Expression of HBcAg in hepatocytes and its association with the efficacy of antiviral therapy / 临床肝胆病杂志

ObjectiveTo investigate the effect of the expression of HBcAg in hepatocytes on the serum level of HBcAb and seroconversion of HBeAg after antiviral therapy with nucleos(t)ide analogues (NUCs). MethodsSerum samples and liver tissue paraffin sections were collected from 101 chronic hepatitis B (CHB) patients who received antiviral therapy with NUCs in Nanfang Hospital and Panyu Central Hospital from January 2015 to June 2018. ELISA was used to measure the serum level of HBcAb, and immunohistochemistry was used to measure the expression of HBcAg in the liver. The GEO database (GSE96851) was analyzed to obtain differentially expressed genes in the liver of patients with HBcAg-positive hepatitis. The two-independent-samples t test was used for comparison of continuous data between two groups; the multiple-independent-samples nonparametric Kruskal-Wallis H test was used for comparison of continuous data between multiple groups, and Dunnett method was used for further comparisons; the chi-square test was used for comparison of categorical data between groups. ResultsThe expression pattern of HBcAg in hepatocytes was classified as absent expression, nuclear expression, cytoplasmic expression, and nuclear/cytoplasmic expression, and according to expression level, HBcAg expression was classified as grades Ⅰ, Ⅱ, Ⅲ, and Ⅳ expression. HBeAg seroconversion rates after 96 weeks of antiviral therapy were 5.88%, 16.67%, 22.73%, and 24.24%, respectively, in the patients with absent expression, nuclear expression, cytoplasmic expression, and nuclear/cytoplasmic expression (χ2=4753, P=0.037), and HBeAg seroconversion rates after 96 weeks of antiviral therapy were 5.88%, 13.04%, 27.59%, and 26.67%, respectively, in the patients with grade Ⅰ, Ⅱ, Ⅲ, and Ⅳ expression (χ2=6.580, P=0.016). There were significant differences in the serum levels of HBcAb-IgM and total HBcAb between the patients with absent expression, nuclear expression, cytoplasmic expression, and nuclear/cytoplasmic expression of HBcAg (HBcAb-IgM: H=9.760, P=0.021; total HBcAb: H=21.46, P＜0.001), and there were also significant differences in the serum levels of HBcAb-IgM and total HBcAb between the patients with grade Ⅰ, Ⅱ, Ⅲ, and IV expression of HBcAg (HBcAb-IgM: H=18.80, P＜0.001; total HBcAb: H=26.03, P＜0.001). The analysis of differentially expressed genes in the liver showed that the expression of antibody-related genes was upregulated in the liver of patients with HBcAg-positive acute liver failure. ConclusionThe expression pattern and level of HBcAg in the cytoplasm of hepatocytes are associated with serum HBcAb, and the measurement of HBcAg may help to predict the efficacy of antiviral therapy with NUCs.

Change and significance of HBcAg-specific Th22 cells in patients with hepatitis B virus infection / 临床肝胆病杂志

ObjectiveTo investigate the changes of Th22 cells, interleukin-22 (IL-22), and transcription factor aryl hydrocarbon receptor (AhR) in patients with hepatitis B virus (HBV) infection and their correlation with clinical indices. MethodsA total of 11 patients with acute hepatitis B (AHB) and 38 patients with chronic hepatitis B (CHB) who attended Eighth Hospital of Xi’an from March 2018 to March 2019 were enrolled as AHB group and CHB group, respectively, and 16 healthy controls were enrolled as HC group. The patients with CHB received tenofovir disoproxil fumarate (TDF) antiviral therapy. Peripheral blood samples were collected for AHB patients at baseline and 6 months after discharge, and peripheral blood samples were collected for CHB patients at baseline and at months 6 and 12 of treatment; peripheral blood mononuclear cells (PBMCs) and plasma were isolated, then PBMCs were stimulated with phorbol ester+ionomycin or recombinant HBcAg, and flow cytometry was used to measure nonspecific CD3+CD4+IL-22+ Th22 cells and HBcAg-specific Th22 cells. ELISA was used to measure the plasma level of IL-22, and quantitative real-time PCR was used to measure the mRNA expression of AhR in PBMCs. The t-test and the paired t-test were used for comparison of normally distributed continuous data between two groups; a one-way analysis of variance was used for comparison of normally distributed continuous data between multiple groups, and the least significant difference t-test was used for further comparison between two groups; the Kruskal-Wallis H test was used for comparison of non-normally distributed continuous data between multiple groups. The chi-square test was used for comparison of categorical data between groups. A Spearman correlation analysis was used to investigate correlation. ResultsThe AHB group had a significantly higher percentage of nonspecific Th22 cells than the CHB group (2.86%±0.45% vs 1.39%±0.33%, t=11.80, P＜0.001) and the HC group (2.86%±0.45% vs 0.80%±0.13%, t=17.30, P＜0.001), and the CHB group also had a significantly higher percentage of nonspecific Th22 cells than the HC group (t=6.825, P＜0.001). The AHB group had a significantly higher percentage of HBcAg-specific Th22 cells than the CHB group (2.97%±0.52% vs 1.22%±0.22%, t=16.58, P＜0.001). The AHB group had a significantly higher plasma level of IL-22 than the CHB group (130.7±39.97 pg/ml vs 66.59±20.83 pg/ml, t=7.176, P＜0.001) and the HC group (130.7±39.97 pg/ml vs 50.63±11.07 pg/ml, t=7.662, P＜0.001), and the CHB group also had a significantly higher plasma level of IL-22 than the HC group (t=2.887, P=0.006). The AHB group had significantly higher mRNA expression of AhR than the CHB group (11.45±3.03 vs 4.81±125, t=10.85, P＜0.0001) and the HC group (11.45±3.03 vs 1.10±0.17, t=13.75, P＜0.001), and the CHB group also had significantly higher mRNA expression of AhR than the HC group (t=11.77, P＜0.001). In both AHB and CHB patients, the percentage of HBcAg-specific Th22 cells was positively correlated with alanine aminotransferase (ALT) level (r=0.638 and 0.830, P=0035 and 0002), and the plasma level of IL-22 was also positively correlated with ALT level (r=0.552 and 0.431, P=0.001 and 0.007). The AHB patients were followed up at 6 months after discharge, and there were significant reductions in the percentage of HBcAg-specific Th22 cells (2.79%±0.56%, t=3.055, P=0.012) and the plasma level of IL-22 (105.8±25.23 pg/ml, t=2.362, P=0.040) from baseline. All CHB patients received TDF antiviral therapy and were followed up at months 6 and 12 of treatment, and there were significant reductions in the percentage of HBcAg-specific Th22 cells (t=4.353 and 3.927, all P＜0.001) and the plasma level of IL-22 (t=4426 and 4.810, both P＜0.0001) from baseline to months 6 and 12 of treatment. ConclusionHBcAg-specific Th22 cells and IL-22 are closely associated with inflammatory response to HBV infection.

Hepatitis B virus core-related antigen: A promising new serum marker for hepatitis B virus / 临床肝胆病杂志

Hepatitis B virus core-related antigen (HBcrAg) is a new serum biomarker for hepatitis B virus (HBV) and is composed of several antigens encoded by the pre-C/C region gene of HBV, including HBcAg, HBeAg, and P22cr precursor protein. There is a poor correlation between HBcrAg and HBsAg and they cannot replace each other. Serum HBcrAg level can reflect the content and transcriptional activity of cccDNA in hepatocytes of patients with chronic hepatitis B, as well as the transcriptional activity of integrated HBV DNA. In addition, HBcrAg can be used to evaluate the antiviral effect of nucleos(t)ide analogues and pegylated interferon- and predict recurrence risk after withdrawal of nucleos(t)ide analogues and the development risk and recurrence of hepatocellular carcinoma after surgery. Therefore, serum HBcrAg is a promising new serum marker for HBV.

Correlation of HBV genotyping and drug resistance mutation in Hohhot,Inner Mongolia / 中国基层医药

Objective To investigate the relationship between HBV -DNA load and serum markers in chronic hepatitis B( CHB) patients in Hohhot,Inner Mongolia,and to explore the mutation of HBV genotype and nucleoside analogue.Methods From January 2015 to December 2017,one hundred and ninety-three CHB patients hospitalized in the People＇s Hospital of Inner Mongolia were selected randomly.The clinical diagnostic criteria for all admitted patients were based on the " Guidelines for the Prevention and Treatment of Chronic Hepatitis B" jointly formulated by the Infectious Diseases Society of 2010. The HBV -DNA load of HBV was detected by real -time quantitative PCR,and the correlation between HBV -DNA load and serum markers was analyzed. Seventy -nine patients were selected from 193 hospitalized patients,PCR-reverse dot blot hybridization was used to analyze HBV genotyping and the drug resistance mutations of different genotypes.Results The differences of HBeAb level and HBV-DNA load between HBeAg positive patients and negative patients were statistically significant(all P<0.001). Of 79 serum specimens of HBV infected people,9 cases(11.4% ) were B genotypes,and 70 cases of C genotype (88.6% ).Of them,25 cases had different loci variation,the rate of variation was 31.6% (25/79),with the unit point rtS213T mutation dominated,accounting for about 24.0% (6/25).Conclusion In Hohhot Inner Mongolia patients with CHB,HBV-DNA load with HBeAg and HBe Ab level are correlated;genotype in patients including B type and C type,which is mainly genotype C;patients with CHB mainly had drug resistance to lamivudine and adefovir dipivoxil, mutations including rtS213T,and hybrid mutation.

Association of serum HBeAg,expression intensity of HBsAg and HBcAg in hepatic tissue with clinical characteristics in 317 chronic hepatitis B patients / 重庆医学

Objective The relationship was analyzed between clinic and the expression intensity of HBsAg and HBcAg with in the hepatic tissue from the serum HBeAg negative group and the positive group.Methods A total of 317 liver biopsy specimens were divided into the HBeAg negative group and the positive group,and the relationship was analyzed between the expression inten sity of HBsAg and HBcAg within the hepatic tissue and their age,gender,ALT level,serum HBV-DNA load,hepatic inflammatory activity grading and fibrosis staging in the two groups.Results Age,ALT level,hepatic inflammatory activity grading and fibrosis of the serum HBeAg negative patients were greater than those of the serum HBeAg positive patients,while their serum HBV-DNA load and the expression intensity of HBcAg within the hepatic tissue were lower than those of the serum HBeAg positive patients (P＜0.05).The expression intensity of HBsAg within the hepatic tissue between the serum HBeAg patients and the serum HBeAg positive patients was not significantly different,and it was not correlated with age,ALT level,hepatic inflammatory grading and fi brosis staging (P＞0.05).After the serum HBeAg turned negative,the expression intensity of HBcAg within the hepatic tissue was decreased (P=0.00,t=12 349.0),and it became positively correlated with the serum HBV DNA load(P=0.007,r=0.251) and its negative correlation with the hepatic inflammatory activity and fibrosis was weakened.Conclusion After the serum HBeAg turned negative,other antigenic components of HBV may still maintain the adequately active immune status within the hepatic tis sue of organisms.After the serum HBeAg turned negative,the expression intensity of HBcAg within the hepatic tissue was de creased and became positively correlated with the serum HBV DNA,while its negative correlation with the hepatic inflammatory activity grade and fibrosis stagings was weakened.

Expression of Hepatocyte Hepatitis B Core Antigen and Hepatitis B Surface Antigen as a Marker in the Management of Chronic Hepatitis B Patients

BACKGROUND/AIMS: We aimed to clarify the association of hepatitis B surface antigen (HBsAg)/hepatitis B core antigen (HBcAg) with the disease status and treatment response in patients with chronic hepatitis B (CHB). METHODS: We investigated 171 biopsy-proven entecavir-treated CHB patients (109 hepatitis B e antigen [HBeAg]-positive, 62 HBeAg-negative). HBcAg expression was positive when ≥10% of hepatocytes stained, and classified into nuclear, mixed, and cytoplasmic patterns. HBsAg expressions were intracytoplasmic (diffuse, globular, and submembranous) and membranous. The histologic activity index (HAI) and fibrosis stage followed Ishak system. RESULTS: In HBeAg-positive patients, older age, increased HAI score, advanced fibrosis, and reduced viral load were observed when HBcAg expression shifted from nucleus to cytoplasm in HBcAg-positive patients, and HBsAg expression from non-submembranous to submembranous in HBcAg-negative patients (all, p<0.05). In HBeAg-negative patients, only intracytoplasmic HBsAg expression patterns had clinical relevance with decreased ALT levels and viremia. In HBeAg-positive patients without favorable predictors of virologic response, negative HBcAg and membranous HBsAg expression predicted greater virologic response (both, p<0.05). The probability of HBeAg seroclearance was higher in patients with increased HAI or lacking HBcAg expression (both, p<0.05). Higher serum HBsAg levels and hepatocyte HBcAg positivity were associated with reduced serum HBsAg during first and post-first year treatment, respectively (both, p<0.05). CONCLUSIONS: Hepatocyte HBcAg/HBsAg expression is a good marker for disease status and predicting treatment response.

Prevalence of occult hepatitis b infection in iranian cancer patients before chemotherapy treatment / Prevalência de infecção oculta da hepatite B em pacientes iranianos com câncer antes do tratamento de quimioterapia

ABSTRACT Background Occult hepatitis B infection is characterized by negative hepatitis B surface antigen (HBsAg) and also detectable hepatitis B virus (HBV) -DNA, with or without hepatitis B core antibody (anti-HBc). HBV reactivation in individuals under immunosuppressive therapy is critical, occurring in occult HBV. Objective In this study, we aimed to determine the prevalence of occult HBV infection among hepatitis B surface antigen negative in cancer patients before receiving chemotherapy. Methods Sera from 204 cancer patients who were negative for HBsAg, were tested for anti-HBc antibodies. The samples that were negative for HBsAg but positive for anti-HBc also examined for HBV-DNA by polymerase chain reaction (PCR). Results Of the 204 HBsAg negative blood samples, 11 (5.4%) samples were positive for anti-HBc antibodies. HBV-DNA was detected in 9/11 (81%) of anti-HBc positive samples. Occult HBV infection in hematological cancers was more than solid cancers, 4.8% and 4.3% respectively. There was no significant difference in HBc antibody positivity based on vaccination, previous blood transfusions, history of familial hepatitis or biochemical parameters (ALT, AST, total and direct bilirubin levels) (P>0.05). Conclusion Screening of occult HBV infection by HBsAg, HBV DNA and anti HB core antibody should be suggested as a routine investigation in cancer patients before receiving chemotherapy.

RESUMO Contexto A infecção oculta da hepatite B caracteriza-se por antígeno de superfície da hepatite B (AgHBs) negativo com vírus detectável da hepatite B (HBV) -DNA, com ou sem anticorpo de núcleo da hepatite B (anti-HBc). A reativação do HBV em indivíduos sob terapia imunossupressora é crítica, originando a infecção oculta pelo VHB. Objetivo Este estudo teve como objetivo determinar a prevalência de infecção oculta pelo VHB entre em pacientes com câncer e com antígeno de superfície da hepatite B negativo antes de receber quimioterapia. Métodos Soro de 204 pacientes com câncer que foram negativos para AgHBs, foram testados para anticorpos anti-HBc. As amostras que foram negativos para AgHBs, mas positivo para anti-HBc foram também examinadas para HBV-DNA, por reação em cadeia da polimerase. Resultados Entre 204 amostras de sangue AgHBs negativas, 11 (5,4%) foram positivos para anticorpos anti-HBc. HBV-DNA foi detectado em 9/11 (81%) das amostras positivas de anti-HBc. Infecção oculta de VHB em câncer hematológico foi maior que em cânceres sólidos, 4,8% e 4,3% respectivamente. Não houve diferença significativa na positividade anti-HBc, com base na vacinação, transfusões de sangue anteriores, história de hepatite familiar ou parâmetros bioquímicos (ALT, AST, total e níveis de bilirrubina total) (P & gt; 0,05). Conclusão A triagem de infecção oculta por AgHBs, HBV-DNA e anti-anticorpo de núcleo HB deve ser sugerida como uma investigação de rotina em pacientes com câncer antes de receber a quimioterapia.

An analysis of medication rule for HBeAg-positive chronic hepatitis B based on association rule and complex system entropy clustering / 临床肝胆病杂志

ObjectiveTo investigate the features of traditional Chinese medicine (TCM) and prescription rules in the treatment of HBeAg-positive chronic hepatitis B (CHB). MethodsA systematic search was performed for the articles on the TCM diagnosis and treatment of HBeAg-positive CHB, the information of TCM diagnosis and treatment in medical records were extracted, and a database was established after data standardization. The Traditional Chinese Medicine Inheritance Support System was used to investigate the medication rule. ResultsA total of 100 articles with 135 medical records were included in this study. A total of 220 types of Chinese materia medica were used, among which Bupleurum chinense, Salvia miltiorrhiza, Radix Glycyrrhizae, Atractylodes macrocephala Koidz., and Poria cocos were frequently used. As for the meridian entry of drugs, liver meridian, spleen meridian, and stomach meridian were commonly used. The analysis showed that the core drugs for HBeAg-positive CHB were Bupleurum chinense, Atractylodes macrocephala Koidz., Radix Paeoniae Alba, Radix Curcumae, Salvia miltiorrhiza, Radix Glycyrrhizae, Poria cocos, Polygonum cuspidatum, Astragalus membranaceus, Herba Artemisiae Scopariae, and Hedyotis diffusa. A total of 20 core drug combinations were deduced based on complex system entropy clustering, and 10 new prescriptions were obtained using unsupervised entropy hierarchical clustering. ConclusionIn this study, literature mining and inductive analysis show that the syndromes of stagnation of liver qi and spleen deficiency and liver and gallbladder damp-heat are common syndrome types of HBeAg-positive CHB. The medicine suits and prescriptions refined in this study can be used for reference in clinical practice.

Nuclear Expression of Hepatitis B Virus X Protein Is Associated with Recurrence of Early-Stage Hepatocellular Carcinomas: Role of Viral Protein in Tumor Recurrence

BACKGROUND: Hepatitis B virus (HBV) plays well-known roles in tumorigenesis of hepatocellular carcinoma (HCC) in infected patients. However, HBV-associated protein status in tumor tissues and the relevance to tumor behavior has not been reported. Our study aimed to examine the expression of HBV-associated proteins in HCC and adjacent nontumorous tissue and their clinicopathologic implication in HCC patients. METHODS: HBV surface antigen (HBsAg), HBV core antigen (HBcAg), and HBV X protein (HBx) were assessed in 328 HBV-associated HCCs and in 155 matched nontumorous tissues by immunohistochemistry staining. RESULTS: The positive rates of HBsAg and cytoplasmic HBx staining in tumor tissue were lower than those in nontumorous tissue (7.3% vs. 57.4%, p < .001; 43.4% vs. 81.3%, p < .001). Conversely, nuclear HBx was detected more frequently in tumors than in nontumorous tissue (52.1% vs. 30.3%, p < .001). HCCs expressing HBsAg, HBcAg, or cytoplasmic HBx had smaller size; lower Edmondson-Steiner (ES) nuclear grade, pT stage, and serum alpha-fetoprotein, and less angioinvasion than HCCs not expressing HBV-associated proteins. Exceptionally, nuclear HBx-positive HCCs showed higher ES nuclear grade and more frequent large-vessel invasion than did nuclear HBx-negative HCCs. In survival analysis, only nuclear HBx-positive HCCs had shorter disease-free survival than nuclear HBx-negative HCCs in pT1 and ES nuclear grade 1-2 HCC subgroup (median, 126 months vs. 35 months; p = .015). CONCLUSIONS: Our data confirmed that expression of normal HBV-associated proteins generally decreases in tumor cells in comparison to nontumorous hepatocytes, with the exception of nuclear HBx, which suggests that nuclear HBx plays a role in recurrence of well-differentiated and early-stage HCCs.

The association between hepatic steatosis and HBsAg and HBcAg in chronic hepatitis B patients / 中华消化杂志

ObjectiveTo explore the association between hepatic steatosis and the liver tissue expression of HBsAg and HBcAg in chronic hepatitis B (CHB) patients.MethodsFrom January 2005 to June 2008,a total of 147 CHB patients with hepatic steatosis diagnosed by liver biopsy and other 149 CHB patients without hepatic steatosis but with similar HBV DNA titer were enrolled.The differences of HBsAg and HBcAg immunostaining and liver injury in these two groups were compared.The data were analysed using t test and chi square test.ResultsCompared with non-steatosis group,the average age and body weight index of hepatic steatosis group were higher (t values were -3.31and -6.57,both P＜0.01).The percentage of moderate to severe hepatic inflammation in liver,obvious hepatic fibrosis and the strong positive HBsAg staining was lower (30.6％ vs 15.4％ ； 26.5％vs 12.8％； 23.1 ％ vs 6.7 ％； x2=9.63,8.92,15.76； all P＜0.01),and the percentage of strong positive HBcAg staining was also in downtrend.Compared with degree F1 and F2 of liver steatosis,the percentage of HBsAg and HBcAg strong positive staining in liver tissues of degree F3 and F4 was in downtrend.ConclusionsHepatic steatosis affected the expression of HBsAg and HBcAg in liver tissue of CHB patients.As hepatic steatosis appeared and became more severe,both expression of HBsAg and HBcAg and the degree of liver injury were in downtrend.

The distribution and it's clinical significance of hepatitis B core antigen in hepatocytes of chronic hepatitis B virus infected patients / 中华传染病杂志

Objective To explore the distribution and clinical significance of hepatitis B core antigen( HBcAg) in the hepatocytes of chronic hepatitis B virus infected patients. Methods Paraffin sections were made from 41 liver biopsy samples obtained from chronic hepatitis B virus infected patients. The immuno-fluorescence confocal technique was utilized to analyze the expression level and localization of HBcAg in hepatocytes. The data were analyzed by using Kruskal Wallis test and chisquare test. Results HBcAg expression were detected in 36 (87. 8%) patients, among whom 23 cases had moderate abnormal liver function, 10 with mild abnormal liver function and 3 with normal liver function. Among the cases with moderate abnormal liver function, 6 cases showed the simple membrane-type HBcAg expression, 17 cases showed mixed cytosolic-type and membrane-type HBcAg expression without the nuclear-type expression. Twelve cases with mild abnormal liver function only showed simple cytosolic-type HBcAg expression without membrane-type or nuclear-type expression. In the three patients with normal liver function, HBcAg was expressed in cytoplasm and nuclear but not on membrane. The correlation between HBcAg expression pattern and liver function was statistically significant (χ2 =10. 60, P＜0.01). Conclusion HBcAg expression is directly correlated with liver injury in chronic hepatitis B virus infected patients, which indicates that membrane expressed HBcAg is the target antigen during the immuno-attack of liver.

Study on the immune tolerance status and blood biochemical index of hepatitis B virus transgenic mice / 中华传染病杂志

ObjectiveTo research the hepatitis B virus (HBV) replication and immune tolerance status of transgenic mice for elucidating the pathogenesis of hepatitis B and evaluating new drugs against HBV.Methods SPE grade HBsAg negative nontransgenic and transgenic mice with the same genetic background were recruited in this study.HBsAg,HBeAg and HBV DNA were detected by chemiluminescent method.Pre-S1 and HBcAg were detected by enzyme linked immunosorbont assay (ELISA).Liver pathology was examined and HBsAg expressions at different stages were determined by immunohistochemical staining.The lymphocyte proliferation of mice was detected by flow cytometry and interferon (IFN)γ-producing T lymphocytes was determined by enzyme linked immunospot (ELISPOT).The expressions of Toll-like receptor (TLR)2 and TLR9 in splenocyte suspension and splenic dendrite cells (DC) were determined by double-labeling immunofluorescence.The data were analyzed by t test and F test.ResultsHBsAg,preS1,HBeAg,HBcAg were expressed and HBV DNA was replicated in HBV transgenic mice,while anti-HBs,anti-HBc,and anti-HBe were all negative.There were no obvious pathological changes in liver tissues.HBsAg was expressed in cytoplasm and HBcAg in nucleus of hepatocytes.After stimulated with HBsAg,T lymphocyte proliferation capacity of HBV transgenic mice was (697.6±67.3) cpm,which was much lower than that of nontransgenic mice [( 1315.5 ±191.6) cpm].The number of spot forming cells of IFNγ-producing splenocytes from transgenic mice after HBsAg stimulation was 8.25 ± 1.10,which was obviously lower than that of nontransgenic mice (28.50±4.21) (F=155.967,P=0.000).The expressions of CD11c+,TLR2 and TLR9 on DC from both HBV transgenic and nontransgenic mice were not different significantly (all P＞0.05).The HBsAg expressions in liver tissues were observed in 18-day-old fetal mice and 1-day-old newborn mice.ConclusionsThe HBV transgenic mice can express HBV-related antigens,and are immune tolerant to the antigens.The innate and acquired immunity of the HBV transgenic mice are normal,which is similar to chronic asymptomatic HBV carriers of human.Therefore,HBV transgenic mouse is an ideal animal model.

Endemic hepatitis b and c virus infection in a brazilian eastern amazon region

CONTEXT: Hepatitis B and hepatitis C infection has been an important cause of morbidity and mortality around the world. However there are few investigations regarding the prevalence and possible risk factors for these diseases in Brazil, particularly in Amazon region, where there are some endemic focus. OBJECTIVES: To determine the prevalence of hepatitis B and hepatitis C in the city of Buriticupu, MA, located in the Brazilian Eastern Amazon region, and try to explore the risk factors for these infections in that area. METHODS: Two hundred forty three subjects (46.5 percent male and 53.5 percent female) were investigated. RESULTS: The prevalence of past or current infection of hepatitis B and C virus was, respectively, 40.74 percent and 5.76 percent. Positivity for HBsAg was found in 2.88 percent of the subjects. The prevalence of current infection or chronic virus carriers found was 2.88 percent (HBsAg). There was a statistically significant relationship between the sera-prevalence of anti-HBc and the distance of the residence from the city center which may reflect an indirect association between the infection and precarious conditions of existence. Individuals with age equal or greater than 60 years were also more likely to be anti-HBc positive which could only reflect that older people have a longer history of exposure to hepatitis B infection. The prevalence of hepatitis C is higher than the worldwide estimate. CONCLUSION: Buriticupu may be considered endemic for hepatitis B and C. Hepatitis B infection could be related to precarious living conditions and old age. Hepatitis C was not associated with the variables investigated in the present investigation.

CONTEXTO: Infecção por hepatites B e C tem sido causa importante de morbimortalidade em todo o mundo. Entretanto, há poucas investigações sobre a prevalência e possíveis fatores de risco relacionados a tais doenças no Brasil, especialmente na região amazônica, onde há algumas regiões endêmicas para tais quadros clínicos. OBJETIVOS: Detectar a prevalência de hepatites B e C na cidade de Buriticupu, MA, localizada na região leste da Amazônia brasileira, e tentar investigar seus fatores de risco nessa área. MÉTODOS: Duzentos e quarenta e três indivíduos (46,5 por cento masculinos e 53,5 por cento femininos) foram investigados. RESULTADOS: A prevalência de hepatite C foi de 5,71 por cento (anti-HCV) e a de hepatite B foi de 40,74 por cento (anti-HBc). A prevalência de indivíduos com infecção atual ou com infecção crônica foi de 2,80 por cento (HBsAg). Houve relação estatisticamente significante entre anti-HBc e a distância da residência dos indivíduos do centro da cidade, o que pode refletir uma associação indireta entre tal quadro infeccioso e condições precárias de existência. Indivíduos com idade igual ou maior a 60 anos também apresentaram maior chance de apresentarem sorologia para anti-HBc, o que pode refletir apenas que pessoas mais velhas apresentam história maior de exposição à infecção. A prevalência de hepatite C é maior do que a mundial estimada. CONCLUSÃO: Buriticupu pode ser considerada endêmica para hepatites B e C. Hepatite B pode estar relacionada com precárias condições de vida e idade avançada. Hepatite C não foi associada com as variáveis investigadas na presente investigação.

Effects of PTD-HBcAg induced murine bone marrow-derived dendritic cells maturation on T lymphocyte proliferation in vitro / 中华传染病杂志

Objective To observe the effects of PTD-hepatitis B core antigen (HBcAg) induced murine bone marrow-derived dendritic cells (DCs) maturation on T lymphocyte proliferation in vitro.Methods Bone marrow derived DCs isolated from BALB/c mice were cultured with recombinant granu|ocyte-macrophage colony-stimulating factor (rGM-CSF) and recombinant interleutin-4 (rIL-4)for 5 days.Tumor necrosis factor (TNF)-a,HBcAg and PTD-HBcAg were added to induce DCs maturation.The distribution and localization of intracellular immunofluorescence were observed by confocal microscopy,and DCs phenotypes were analyzed by flow cytometry.The level of IL-12 p70 in the supernatant was detected by enzyme linked immunosorhent assay (ELISA).The proliferation of T lymphocytes was performed by using cell counting kit-8 (CCK-8).All data were analyzed using t test.Results DCs were cultured and identified successfully.Recombinant PTD-HBcAg could penetrate into DCs cytoplasm while recombinant HBcAg was detected on the surface of cells.DCs surface molecules,such as CD80,CD86 and major histocompability complex (MHC) II were upregulated by PTDHBcAg;IL-12 p70 levels induced by 50 mg/L and 100 mg/L recombinant PTD-HBcAg were (142.50±18.31) ng/L and (124.30±15.12) ng/L,respectively,which were significantly higher than those induced by recombinant HBcAg [(42.31±4.21 ) ng/L,t = 9.234 and 9.045,respectively,P＜0.05].The proliferation of T lymphocytes induced by PTD-HBcAg was much higher than that in HBcAg group or positive control TNF-a group.Conclusions PTD-HBcAg could penetrate membrane of DCs and promote the differentiation and maturation of DCs.PTD-HBcAg could up-regulate the expressions of costimulatory molecules on cell surface of DCs,and enhance the ability of DCs on stimulating T lymphocytes proliferation and IL-12 p70 production.

Detection of hepatitis B virus DNA in sera from 18 alcoholic carriers of anti-HBc alone and response to a single dose of hepatitis B vaccine / Pesquisa de DNA do vírus da hepatite B no soro de 18 alcoolistas portadores de anti-HBc isolado e resposta a uma dose da vacina para hepatite B

To evaluate the possibility of occult hepatitis B virus (HBV) infection in alcoholics carriers of "anti-HBc alone", and to verify the behavior of this serological pattern after a single dose of hepatitis B vaccine, 18 alcoholics who had this serological profile were evaluated by the polymerase chain reaction method, and 17 of them were vaccined. All were negative for HBV DNA. Nine (52.9 percent) of those vaccined had anamnestic response, mainly those with positive anti-HBe (8/10; 80 percent). "Anti-HBc alone" was compatible with low levels of anti-HBs in half of the patients, and probably with false positive results for anti-HBc in the others.

Para avaliar a possibilidade de infecção oculta pelo vírus da hepatite B em alcoolistas com "anti-HBc isolado" e a resposta a uma dose da vacina para a hepatite B, 18 alcoolistas com este perfil sorológico foram avaliados pelo método de reação em cadeia da polimerase e 17 deles foram vacinados. Todos tiveram negativos os exames para o VHB DNA. Nove (52,9 por cento) dos vacinados tiveram resposta anamnéstica, principalmente aqueles com anti-HBe positivo (8/10; 80 por cento). "Anti-HBc isolado" foi compatível com baixos títulos de anti-HBs em metade dos pacientes, e provavelmente com resultado falso-positivo para o anti-HBc nos demais.

Trans-activator of transcription protein transduction domain mediated cell membrane penetration of HBcAg / 中华传染病杂志

0bjective To observe the cell membrane penetration of protein transduction domain (PTD)-HBeAg fusion protein in vitro.Methods The sequence of trans-activator of transcription (Tat)-PTD was synthesized and the whole HBcAg gene was amplified by polymerase chain reaction (PCR).Overlap extension PCR was employed to fuse Tat-PTD and whole HBcAg gene.Then the fusion gene was cloned into prokaryotic expression vector pMAL-c2X.The correct vector was transformed into E.coli Rosetta-gamiTM 2(DE3),and the protein was induced by isopropyl β-D-1-thiogalactopyranoside(IPTG).Western blot was used tO identify the protein. Furthermore,the fusion protein PTD-HBcAg was purified by affinity chromatography.HBcAg protein expressed using the same methods was employed as eontr0l.The purified protein was added tO HuH-7 cell culture,then the transduction of PTD-HBcAg and HBcAg in cells were detected by indirect immunofluorescence assay (IFA).Results The fusion protein was effectively expressed in E. Coli and purified by affinity chromatography.Both purified PTD-HBcAg and HBcAg could be recognized by HBeAg monoclonal antibody in Western blot analysis.IFA visualization showed that PTD-HBeAg could be introduced into HUH-7 ceils while HBcAg only could not be detected in cells.Conclusions PTD-HBcAg fusion protein can be expressed effectively and purified in prokaryotic expression system.PTD could mediate HBcAg penetrating eell membrane into the cells.

The expression profile of Toll-like receptor on monocyte-derived dendritic cells in patients with chronic hepatitis B virus infection / 中华传染病杂志

Objective To evaluate the expression of Toll-like receptor(TLR)on the monocyte- derived dendritic cells(DC)from chronic hepatitis B(CHB)patients and to analyze the expression pro- file and significance of the TLR such as TLR3,TLR4,TLR?,TLR8 and TLRg,which are associat- ed with immune response to viral infection.Methods Peripheral blood mononuclear cell(PBMC) centrifugated by the hydroxyethyl starch(HES)centrifugation were cultured and induced into DC by granulocyte-maerophage colony stimulating factor(GM-CSF)and interleukin-4(IL-4),and their mor- phology and phenotype were detected by the inverted microscope and flow cytometry respectively. Monocyte-derived DC were obtained from 10 chronically hepatitis B virus(HBV)-infected patients and 15 healthy volunteers.TLR3,TLR4,TLR7,TLRS,TLR9 expression on immature and mature DC were analyzed by FACS Calibur.DC was pulsed with HBcAg on day 3 and 5,then DC maturation and ability to process HBcAg and to stimulate autogeneic T cells were evaluated.Results Monocyte- derived DC developed different TLR expression patterns as they went through different maturation stages.TLR7,TLR8 expressions on immature DC and TLR3,TLR7 expressions on mature DC were lower in CHB than in control(for TLR7,TLR8 expression on immature DC:75.9%,1.0%vs 98.4%,15.4%,P

Observations on immune responses in mice induced by co immunizations of DNA vaccine of HBcAg and plasmids encoding interleukin 12 and interleukin 18 / 中华传染病杂志

Objective To observe the specific immune responses in mice induced by co immunization of DNA vaccine of HBcAg and plasmids encoding interleukin 12 and interleukin 18. Methods The mice were divided into following groups: vector alone, DNA vaccine of HBcAg alone, DNA vaccine of HBcAg plus plasmid of interleukin 12, DNA vaccine of HBcAg plus plasmid of interleukin 18, and DNA vaccine of HBcAg plus plasmids of interleukin 12 and interleukin 18. The mice were immunized with above DNA constructs by intramuscular injections. The levels of anti HBc and its isotypes(IgG1,IgG2a) in sera, and the level of IFN ? in supernatant of spleno lymphocyte cultures were measured by ELISA methods. CTL acti vities of spleno lymphocyte were detected with LDH release assay. Results Mice in all groups except for vector alone were sera positive for anti HBc. Comparing with group of DNA vaccine of HBcAg alone, groups of DNA vaccine of HBcAg plus plasmid of interleukin 12, DNA vaccine of HBcAg plus plasmid of interleukin 18, and DNA vaccine of HBcAg plus plasmid of interleukin 12 and interleukin 18 showed much higher end point titers of anti HBc( P

Humoral and cellular immunogenecity of nucleic acid vaccine of hepatitis B core antigen in rhesus monkeys / 中华传染病杂志

Objective To observe humoral and cellular immunogenecity of nucleic acid vaccine of hepatitis B core antigen (HBcAg) in rhesus monkeys. Methods Rhesus monkeys of the experimental group and the control group received intramuscular injections of a HBcAg nucleic acid vaccine(pJW4303/HBc) and a control plasmid (pJW4303), respectively. Anti HBc titers, isotypes of anti HBc IgG in sera of the rhesus monkeys pre and post vaccinations, and IFN ? as well as IL 4 levels in the culture supernatant of PBMC isolated from the monkeys were detected by an enzyme linked immunoabsorbent assay. HBcAg specific proliferation activities of PBMC in the monkeys were measured by 3H TdR incorporation assay. Results It was observed in rhesus monkeys of experimental group an obvious anti HBc response after immunization with HBcAg nucleic acid vaccine. The major isotypes of anti HBc IgG was IgG2 and IFN ? was predominant compared with IL 4 in the culture supernatant of rhesus monkeys' PBMC, both indicating Th1 type of immune responses. HBcAg specific proliferation activities of PBMC in the experimental group were significantly stronger than those in the control group. Conclusions The nucleic acid vaccine based on HBcAg shows a good humoral and cellular immunogenecity in rhesus monkeys.

Expression of hepatitis B virus core gene in Pichia pastoris / 中国病理生理杂志

AIM: To study the expression of hepatitis B virus core gene in Pichia pastoris and to obtain high-level expressed recombinant HBcAg with good immunoreactivity and high specificity. METHODS: HBV core gene was amplified by PCR from plasmid pHBV1 which contained HBV whole DNA sequence. The PCR product was cloned into pGEM-T vector by TA cloning strategy. After confirmed by DNA sequence analysis, the gene of interest was inserted into the yeast expression vector pPIC9. The recombinant plasmid pPIC9-cAg was constructed and transformed into GS115 by electroporation. The recombinant yeast GS115 was induced by 0.5% methanol. The expressed product was analysed by SDS-PAGE,Western blot and ELISA. RESULTS: The restriction analysis and DNA sequence analysis proved that HBV core gene had already been cloned to yeast expression plasmid pPIC9. The expressed HBcAg existed in SDS-PAGE. Good immunoreactivity and high specificity of the recombinant HBcAg have been proved by ELISA and Western blot. The titre of the recombinant HBcAg in the cell lysate was 1∶ 12 800 . CONCLUSION: The recombinant plasmid pPIC9-cAg was successfully constructed. The recombinant HBcAg with good immunoreactivity and high specificity was successfully expressed in Pichia pastoris expression system and can be applied to further developing HBcAb immunoassay. [