Development and evaluation of an inactivated bivalent vaccine against duck viral hepatitis / 生物工程学报

The rapid mutation and widely spread of duck hepatitis A virus (DHAV) lead to the vast economic loss of the duck industry. To prepare and evaluate bivalent inactivated vaccine laboratory products of DHAV, 6 strains were screened from 201 DHAV-1 strains and 38 DHAV-3 strains by using serotype epidemiological analysis in most of the duck factory. Vaccine candidate strains were selected by ELD50 and LD50 tests in the 6 strains. Continuously passaged, the 5th passaged duck embryos bodies grinding fluid was selected as vaccine virus seeds. The virus seeds were treated with formaldehyde and water in oil in water (W/O/W) emulsions, making into three batches of two bivalent inactivated vaccine laboratory products. The safety test, antibody neutralization test, challenged protection and cross immune protection experiment suggested that the vaccines possessed good safety, and neutralizing antibodies were detected at 7th day and the challenged protection rate reached 90% to 100% at the 14th and 21st day. Moreover, immune duration of ducklings lasted more than five weeks. However, cross-immunity protection experiments with DHAV-SH and DHAV-FS only had 20%-30%. The two bivalent inactivated vaccine laboratory products of duck viral hepatitis were effective and reliable, providing a new method as well as a new product for DHAV prevention and control.

Fowl adenovirus Group I as a causal agent of inclusion body hepatitis/hydropericardium syndrome (IBH/HPS) outbreak in brazilian broiler flocks / Adenovirus aviário Ggrupo I como um agente causal da síndrome da hepatite por corpúsculo de inclusão/hidropericárdio (IBH/HPS) em lotes de frangos de corte

Commercial broiler flocks from a farm located in the State of São Paulo, Brazil, presented diarrhea, depression, increased mortality and poor weight gain. Upon post-mortem examination, classical signs of Inclusion Body Hepatitis/Hydropericardium Syndrome (IBH/HPS) were observed, including enlarged pale yellow-colored livers and straw-colored liquid in the pericardial sac. In addition, gross lesions were also observed in the kidneys, pancreas, thymus, intestines and gallbladder. Samples of these organs were analyzed by PCR for the detection of the hexon gene of the Fowl Adenovirus (FAdVs) Group I. The results were positive for both flocks (A and B) assayed by PCR. The macroscopic lesions associated with the detection of FAdV Group I by PCR in several of these affected organs allowed for the identification of IBH/HPS. In fact, this is the first report in Brazil of IBH/HPS in broilers, which identifies FAdVs group I as a causal agent of the disease. These findings may contribute to the worldwide epidemiology of the adenovirus-mediated hepatitis/hydropericardium syndrome...

Lotes comerciais de frangos de uma granja localizada no Estado de São Paulo, Brasil, apresentavam diarreia, depressão, aumento de mortalidade e baixo ganho de peso. Após o exame post-mortem, sinais clássicos da síndrome de hepatite por corpúsculo de inclusão/hidropericárdio (IBH/HPS) foram observados incluindo hepatomegalia com aspecto amarelado pálido e líquido de coloração amarelo palha no saco pericárdio. Além disso, as alterações macroscópicas foram também observadas nos rins, pâncreas, timo, intestinos e vesícula biliar. Amostras destes órgãos foram analisadas pela técnica de PCR para detectar o adenovírus aviário do grupo I através do gene Hexon. Os resultados foram positivos para ambos os lotes (A e B) utilizando-se a técnica de PCR. As lesões macroscópicas associadas à detecção do adenovírus aviário do grupo I pela técnica de PCR em vários destes órgãos acometidos permitiu a identificação da síndrome de hepatite/hidropericárdio em frangos no Brasil. Ao nosso conhecimento, este é a primeira descrição da síndrome de hepatite/hidropericárdio causado por adenovírus aviário do grupo I, no Brasil. Estes achados podem contribuir com a epidemiologia mundial do adenovírus mediando a síndrome de hepatite/hidropericárdio...

Development and validation of a genotype 3 recombinant protein-based immunoassay for hepatitis E virus serology in swine

Hepatitis E virus (HEV) is classified within the family Hepeviridae, genus Hepevirus. HEV genotype 3 (Gt3) infections are endemic in pigs in Western Europe and in North and South America and cause zoonotic infections in humans. Several serological assays to detect HEV antibodies in pigs have been developed, at first mainly based on HEV genotype 1 (Gt1) antigens. To develop a sensitive HEV Gt3 ELISA, a recombinant baculovirus expression product of HEV Gt3 open reading frame-2 was produced and coated onto polystyrene ELISA plates. After incubation of porcine sera, bound HEV antibodies were detected with anti-porcine anti-IgG and anti-IgM conjugates. For primary estimation of sensitivity and specificity of the assay, sets of sera were used from pigs experimentally infected with HEV Gt3. For further validation of the assay and to set the cutoff value, a batch of 1100 pig sera was used. All pig sera were tested using the developed HEV Gt3 assay and two other serologic assays based on HEV Gt1 antigens. Since there is no gold standard available for HEV antibody testing, further validation and a definite setting of the cutoff of the developed HEV Gt3 assay were performed using a statistical approach based on Bayes' theorem. The developed and validated HEV antibody assay showed effective detection of HEV-specific antibodies. This assay can contribute to an improved detection of HEV antibodies and enable more reliable estimates of the prevalence of HEV Gt3 in swine in different regions.

SOCS3 expression correlates with severity of inflammation in mouse hepatitis virus strain 3-induced acute liver failure and HBV-ACLF / 华中科技大学学报（医学）（英德文版）

Recently, suppressor of cytokine signaling-3 (SOCS3) has been shown to be an inducible endogenous negative regulator of Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway which is relevant in inflammatory response, while its functions in acute liver failure and HBV-induced acute-on-chronic liver failure (HBV-ACLF) have not been fully elucidated. In this study, we explored the role of SOCS3 in the development of mouse hepatitis virus strain 3 (MHV-3)-induced acute liver failure and its expression in liver and peripheral blood mononuclear cells (PBMCs) of patients with HBV-ACLF. Inflammation-related gene expression was detected by real-time PCR, immunohistochemistry and Western blotting. The correlation between SOCS3 level and liver injury was studied. Our results showed that the SOCS3 expression was significantly elevated in both the liver tissue and PBMCs from patients with HBV-ACLF compared to mild chronic hepatitis B (CHB). Moreover, a time course study showed that SOCS3 level was increased remarkably in the liver of BALB/cJ mice at 72 h post-infection. Pro-inflammatory cytokines, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α, were also increased significantly at 72 h post-infection. There was a close correlation between hepatic SOCS3 level and IL-6, and the severity of liver injury defined by alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, respectively. These data suggested that SOCS3 may play a pivotal role in the pathogenesis of MHV-3-induced acute liver failure and HBV-ACLF.

CD69+NK cells contribute to the murine hepatitis virus strain 3-induced murine hepatitis / 华中科技大学学报（医学）（英德文版）

The role of hepatic CD69+ natural killer (NK) cells in virus-induced severe liver injury and subsequent hepatic failure is not well defined. In this study, a mouse model of fulminant liver failure (FHF) induced by murine hepatitis virus strain 3 (MHV-3) was used to study the role of hepatic CD69+NK cells in the development of FHF. The CD69 expression in NK cells in the liver, spleen, bone marrow and peripheral blood was detected by using flow cytometry. The correlation between the CD69 level in hepatic NK cells and liver injury was studied. The functional marker (CD107a), and activating and inhibitory receptor (NKG2D and NKG2A) expressed on CD69+NK cells and CD69-NK cells were detected by using flow cytometry. Pro-inflammatory cytokines (IL-9, IFN-γ and TNF-α) were also examined by using intracellular staining. After MHV-3 infection, the number of CD69+NK cells in the liver of BALB/cJ mice was increased markedly and peaked at 72 h post-infection. Similar changes were also observed in the spleen, bone marrow and peripheral blood. Meanwhile, the CD69 expression in hepatic NK cells was highly correlated with the serum level of ALT and AST. The expression of CD107a and NKG2D, as well as the production of TNF-α, IFN-γ and IL-9 in hepatic CD69+NK cells was all significantly up-regulated during 48-72 h post-infection. In contrast, the NKG2A expression was increased in hepatic CD69-NK cells but not in CD69+NK cells. These results suggested that hepatic CD69+NK cells play a pivotal role in the pathogenesis of FHF by enhancing degranulation and cytotoxic ability of NK cells and increasing the production of pro-inflammatory cytokines.

Sequence analysis of VP1 gene of the duck hepatitis A virus type 3 strains isolated from Shandong Province of China in 2012 / 病毒学报

To reveal the genetic variation of the viral protein 1 (VP1) gene of the duck hepatitis A virus type 3 (DHAV-3), the VP1 gene of 13 virulent DHAV-3 strains isolated from Shandong province of China in 2012 were amplified by RT-PCR, sequenced and analyzed. The results showed that all the VP1 genes of the 13 isolates contained 720 nucleotides encoding 240 amino acids, and shared with nucleotide identities of 94. 6%-99.9% and amino acid identities of 95.0%-100%. The nucleotide and amino acid sequence homologies between the 13 DHAV-3 isolates and other 31 DHAV-3 reference strains were 92.5%-100% and 90. 8%-100%, respectively. Phylogenetic analysis showed that the VP1 gene of DHAV-3 had distinct geographical characteristics. Distribution of genotypes of the 44 DHAV-3 strains was as follows: except the vaccine strain B63, all the other Chinese isolates belonged to genotype I (GI), Vietnamese wild isolates mainly belonged to subtype 1 (S1) of genotype II (GII), and all Korean isolates belonged to subtype 2 (S2) of GII.

Research on the gene structure of duck hepatitis B virus and its encoding proteins / 病毒学报

Duck hepatitis B virus (DHBV) belongs to the Avihepadnavirus genus of the Hepadnaviridae, and it not only has the same replication pattern, but also has the similar genomic and antigenic structures to Hepatitis B virus (HBV). The genome of DHBV is a partially double-stranded closed circular DNA. The genome consists of three distinct open reading frames (ORFs): ORF-PreS/S, ORF-PreC/C and ORF-P, which all locate on the negative DNA strand and encode four separate proteins. The ORF-PreS/S encodes envelope proteins L and S, and the ORF-PreC/C and ORF-P encode capsid proteins C and polymerase proteins P, respectively. The characteristics of genome structure,viral proteins features and functions were described in this review in order to provide useful information for the further study of DHBV and the duck model infected by DHBV.

Overview on duck virus hepatitis A / 生物工程学报

This article describes the nomenclature, history and genetic evolution of duck hepatitis A virus, and updates the epidemiology, clinical symptom and surveillances of duck virus hepatitis A. It also summarizes the present status and progress of duck virus hepatitis A and illustrated the necessity and urgency of its research, which provides rationale for the control of duck hepatitis A virus disease in China.

Contribution of IL-17 to mouse hepatitis virus strain 3-induced acute liver failure / 华中科技大学学报（医学）（英德文版）

Recently, the Th17 cells and IL-17 have been shown to play a critical role in the immune-mediated liver injury in hepatitis B, while their functions in acute liver failure have not been well elucidated yet. In this study, we primarily investigated the role of IL-17 in the development of mouse hepatitis virus strain 3 (MHV-3)-induced acute liver failure. IL-17 mRNA levels in liver tissue were quantified by using quantitative real-time polymerase chain reaction, and cytokine IL-17 levels in liver tissue and serum were determined by using ELISA in MHV-3-induced murine fulminant hepatitis model. The IL-17 expression levels on CD4(+)T and CD8(+)T cells were determined by using flow cytometry. The correlation between IL-17 level and liver injury was studied. Th17 associated cytokines were also investigated by intracellular staining. Our results showed that the IL-17 expression was significantly elevated in the liver and serum of BALB/cJ mice infected with MHV-3. Moreover, a time course study showed that the percentage of both IL-17-producing CD4(+)T cells and IL-17-producing CD8(+)T cells was increased remarkably in the liver starting from 48 h and peaked at 72 h post-infection. There was a close correlation between hepatic or serum IL-17 concentration and the severity of liver injury defined by ALT level, respectively. Th17 associated cytokines, IL-6, IL-21 and IL-22, were also increased significantly at 72 h post-infection. It was concluded that IL-17 may contribute to the pathogenesis of MHV-3-induced acute liver failure.

Inhibitory effect of total saponins isolated from Taraphochlamys affinis on duck hepatitis B virus replication / 中国中药杂志

It has been previously shown that Taraphochlamys affinis possessed anti-hepatitis B virus (HBV) activities. To identify the active ingredients, the total saponins (TSTA) were isolated from T. affinis and the inhibitory effect of TSTA on HBV in the duck HBV model was examined. The results showed that serum levels of DHBV-DNA decreased in all ducks treated with TSTA (1.0 and 2.0 g x kg(-1) x d(-1)) and lamivudine (3TC) (50 mg x kg(-1) x d(-1)) during treatment, but 7 days after the cessation of treatment (p7) with 3TC, the viral replication level returned to the pretreatment baseline. Contrariwise in ducks treated with TSTA, the effect of DHBV DNA inhibition lasted. Compared with model control group,the alanine aminotransferase (ALT), aspartate aminotransferase (AST) and duck hepatitis B surface antigen (DHBsAg) values of 1.0 and 2.0 g x kg(-1) x d(-1)-dose TSTA groups were significantly lower on 7, 14 days after the treatment (d7, d14) and p7, and at p7, the ALT and DHBsAg levels of 2.0 g x kg(-1) x d(-1)-dose TSTA group was significantly lower than that of 3TC group. Furthermore, significant histological improvement was noted in ducklings of TSTA treatment group 7 days after the withdrawal. The study results demonstrate that TSTA possesses potent anti-HBV activity.

KCTD9, a novel potassium channel related gene, was highly expressed in hepatic NK cells and T cells of fulminant hepatitis mice induced by MHV-3 / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To explore the mechanisms of a novel potassium channel gene named KCTD9 (potassium channel tetramerization domain containing 9) in model of fulminant viral hepatitis induced by murine hepatitis virus 3 (MHV-3).</p><p><b>METHODS</b>78 BALB/cJ mice(6 male) were randomly and equally assigned to two groups, model group of fulminant viral hepatitis induced by MHV3 and its control. 75 C3H/HeJ female mice were done into two groups, 39 for model group of chronic hepatitis induced by MHV3, 36 for control. Various samples including spleen, liver and lymphocytes from mice of two model groups and the controls were examined for KCTD9 expression by real time quantitative PCR and Immunohistochemistry. Independent-samples T test or one-way ANOVA were carried out in different groups.</p><p><b>RESULTS</b>Increased expressions of KCTD9 mRNA was observed in livers of both model mice of fulminant viral hepatitis and chronic hepatitis. Compared with the control mice, the expressions of KCTD9 mRNA were up-regulated by 577.1-, 8.8-, 59.4- and 10.8-fold in hepatic NK cells, CD4+ T cells, CD8+ T cells and splenic NK cells respectively in model mice of fulminant viral hepatitis 48 hr post MHV-3 infection, whereas down-regulation by 43% and 69% in splenic CD4 + T cells and CD8+ T cells were found respectively. In contrast, in model mice of chronic viral hepatitis the expressions of KCTD9 mRNA were down-regulated by 71% and 51% in hepatic CD4+ T cells and NK cells, respectively. The expression of KCTD9 protein was mainly evidenced in infiltrative mononuclear cells of liver as shown by immunohistochemistry. Basal expression was also investigated and showed constitutive expression of KCTD9 in brain, thymus and other organs in BALB/cJ mice.</p><p><b>CONCLUSION</b>A novel potassium channel gene KCTD9 was highly expressed in hepatic NK cells and T cells of fulminant hepatitis mice induced by MHV-3.</p>

review of hepatitis viral infections in Pakistan

A review of published literature on viral hepatitis infections in Pakistan is presented. A total of 220 abstracts available in the Pakmedinet and Medline have been searched. All relevant articles were reviewed to determine the prevalence of hepatitis viral infections in Pakistan. Two hundred and three [203] relevant articles/abstracts including twenty nine supporting references are included in this review. Of the articles on prevalence of hepatitis infection, seven were related to Hepatitis A, fifteen to Hepatitis E while the remaining articles were on frequency of hepatitis B and C in different disease and healthy population groups. These included eight studies on healthy children, three on vertical transmission, nineteen on pregnant women, fifteen on healthy individuals, six on army recruits, thirty one on blood donors, thirteen on health care workers, five on unsafe injections, seventeen on high risk groups, five on patients with provisional diagnosis of hepatitis, thirty three on patients with chronic liver disease, four on genotypes of HBV and five on genotypes of HCV. This review highlights the lack of community-based epidemiological work as the number of subjects studied were predominantly patients, high risk groups and healthy blood donors. High level of Hepatitis A seroconversion was found in children and this viral infection accounts for almost 50%-60% of all cases of acute viral hepatitis in children in Pakistan. Hepatitis E is endemic in the country affecting mostly the adult population and epidemic situations have been reported from many parts of the country. The mean results of HBsAg and Anti-HCV prevalence on the basis of data aggregated from several studies was calculated which shows 2.3% and 2.5% prevalence of HBsAg and Anti-HCV in children, 2.5% and 5.2% among pregnant women, 2.6% and 5.3% in general population, 3.5% and 3.1% in army recruits, 2.4% and 3.6% in blood donors, 6.0% and 5.4% in health care workers, 13.0% and 10.3% in high risk groups, 12.3% and 12.0% in patients with provisional diagnosis of hepatitis and 25.7% and 54% in patients with chronic liver disease respectively. This review has illustrated the high endemicity of hepatitis viral infections in Pakistan where hepatitis B and C potentially account for a serious burden of the disease. This review has triggered the launching of a network intervention for the control of hepatitis viral infectious. This review was used as the basis for the launch of hepatitis programme, but putting it into a formal review took time and the hepatitis program was initiated

Construction of MHV-A59 damp-heat mouse model and analysis of the relevant indices / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the impact of inflammation, water metabolism and immune function on the establishment of a mouse model of damp-heat syndrome with MHV-A59 infection.</p><p><b>METHODS</b>Twenty-four mice were randomly divided into control group, virus group, damp-heat group and model group. The peripheral blood CD4(+) and CD8(+) lymphocytes were detected by flow cytometry, and the serum levels of IFN-γ and IL-4 were assayed by ELISA. The expressions of NF-κB and AQP4 in the liver and stomach were determined using immunohistochemistry.</p><p><b>RESULTS</b>The expression of NF-κB and CD4(+)/CD8(+) ratio in the virus and model groups were significantly higher than those in the damp-heat and control groups, while the expression of AQP4 was significantly higher in the model and damp-heat groups than in the other groups. Compared with the control group, the model group showed a significantly higher ratio of IFN-γ/IL-4.</p><p><b>CONCLUSIONS</b>MHV-A59 virus is the main cause of elevated NF-κB expression and CD4(+)/CD8(+)/ ratio, while damp-heat syndrome is responsible for increased AQP4 expression, and their synergistic effect results in increased IFN-γ/IL-4 ratio. The mouse model established using MHV-A59 virus and the damp-heat factors can mimic damp-heat syndrome described in traditional Chinese medicine theory.</p>

Effects of hyperin on the cccDNA of duck hepatitis B virus and its immunological regulation / 药学学报

The aim of this study is to investigate the effect of hyperin on the cccDNA of duck hepatitis B virus and its immunological regulation. Duck hepatitis B virus (DHBV) infection model and normal mouse spleen lymphocyte were used to evaluate the anti-HBV and immunoregulation effects. The DHBV-DNA of serum was detected at different time points by using serum DOT-BLOT hybridization. Polymerase chain reaction (PCR) was used for the determination of nuclear covalent closed circular DNA (cccDNA). Cytokine secretion was determined by ELISA method. DHBV-DNA were inhibited by hyperin (25 or 50 mg x kg(-1)), while cccDNA of liver could be eliminated efficiently by hyperin (25 or 50 mg x kg(-1), P < 0.05, P < 0.01). The T helper 1 effector cytokine was markedly enhanced by hyperin (25 or 50 microg x mL(-1), P < 0.01). In conclusion, hyperin has anti-HBV activity via multiple targets and pathways, and cccDNA may be one of the important targets.

Role of CD4-CD8- T cells in the murine hepatitis virus type 3 induced chronic viral hepatitis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate role of CD4-CD8- T cells in murine hepatitis virus type 3 (MHV-3) induced chronic viral hepatitis in C3H/Hej mice and to identify their surface markers.</p><p><b>METHODS</b>Thirty C3H/Hej mice received 10 Pfu MHV-3 intraperitoneally, the CD4-CD8- T cells were isolated using magnetic bead sorting on 0, 4, 15, 30, 40 days post MHV-3 infection. The cytotoxic effects of CD4-CD8- T cells on normal and infected hepatocytes, CD8+ T cells and unrelated-virus (murine cytomegalovirus, MCMV) infected CD8+ T cells were examined by non-radioactive cytotoxicity assay. The surface markers of CD4-CD8- T cells were determined by flow cytometry.</p><p><b>RESULTS</b>MHV-3 infected CD4-CD8- T cells showed significant cytotoxic effect on CD8+ T cells, but not on infected hepatocytes or MCMV infected CD8+ T cells. The analysis of cell surface markers demonstrated that the CD4-CD8- T cells are a completely new T cell subset.</p><p><b>CONCLUSIONS</b>CD4-CD8- T cells have significant cytotoxic effect on virus specific CD8+ T cells in MHV-3 infected C3H/Hej mice, which suggests that CD4-CD8- T cells have immune modulatory functions in the development of chronic viral hepatitis. The phenotype of these CD4-CD8- T cells detected by flow cytometry is TCR alpha beta +CD3+CD4- CD8- CD25- CD28- CD30- CD44+.</p>

Establishment of an in vivo model for duck hepatitis B virus infection using Hubei duckling / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To develop a standard duck hepatitis B virus (DHBV) animal model using a local Hubei species of duck, Ma Ya, and use it as an in vivo experimental system to study antiviral strategies against hepatitis B.</p><p><b>METHODS</b>Two-day-old Ma Ya ducklings were experimentally infected via intraperitoneal injection with the DHBV inocula which was collected from the transfected culture supernatant of 1.5-fold-overlength genome recombinant plasmid. Blood samples were taken twice or thrice a week during post-inoculation for 50 days. Viremia was quantified by serum real-time PCR to show the peak. Antiviral treatment of the DHBV-infected ducklings was started 3 d post-inoculation. The animals received oral administration of lamivudine (3TC) at a dose of 25 mg/kg/d for 5 d, followed by a maintenance therapy thrice weekly for 3 more weeks. Serum was quantified to show the viremia peak and liver biopsy specimens were analysed by Southern blotting and in-situ hybridization at the end of antiviral drug treatment.</p><p><b>RESULTS</b>The experimental infection rate of 2-day-old ducklings was 87.5%. Viremia started to be detectable on day 7 and reached a peak on day 11 post-inoculation, followed by a decrease and fluctuations. Four weeks of oral administration of 3TC led to a significant decrease in viremia peak during. This effect was not sustained, as a rebound in viremia was observed after drug withdrawal. Similarly, the analysis of liver biopsies at the end of 3TC treatment showed a marked decrease in DHBV DNA. However, after drug withdrawal a rebound of intrahepatic DHBV DNA was observed in duck livers.</p><p><b>CONCLUSION</b>The Hubei duck model with experimental DHBV infection of transfected supernatant is more suitable for the hepadnavirus biologic research due to its stability and practicability.</p>

The functions of liver natural killer cells in murine fulminant hepatitis induced by murine hepatitis virus strain 3 / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To investigate the role of liver natural killer cells (NK cells) in murine hepatitis virus strain 3 (MHV-3) induced murine fulminant hepatitis.</p><p><b>METHOD</b>Balb/cJ mice (6-8 weeks, female) were intraperitoneally injected with 100 PFU MHV-3. The numbers of NK cells in their livers, spleens, blood and bone marrow and the expression of CD69 on liver NK cells at 0, 24, 48 and 70 h after MHV-3 infection were analyzed by flow cytometry. The cytotoxic activity of liver NK cells was detected by a non-radioactive cytotoxicity assay. The levels of IFN gamma produced by hepatic NK cells were detected by intracellular cytokine staining.</p><p><b>RESULT</b>Following MHV-3 infection, the proportion of liver NK cells in the mice increased remarkably and reached the peak (43.9%+/-2.3%) at 48 h, then kept a high proportion until the mice were sacrificed. The proportion of NK cells in the peripheral blood also significantly increased and reached the peak (18.0%+/-5.4%) at 48 h. However, there were few NK cells in the peripheral blood at 70 h after infection; the ratio was only 1.3%+/-0.6%. In the spleens and bone marrow, the proportions of NK cells were both significantly decreased from 0 h to 48 h and then slightly increased. The expression of CD69 on liver NK cells was highly up-regulated after the infection and the cytotoxic activity of hepatic NK cells at 48 h was also significantly enhanced. In addition, an increase in IFN gamma production by hepatic NK cells was observed at 48 h.</p><p><b>CONCLUSION</b>After MHV-3 infection, NK cells were recruited to the liver quickly, probably from the spleen and bone marrow. Recruited NK cells remarkably express CD69, enhance cytotoxic activity and IFN gamma production, which correlate with the disease severity of fulminant viral hepatitis. Our results suggest that liver NK cells may play a pivotal role in the pathogenesis of fulminant viral hepatitis.</p>

Experimental study on effect of Shenling Yigan Granule in antagonizing duck hepatitis B virus / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study the viral inhibitory effect of Shenling Yigan Granule (SYG) on duck hepatitis B virus (DHBV) in vivo.</p><p><b>METHODS</b>Chongqing ducks infected with DHBV were used. They were randomly divided into five groups, the small-, medium- and high-dose (1.6 g/kg, 3.2 g/kg, 6.4 g g/kg) SYG groups, the lamivudine positive control group, and the model group. The changes of serum DHBV-DNA, DHB-sAg contents and hepatic pathology were observed.</p><p><b>RESULTS</b>The serum content of DHBV-DNA in the three SYG groups and the positive control group was significantly decreased (P < 0.05), while it was rebounded in the latter at day 7 after stopped lamivudine administration. The change of DHBsAg level was insignificantly in all groups. And the hepatic pathological change in the SYG groups and positive control group was slighter than that in the model control group, but showed insignificant difference in comparison between the SYG groups and the model group (P > 0.05).</p><p><b>CONCLUSION</b>SYG has certain in vivo inhibitory effects on DHBV-DNA.</p>

Differential gene expression profiles in acute hepatic failure model in mice infected with MHV-3 virus intervened by anti-hepatic failure compound / 华中科技大学学报（医学）（英德文版）

Differential gene expression profiles in Balb/cJ mouse model of acute hepatic failure infected with MHV-3 virus intervened by anti-hepatic failure compound (AHFC) and the changes of cytokines regulated by genes were investigated. The Balb/cj mice were divided into AHFC-intervened group and control group randomly. Acute hepatic failure model of Balb/cJ mice infected with MHV-3 virus was established. The survival rate in the two groups was observed. It was found that the survival rate in the AHFC-intervened group and control group was 90% and 50% respectively 48 h after intraperitoneal injection of MHV-3 (P<0.05). Before and after the experiment, the cytokines in peripheral blood of the survival mice were determined, and RNA was extracted from survival mouse liver tissue for the analysis of the differential gene expression by a 36 kb mouse oligonucleotide DNA array. In all the genes of microarray there were 332 genes expressed differently in the two groups, in which 234 genes were up-regulated and 78 genes down-regulated. Through clustering analysis, the differential expression of immune related genes, including TNF receptor superfamily, Kctd9, Bcl-2, Fgl2, IL-8, IL-6, IFN-gamma, TNF-alpha etc. might be related with the curative effectiveness of AHFC. It was suggested that AHFC can balance the immune state of mouse model of acute hepatic failure infected with MHV-3 virus mainly through regulating the expression of immune related genes, decrease the immune damage and inhibit liver cell apoptosis of mouse acute hepatic failure model obviously so as to increase the survival rate of mouse models of acute hepatic failure.

Inhibition of binding peptides on replication of duck hepatitis B virus / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To study the inhibitory effect of binding peptides on duck hepatitis B virus (DHBV) replication in duck hepatocytes.</p><p><b>METHODS</b>Specific binding peptides to duck hepatitis B virus polymerase (DHBVP) were screened by phage display technology (PDT), then were sequenced and synthesized. Binding peptides were added into primary culture of duck hepatocytes infected with DHBV in vitro. DHBV-DNA in the cytoplasm, cell nucleus and medium supernatant was assayed over time.</p><p><b>RESULTS</b>Seven binding peptides were obtained after 3-round screening by PDT. Duck primary hepatocytes infected by DHBV were treated with above obtained binding peptides. The DHBV-DNA levels in medium supernatant and cytoplasm of duck hepatocytes treated with synthesized peptides (the 3rd and the 6th peptide) were significantly lower than those of control cells (P<0.05).</p><p><b>CONCLUSION</b>Specific binding peptides to DHBVP could inhibit the replication of DHBV.</p>