Cytosolic retinoic acid-inducible gene I（RIG-I） is an important innate immune RNA sensor and can induce antiviral cytokines, e.g., interferon-β（IFN-β）. Innate immune response to hepatitis B virus（HBV） plays a ...Cytosolic retinoic acid-inducible gene I（RIG-I） is an important innate immune RNA sensor and can induce antiviral cytokines, e.g., interferon-β（IFN-β）. Innate immune response to hepatitis B virus（HBV） plays a pivotal role in viral clearance and persistence. However, knowledge of the role that RIG-I plays in HBV infection is limited. The woodchuck is a valuable model for studying HBV infection. To characterize the molecular basis of woodchuck RIG-I（w RIG-I）, we analyzed the complete coding sequences（CDSs） of w RIG-I, containing 2778 base pairs that encode 925 amino acids. The deduced w RIG-I protein was 106.847 k D with a theoretical isoelectric point（p I） of 6.07, and contained three important functional structures [caspase activation and recruitment domains（CARDs）, DEx D/H-box helicases, and a repressor domain（RD）]. In woodchuck fibroblastoma cell line（WH12/6）, w RIG-I-targeted small interfering RNA（si RNA） down-regulated RIG-I and its downstrean effector–IFN-β transcripts under RIG-I＇ ligand, 5＇-ppp double stranded RNA（ds RNA） stimulation. We also measured m RNA levels of w RIG-I in different tissues from healthy woodchucks and in the livers from woodchuck hepatitis virus（WHV）-infected woodchucks. The basal expression levels of w RIG-I were abundant in the kidney and liver. Importantly, w RIG-I was significantly up-regulated in acutely infected woodchuck livers, suggesting that RIG-I might be involved in WHV infection. These results may characterize RIG-I in the woodchuck model, providing a strong basis for further study on RIG-I-mediated innate immunity in HBV infection.展开更多
Objective： To observe the anti-virus effects of andrographolide （AD） on the retinoic acid-inducible gene-I （RIG-I）-Iike receptors （RLRs） signaling pathway when immunological cells were infected with HIN1. Meth...Objective： To observe the anti-virus effects of andrographolide （AD） on the retinoic acid-inducible gene-I （RIG-I）-Iike receptors （RLRs） signaling pathway when immunological cells were infected with HIN1. Methods： Leukomonocyte was obtained from umbilical cord blood by Ficoll density gradient centrifugation, and immunological cells were harvested after cytokines stimulation. Virus infected cell model was established by H1N1 co-cultured with normal human bronchial epithelial cell line （16HBE）. The optimal concentration of AD was defined by methyl-thiazolyl-tetrazolium （MTT） assay. After the virus infected cell model was established, AD was added into the medium as a treatment intervention. After 24-h co-culture, cell supernatant was collected for interferon gamma （IFN- ~, ） and interleukin-4 （IL-4） enzyme-linked immunosorbent assay （ELISA） detection while immunological cells for real-time polymerase chain reaction （RT-PCR）. Results： The optimal concentration of AD for anti-virus effect was 250 μg/mL. IL-4 and IFN-γ in the supernatant and mRNA levels in RLRs pathway increased when cells was infected by virus, RIG-I, IFN-13 promoter stimulator-1 （IPS-1）, interferon regulatory factor （IRF）-7, IRF-3 and nuclear transcription factor K B （NF- K B） mRNA levels increased significantly （P〈0.05）. When AD was added into co-culture medium, the levels of IL-4 and IFN-γ were lower than those in the non-interference groups and the mRNA expression levels decreased, RIG-I, IPS-1, IRF-7, IRF-3 and NF- K B decreased significantly in each group with significant statistic differences （P〈0.05）. Conclusions： The RLRs mediated viral recognition provided a potential molecular target for acute viral infections and andrographolide could ameliorate H1N1 virus-induced cell mortality. And the antiviral effects might be related to its inhibition of viral-induced activation of the RLRs signaling pathway.展开更多
AIM To investigate the effect of(-)-epigallocatechin-3-gallate(EGCG) on polyinosinic-polycytidylic acid(poly I:C)-triggered intracellular innate immunity against hepatitis C virus(HCV) in hepatocytes. METHODS A cell c...AIM To investigate the effect of(-)-epigallocatechin-3-gallate(EGCG) on polyinosinic-polycytidylic acid(poly I:C)-triggered intracellular innate immunity against hepatitis C virus(HCV) in hepatocytes. METHODS A cell culture model of HCV infection was generated by infecting a hepatoma cell line, Huh7, with HCV JFH-1 strain(JFH-1-Huh7). Poly I:C with a high molecular weight and EGCG were used to stimulate the JFH-1-Huh7 cells. Real-time reverse transcription-polymerase chain reaction was used to detect the expression levels of intracellular m RNAs and of intracellular and extracellular HCV RNA. Enzyme-linked immunosorbent assay was used to evaluate the interferon(IFN)-λ1 protein level in the cell culture supernatant. Immunostaining was used to examine HCV core protein expression in Huh7 cells.RESULTS Our recent study showed that HCV replication could impair poly I:C-triggered intracellular innate immune responses in hepatocytes. In the current study, we showed that EGCG treatment significantly increased the poly I:C-induced expression of Toll-like receptor 3(TLR3), retinoic acid-inducible gene I, and IFN-λ1 in JFH-1-Huh7 cells. In addition, supplementation with EGCG increased the poly I:C-mediated antiviral activity in JFH-1-Huh7 cells at the intracellular and extracellular HCV RNA and protein levels. Further investigation of the mechanisms showed that EGCG treatment significantly enhanced the poly I:C-induced expression of IFN-regulatory factor 9 and several antiviral IFNstimulated genes, including ISG15, ISG56, myxovirus resistance A, and 2'-5'-oligoadenylate synthetase 1, which encode the key antiviral elements in the IFN signaling pathway. CONCLUSION Our observations provide experimental evidence that EGCG has the ability to enhance poly I:C-induced intracellular antiviral innate immunity against HCV replication in hepatocytes.展开更多
Hepatitis C virus(HCV)frequently elicits only mild immune responses so that it can often establish chronic infection.In this case HCV antigens persist and continue to stimulate the immune system.Antigen persistence th...Hepatitis C virus(HCV)frequently elicits only mild immune responses so that it can often establish chronic infection.In this case HCV antigens persist and continue to stimulate the immune system.Antigen persistence then leads to profound changes in the infected host’s immune responsiveness,and eventually contributes to the pathology of chronic hepatitis.This topic highlight summarizes changes associated with chronic hepatitis C concerning innate immunity(interferons,natural killer cells),adaptive immune responses(immunoglobulins,T cells,and mechanisms of immune regulation(regulatory T cells).Our overview clarifies that a strong anti-HCV immune response is frequently associated with acute severe tissue damage.In chronic hepatitis C,however,the effector arms of the immune system either become refractory to activation or take over regulatory functions.Taken together these changes in immunity may lead to persistent liver damage and cirrhosis.Consequently,effector arms of the immune system will not only be considered with respect to antiviral defence but also as pivotal mechanisms of inflammation,necrosis and progression to cirrhosis.Thus,avoiding Scylla-a strong,sustained antiviral immune response with inital tissue damage-takes the infected host to virus-triggered immunopathology,which ultimately leads to cirrhosis and liver cancerthe realm of Charybdis.展开更多
基金supported by grants from the National Science and Technology Major Project for Infectious Diseases of China(No.2012ZX10004503)the National Natural Science Foundation of China(No.81101248,No.81261120397 and No.81371828)the Deutsche Forschungs-gemeinschaft(Transregio TRR60)
文摘Cytosolic retinoic acid-inducible gene I（RIG-I） is an important innate immune RNA sensor and can induce antiviral cytokines, e.g., interferon-β（IFN-β）. Innate immune response to hepatitis B virus（HBV） plays a pivotal role in viral clearance and persistence. However, knowledge of the role that RIG-I plays in HBV infection is limited. The woodchuck is a valuable model for studying HBV infection. To characterize the molecular basis of woodchuck RIG-I（w RIG-I）, we analyzed the complete coding sequences（CDSs） of w RIG-I, containing 2778 base pairs that encode 925 amino acids. The deduced w RIG-I protein was 106.847 k D with a theoretical isoelectric point（p I） of 6.07, and contained three important functional structures [caspase activation and recruitment domains（CARDs）, DEx D/H-box helicases, and a repressor domain（RD）]. In woodchuck fibroblastoma cell line（WH12/6）, w RIG-I-targeted small interfering RNA（si RNA） down-regulated RIG-I and its downstrean effector–IFN-β transcripts under RIG-I＇ ligand, 5＇-ppp double stranded RNA（ds RNA） stimulation. We also measured m RNA levels of w RIG-I in different tissues from healthy woodchucks and in the livers from woodchuck hepatitis virus（WHV）-infected woodchucks. The basal expression levels of w RIG-I were abundant in the kidney and liver. Importantly, w RIG-I was significantly up-regulated in acutely infected woodchuck livers, suggesting that RIG-I might be involved in WHV infection. These results may characterize RIG-I in the woodchuck model, providing a strong basis for further study on RIG-I-mediated innate immunity in HBV infection.
基金Supported by the National Natural Science Foundation of China(No.81273616 and 30973693)the Doctoral Fund of the Ministryof Education 2010(No.20104401110003)the Natural Science Foundation of Guangdong Province(No.S2013010013434)
文摘Objective： To observe the anti-virus effects of andrographolide （AD） on the retinoic acid-inducible gene-I （RIG-I）-Iike receptors （RLRs） signaling pathway when immunological cells were infected with HIN1. Methods： Leukomonocyte was obtained from umbilical cord blood by Ficoll density gradient centrifugation, and immunological cells were harvested after cytokines stimulation. Virus infected cell model was established by H1N1 co-cultured with normal human bronchial epithelial cell line （16HBE）. The optimal concentration of AD was defined by methyl-thiazolyl-tetrazolium （MTT） assay. After the virus infected cell model was established, AD was added into the medium as a treatment intervention. After 24-h co-culture, cell supernatant was collected for interferon gamma （IFN- ~, ） and interleukin-4 （IL-4） enzyme-linked immunosorbent assay （ELISA） detection while immunological cells for real-time polymerase chain reaction （RT-PCR）. Results： The optimal concentration of AD for anti-virus effect was 250 μg/mL. IL-4 and IFN-γ in the supernatant and mRNA levels in RLRs pathway increased when cells was infected by virus, RIG-I, IFN-13 promoter stimulator-1 （IPS-1）, interferon regulatory factor （IRF）-7, IRF-3 and nuclear transcription factor K B （NF- K B） mRNA levels increased significantly （P〈0.05）. When AD was added into co-culture medium, the levels of IL-4 and IFN-γ were lower than those in the non-interference groups and the mRNA expression levels decreased, RIG-I, IPS-1, IRF-7, IRF-3 and NF- K B decreased significantly in each group with significant statistic differences （P〈0.05）. Conclusions： The RLRs mediated viral recognition provided a potential molecular target for acute viral infections and andrographolide could ameliorate H1N1 virus-induced cell mortality. And the antiviral effects might be related to its inhibition of viral-induced activation of the RLRs signaling pathway.
基金Supported by the National Natural Science Foundation of China,No.81500449the Natural Science Foundation of Shanghai,No.14ZR1434200+2 种基金Shanghai Municipal Commission of Health and Family Planning,No.20144Y0175the Scientific Research Foundation for the Returned Overseas Chinese Scholarsthe State Education Ministry of China,No.20150909-6
文摘AIM To investigate the effect of(-)-epigallocatechin-3-gallate(EGCG) on polyinosinic-polycytidylic acid(poly I:C)-triggered intracellular innate immunity against hepatitis C virus(HCV) in hepatocytes. METHODS A cell culture model of HCV infection was generated by infecting a hepatoma cell line, Huh7, with HCV JFH-1 strain(JFH-1-Huh7). Poly I:C with a high molecular weight and EGCG were used to stimulate the JFH-1-Huh7 cells. Real-time reverse transcription-polymerase chain reaction was used to detect the expression levels of intracellular m RNAs and of intracellular and extracellular HCV RNA. Enzyme-linked immunosorbent assay was used to evaluate the interferon(IFN)-λ1 protein level in the cell culture supernatant. Immunostaining was used to examine HCV core protein expression in Huh7 cells.RESULTS Our recent study showed that HCV replication could impair poly I:C-triggered intracellular innate immune responses in hepatocytes. In the current study, we showed that EGCG treatment significantly increased the poly I:C-induced expression of Toll-like receptor 3(TLR3), retinoic acid-inducible gene I, and IFN-λ1 in JFH-1-Huh7 cells. In addition, supplementation with EGCG increased the poly I:C-mediated antiviral activity in JFH-1-Huh7 cells at the intracellular and extracellular HCV RNA and protein levels. Further investigation of the mechanisms showed that EGCG treatment significantly enhanced the poly I:C-induced expression of IFN-regulatory factor 9 and several antiviral IFNstimulated genes, including ISG15, ISG56, myxovirus resistance A, and 2'-5'-oligoadenylate synthetase 1, which encode the key antiviral elements in the IFN signaling pathway. CONCLUSION Our observations provide experimental evidence that EGCG has the ability to enhance poly I:C-induced intracellular antiviral innate immunity against HCV replication in hepatocytes.
文摘Hepatitis C virus(HCV)frequently elicits only mild immune responses so that it can often establish chronic infection.In this case HCV antigens persist and continue to stimulate the immune system.Antigen persistence then leads to profound changes in the infected host’s immune responsiveness,and eventually contributes to the pathology of chronic hepatitis.This topic highlight summarizes changes associated with chronic hepatitis C concerning innate immunity(interferons,natural killer cells),adaptive immune responses(immunoglobulins,T cells,and mechanisms of immune regulation(regulatory T cells).Our overview clarifies that a strong anti-HCV immune response is frequently associated with acute severe tissue damage.In chronic hepatitis C,however,the effector arms of the immune system either become refractory to activation or take over regulatory functions.Taken together these changes in immunity may lead to persistent liver damage and cirrhosis.Consequently,effector arms of the immune system will not only be considered with respect to antiviral defence but also as pivotal mechanisms of inflammation,necrosis and progression to cirrhosis.Thus,avoiding Scylla-a strong,sustained antiviral immune response with inital tissue damage-takes the infected host to virus-triggered immunopathology,which ultimately leads to cirrhosis and liver cancerthe realm of Charybdis.
