Chengliang Zhu

Influenza A Virus Induces Interleukin-27 through Cyclooxygenase-2 and Protein Kinase A Signaling

Background: Interleukin (IL)-27 is significantly elevated in influenza patients. Results: IL-27 inhibits influenza A virus (IAV) replication and its expression is mediated by cyclooxygenase-2, protein kinase A (PKA), and the cAMP-response element-binding protein pathway. Conclusion: IL-27 exerts antiviral function through activation of STAT1/2/3 and PKR phosphorylation. Significance: IL-27 is one host immune factor produced in response to IAV infection. We previously reported that IL-27, which belongs to the IL-12 family of cytokines, is elevated in the serum of patients infected with influenza A virus (IAV). Here, we show that the expression of IL-27 was significantly up-regulated in A549 human lung epithelial cells and human peripheral blood mononuclear cells infected with IAV. Additionally, IAV triggered IL-27 expression through protein kinase A and cAMP-response element-binding protein signaling, which was mediated by cyclooxygenase-2-derived prostaglandin E2. IL-27 inhibited IAV replication by STAT1/2/3 phosphorylation and activated antiviral factor protein kinase R phosphorylation. Clinical analysis showed that IL-27 levels were significantly elevated in a cohort of patients infected with IAV compared with healthy individuals and that circulating IL-27 levels were tightly and positively correlated with prostaglandin E2 levels. These results indicate that IL-27 expression is one host immune factor produced in response to IAV infection and that elevated IL-27 levels inhibit viral replication.

Hepatitis B virus enhances interleukin-27 expression both in vivo and in vitro

The immune system plays important roles in determining the outcomes of hepatitis B virus (HBV) infection. Interleukin-27 (IL-27) is a recently identified pro-inflammatory cytokine produced by macrophages, dendritic cells, and epithelial cells. To determine the correlation between HBV infection and IL-27 expression, we investigated the serum IL-27 levels in patients with HBV infection and in healthy individuals. Results showed that IL-27 was significantly elevated in patients as compared to healthy individuals (P<0.001). IL-27 was also detected at higher levels in patients with liver cirrhosis or hepatocellular carcinoma than those with acute hepatitis B or chronic hepatitis B (P<0.05). We also found that IL-27 expression was influenced by HBV e antigen. In addition, our in vitro studies demonstrated that IL-27 promoter activity, mRNA and protein expression were all stimulated in cells transfected with infectious HBV clone.

Borna Disease Virus P Protein Affects Neural Transmission through Interactions with Gamma-Aminobutyric Acid Receptor-Associated Protein

ABSTRACT Borna disease virus (BDV) is one of the infectious agents that causes diseases of the central nervous system in a wide range of vertebrate species and, perhaps, in humans. The phosphoprotein (P) of BDV, an essential cofactor of virus RNA-dependent RNA polymerase, is required for virus replication. In this study, we identified the gamma-aminobutyric acid receptor-associated protein (GABARAP) with functions in neurobiology as one of the viral P protein-interacting cellular factors by using an approach of phage display-based protein-protein interaction analysis. Direct binding between GABARAP and P protein was confirmed by coimmunoprecipitation, protein pull-down, and mammalian two-hybrid analyses. GABARAP originally was identified as a linker between the gamma-aminobutyric acid receptor (GABAR) and the microtubule to regulate receptor trafficking and plays important roles in the regulation of the inhibitory neural transmitter gamma-aminobutyric acid (GABA). We showed that GABARAP colocalizes with P protein in the cells infected with BDV or transfected with the P gene, which resulted in shifting the localization of GABARAP from the cytosol to the nucleus. We further demonstrated that P protein blocks the trafficking of GABAR, a principal GABA-gated ion channel that plays important roles in neural transmission, to the surface of cells infected with BDV or transfected with the P gene. We proposed that during BDV infection, P protein binds to GABARAP, shifts the distribution of GABARAP from the cytoplasm to the nucleus, and disrupts the trafficking of GABARs to the cell membranes, which may result in the inhibition of GABA-induced currents and in the enhancement of hyperactivity and anxiety.

Inducible Interleukin 32 (IL-32) Exerts Extensive Antiviral Function via Selective Stimulation of Interferon λ1 (IFN-λ1)

Background: Interleukin-32 participates in responses to viral infection, but how virus induces its expression and the mechanisms of its antiviral activities remain unclear. Results: Interleukin-32 selectively stimulates IFN-λ1 via activating distinct regulatory elements in the promoter. Conclusion: Interleukin-32 promotes IFN-λ1-mediated antiviral response. Significance: Interleukin-32 is one immune response factor and plays an important role in the control of viral infection. Interleukin (IL)-32 has been recognized as a proinflammatory cytokine that participates in responses to viral infection. However, little is known about how IL-32 is induced in response to viral infection and the mechanisms of IL-32-mediated antiviral activities. We discovered that IL-32 is elevated by hepatitis B virus (HBV) infection both in vitro and in vivo and that HBV induced IL-32 expression at the level of both transcription and post-transcription. Furthermore, microRNA-29b was found to be a key factor in HBV-regulated IL-32 expression by directly targeting the mRNA 3′-untranslated region of IL-32. Antiviral analysis showed that IL-32 was not sufficient to alter HBV replication in HepG2.2.15 cells. To mimic the viremic phase of viral infection, freshly isolated peripheral blood mononuclear cells were treated with IL-32γ, the secretory isoform, and the supernatants were used for antiviral assays. Surprisingly, these supernatants exhibited extensive antiviral activity against multiplex viruses besides HBV. Thus, we speculated that the IL-32γ-treated peripheral blood mononuclear cells produced and secreted an unknown antiviral factor. Using antibody neutralization assays, we identified the factor as interferon (IFN)-λ1 and not IFN-α. Further studies indicated that IL-32γ effectively inhibited HBV replication in a hydrodynamic injection mouse model. Clinical data showed that elevated levels of IFN-λ1 both in serum and liver tissue of HBV patients were positively correlated to the increased levels of IL-32. Our results demonstrate that elevated IL-32 levels during viral infection mediate antiviral effects by stimulating the expression of IFN-λ1.

Hepatitis C virus NS3 protein enhances cancer cell invasion by activating matrix metalloproteinase-9 and cyclooxygenase-2 through ERK/p38/NF-κB signal cascade

Hepatitis C virus (HCV) infection causes acute and chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma (HCC). However, the mechanisms by which HCV causes the diseases are largely unknown. Here, we elucidated the effects of HCV on the invasion and migration of hepatoma cells, with the aim to reveal the mechanism by which HCV infection induces HCC. We initially showed that matrix metalloproteinase-9 (MMP-9) was elevated in the sera of HCV-infected patients, and demonstrated that HCV nonstructural protein 3 (NS3) activated MMP-9 transcription through nuclear factor-κB (NF-κB) by stimulating translocation of NF-κB from cytosol to the nucleus to enhance its binding to MMP-9 promoter. In addition, cyclooxygenase-2 (COX-2) and extracellular signal-regulated kinase (ERK1/2)/mitogen-activated protein kinase (p38) pathway were involved in HCV-activated MMP-9 expression. Moreover, NS3 enhanced hepatoma cell invasion and migration through MMP-9 and COX-2. Thus, this study provides new insights into the roles of HCV NS3, MMP-9 and COX-2 in regulating cancer cell invasion.

IL-27, a Cytokine, and IFN-λ1, a Type III IFN, Are Coordinated To Regulate Virus Replication through Type I IFN

IL-27, a member of the IL-12 family, plays a critical role in the control of innate and adaptive immune responses. IFN-λ1, a member of the type III IFN family, shows antiviral abilities. In this study, we investigated the effects of IL-27 and IFN-λ1 on the replication of hepatitis B virus (HBV), a major pathogen associated with a high risk for cirrhosis, liver failure, and hepatocellular carcinoma. We revealed that HBV infection activates IL-27 expression and IFN-λ1 production and demonstrated that viral-activated IL-27 and IFN-λ1 are coordinated to inhibit HBV replication. Initially, HBV infection upregulates IL-27 expression, which, in turn, stimulates IFN-λ1 production through regulating ERK1/2 signaling and by enhancing NF-κB nuclear translocation to bind to the IFN-λ1 promoter. Moreover, IL-27–activated IFN-λ1 upregulates IFN-λ1 receptor (IL-28R1 and IL-10Rβ) activity, resulting in the activation of the STAT1/2 pathway, which, in turn, induces the expression of IFN-stimulated genes, including IFN-inducible dsRNA-activated protein kinase, oligoadenylate synthetase 1, and IFN-induced GTP-binding protein 1 and, finally, inhibits HBV protein expression and viral capsid–associated DNA replication. More interestingly, we also revealed that type I IFN (IFN-α) is also involved in the downregulation of HBV replication mediated by IL-27. Thus, we identified a previously unknown mechanism by which IL-27 and IFN-λ1 are coordinated to regulate virus replication through type I IFN.

Association of functional polymorphism of ApoB promoter with hepatitis C virus infection

BACKGROUND Hepatitis C virus (HCV) binds to apolipoprotein B (ApoB) and low-density lipoprotein (LDL) before entering into hepatocytes. ApoB promoter polymorphisms influence the levels of ApoB and LDL in blood. We investigated the correlations between ApoB promoter polymorphism and HCV infection. METHODS ApoB promoter polymorphism was analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 243 HCV-infected patients and 346 healthy individuals. Serum lipids concentrations were measured in all subjects. RESULTS ApoB polymorphisms on -516 position in patients were statistically different from healthy individuals (p<0.0001). CC genotype was higher in patients (58.7%) than in healthy individuals (43.1%) (p<0.0001). TT genotype was lower in patients (6.2%) than in healthy group (16.1%) (p<0.0001). C allele frequency was higher (76.3%) in patients than in healthy individuals (63.4%) (p<0.0001). T allele was lower in patients (23.7%) than in controls (36.6%) (p<0.0001). CC genotype was higher in HCV RNA positive patients (63.5%) than in negative ones (45.9%) (p<0.0001). TT genotype was significantly higher in HCV RNA negative patients (19.2%) than in positive individuals (2.6%) (p<0.0001). CONCLUSIONS CC genotype of ApoB promoter at -516 position may increase susceptibility of HCV infection and TT genotype may be associated with viral clearance.

Hepatitis C virus core protein induces hypoxia-inducible factor 1α-mediated vascular endothelial growth factor expression in Huh7.5.1 cells.

Hepatitis C virus (HCV) infection is one of the major causes of hepatocellular carcinoma (HCC). It has been demonstrated that the overexpression of angiogenic factors are associated with the maintenance of liver neoplasia. Hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) are important regulators of angiogenesis and are important in wound healing, the regeneration of new vessels and reproductive functions. The present study investigated the role of the HCV core protein in the induction of HIF-1α and VEGF expression. The HCV core gene and HIF-1α siRNA were transfected into Huh7.5.1 cells. The results demonstrated that the induction of HCV core gene expression in Huh7.5.1 cells leads to the overexpression and stabilization of HIF-1α, and the activation of HIF-1α leads, in turn, to the stimulation of VEGF, which is one of the most important angiogenic factors. These results provide new information to facilitate the understanding of HCC oncogenesis.

Matrix Metalloproteinase 9 Facilitates Hepatitis B Virus Replication through Binding with Type I Interferon (IFN) Receptor 1 To Repress IFN/JAK/STAT Signaling

ABSTRACT Hepatitis B virus (HBV) infection may cause acute hepatitis B, chronic hepatitis B (CHB), liver cirrhosis, and hepatocellular carcinoma (HCC). However, the mechanisms by which HBV evades host immunity and maintains chronic infection are largely unknown. Here, we revealed that matrix metalloproteinase 9 (MMP-9) is activated in peripheral blood mononuclear cells (PBMCs) of HBV-infected patients, and HBV stimulates MMP-9 expression in macrophages and PBMCs isolated from healthy individuals. MMP-9 plays important roles in the breakdown of the extracellular matrix and in the facilitation of tumor progression, invasion, metastasis, and angiogenesis. MMP-9 also regulates respiratory syncytial virus (RSV) replication, but the mechanism underlying such regulation is unknown. We further demonstrated that MMP-9 facilitates HBV replication by repressing the interferon (IFN)/Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway, IFN action, STAT1/2 phosphorylation, and IFN-stimulated gene (ISG) expression. Moreover, MMP-9 binds to type I IFN receptor 1 (IFNAR1) and facilitates IFNAR1 phosphorylation, ubiquitination, subcellular distribution, and degradation to interfere with the binding of IFANR1 to IFN-α. Thus, we identified a novel positive-feedback regulation loop between HBV replication and MMP-9 production. On one hand, HBV activates MMP-9 in infected patients and leukocytes. On the other hand, MMP-9 facilitates HBV replication through repressing IFN/JAK/STAT signaling, IFNAR1 function, and IFN-α action. Therefore, HBV may take the advantage of MMP-9 function to establish or maintain chronic infection. IMPORTANCE Hepatitis B virus (HBV) infection may cause chronic hepatitis B (CHB) and hepatocellular carcinoma (HCC). However, the mechanisms by which HBV maintains chronic infection are largely unknown. Matrix metalloproteinase 9 (MMP-9) plays important roles in the facilitation of tumor progression, invasion, metastasis, and angiogenesis. However, the effects of MMP-9 on HBV replication and pathogenesis are not known. This study reveals that MMP-9 expression is activated in patients with CHB, and HBV stimulates MMP-9 production in PBMCs and macrophages. More interestingly, MMP-9 in turn promotes HBV replication through suppressing IFN-α action. Moreover, MMP-9 interacts with type I interferon receptor 1 (IFNAR1) to disturb the binding of IFN-α to IFNAR1 and facilitate the phosphorylation, ubiquitination, subcellular distribution, and degradation of IFNAR1. Therefore, these results discover a novel role of MMP-9 in viral replication and reveal a new mechanism by which HBV evades host immunity to maintain persistent infection.

The collagen triple helix repeat containing 1 facilitates hepatitis B virus‐associated hepatocellular carcinoma progression by regulating multiple cellular factors and signal cascades

Hepatitis B virus (HBV) infection is one of the major causes of acute and chronic liver diseases, fulminant hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). HCC accounts for more than 85% of primary liver cancers and is the seventh most common cancer and the third leading cause of cancer‐related deaths. However, the mechanism by which HBV induces HCC is largely unknown. Collagen triple helixes repeat containing 1 (CTHRC1) is a secreted protein and has characteristics of a circulating hormone with potentially broad implications for cell metabolism and physiology. CTHRC1 is associated with human cancers, but its effect on HCC is unknown. Here, we revealed that CTHRC1 expression is highly correlated with HCC progression in HBV‐infected patients, and demonstrated that HBV stimulates CTHRC1 expression by activating nuclear factor‐kappa B (NF‐κB) and cAMP response element binding protein (CREB), through extracellular signal‐regulated kinase/c‐Jun N‐terminal kinase (ERK/c‐JNK) pathway. In addition, CTHRC1 activates hypoxia‐inducible factor 1α (HIF‐1α) and vascular endothelial growth factor (VEGF) through regulating phosphoinosmde‐3‐kinase/protein kinase B/mammalian target of rapamycin (PI‐3K/AKT/mTOR) pathway. More interestingly, CTHRC1 enhances colony formation, migration, and invasion of hepatoma cells by regulating p53 and stimulating matrix metalloproteinase‐9 (MMP‐9) expression. In addition, knock‐down of CTHRC1 results in the repression of HBV‐associated carcinogenesis in nude mice. Thus, we revealed a novel mechanism by which HBV facilitates HCC development through activating the oncoprotein CTHRC1, which in turn enhances HBV‐related HCC progression by stimulates colony formation, migration, and invasion of hepatoma cells through regulating multiple cellular factors and signal cascades.