, Zhu

, Wang

, et al. The role of the toll-like receptor TLR4 in hepatitis B virus-associated glomerulonephritis. Arch Virol, 2013; 158:425–433.