RESUMEN
AIM: To build a hepatitis B virus (HBV)-infected human trophoblast cell model in vitro and determine the mechanism of intrauterine HBV infection. METHODS: Serum from hepatitis B-infected patients containing HBV DNA >10(9) was drawn, subsequently inoculated into human trophoblast cells in vitro (JEG3) and passage-cultured. The supernatants and intracellular HBV viral load of inoculated cells were tested by real-time PCR, and HBV DNA was determined by Southern blot. RESULTS: From inoculation of the 1st passage JEG3 cells, the supernatant viral load of the 1st passage was seen increasing over time, which peaked at 120 h, whereas the HBV viral load was seen decreasing gradually in subsequent passages, and tested negative after the 6th passage. In addition, infected cells of HBV DNA were tested by Southern blot, and showed continual expression in the subsequent cell passages 1-5 while passage 6 was negative. HBsAg was tested as positive from different passages 1-5, and its concentration was also seen decreasing with each subsequent passage cultured until the 6th passage when it tested negative. CONCLUSION: HBV could infect human trophoblast cells (JEG3) in vitro, and it showed continual expression in subsequent cell passages 1-5.
Asunto(s)
Virus de la Hepatitis B/crecimiento & desarrollo , Trofoblastos/virología , Southern Blotting , Línea Celular Tumoral , ADN Viral/genética , Humanos , Reacción en Cadena de la Polimerasa , Carga Viral , Cultivo de VirusRESUMEN
OBJECTIVE: To investigated the relationship between the serum levels of Th1/Th2 cytokines and the progress of viral hepatitis C and the outcome of interferon therapy. METHODS: Serum cytokine detection used the method of ELISA. HCV genotype were classified by direct sequencing. HCV RNA loads were determined by fluorescence quantitative PCR. RESULTS: The levels of IL-2 and TGF-beta in serum of patients with chronic hepatitis C were lower hut IL-5 was higher than those of normal control. The level of IL-6 was positively related to the sera level of ALT and was negatively related to sera HCV RNA load. Patients of HCV genotype 1 had higher sera quantities of IL-6 than those of genotype 2 and patients of genotype 2a had lower sera quantities of IL-2 than those of 2b. The levels of IL-2 had the tendency to decrease whereas IL-6 had the tendency to increase when time went on. The level of TGF-beta increased at early phase but decrease later. There were no difference of all cytokines detected between the groups of response and nonresponse before interferon therapy, hut the quantity of serum IFN-gamma were increased after interferon therapy in the response group. CONCLUSION: The tested cytokines co-participate in the pathogenesis of chronic hepatitis C and have the relationship with the clinical manifestations of the patients. There were no correlation between the levels of Th1/Th2 cytokines in the serum before IFN treatment and the result of IFN therapy. Increasing IFN-gamma in the serum induced by IFN treatment is associated with sustained virological response.
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Citocinas/sangre , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/inmunología , Interferones/administración & dosificación , Células TH1/inmunología , Células Th2/inmunología , Adulto , Anciano , Citocinas/inmunología , Femenino , Hepacivirus/genética , Hepacivirus/inmunología , Hepacivirus/aislamiento & purificación , Hepatitis C Crónica/sangre , Humanos , Masculino , Persona de Mediana Edad , Resultado del TratamientoRESUMEN
AIM: To explore the mechanism of intra-uterine transmission, the HBV infection status of placental tissue and in vitro cultured placental trophoblastic cells was tested through in vivo and in vitro experiments. METHODS: A variety of methods, such as ELISA, RT-PCR, IHC staining and immunofluorescent staining were employed to test the HBV marker positive pregnant women's placenta and in vitro cultured placental trophoblastic cells. RESULTS: The HBV DNA levels in pregnant women's serum and fetal cord blood were correlated. For those cord blood samples positive for HBV DNA, their maternal blood levels of HBV DNA were at a high level. The HBsAg IHC staining positive cells could be seen in the placental tissues and the presence of HBV DNA detected. After co-incubating the trophoblastic cells and HBV DNA positive serum in vitro, the expressions of both HBsAg and HBV DNA could be detected. CONCLUSION: The mechanism of HBV intra-uterine infection may be due to that HBV breaches the placental barrier and infects the fetus.
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Virus de la Hepatitis B , Hepatitis B/transmisión , Hepatitis B/virología , Transmisión Vertical de Enfermedad Infecciosa , Placenta/virología , Complicaciones Infecciosas del Embarazo/virología , Útero/virología , Adulto , Células Cultivadas , ADN Viral , Femenino , Sangre Fetal/virología , Antígenos de Superficie de la Hepatitis B , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/inmunología , Humanos , Recién Nacido , Masculino , Embarazo , Trofoblastos/citología , Trofoblastos/virologíaRESUMEN
OBJECTIVE: To investigate the relationship between hepatitis C virus (HCV) genotype, serum viral load and ALT levels, and the factors associated with the viral relapse after IFN treatment in patients with chronic hepatitis C. METHODS: The HCV RNA levels were determined with Cobas Amplicor Monitor Test, version 2.0, and HCV genotypes were examined by means of PCR products of 5' NTR digested with restriction endonucleases. The patients with chronic hepatitis C were treated with PEG-IFN alpha -2a and Roferon-A for 24 weeks. Those with a viral response after 24 week treatment were followed for an additional 24 weeks. The association of clinical characteristics, such as sex, age, the way of the HCV infection, IFN treatment history and platelet counts, and the HCV genotype, virus load and medicine used for the viral relapse after IFN treatment were analyzed. RESULTS: Of the 208 chronic hepatitis C patients, the ALT levels were not related to HCV RNA levels (r = 0.093, P > 0.05). No difference of ALT levels between HCV genotypes was found, and the HCV RNA load was also of no difference between HCV genotype 1 patients and non 1 patients. Of the 119 patients with viral response after 24 week treatment, 58 cases (48.7%) relapsed after another 24 week's follow-up. Relapse was not significantly related to the clinical characteristics, such as sex, age, mode of the infection, treatment history of IFN, AST/ALT ratio, platelet counts and the baseline viral load. Among patients with genotype 1 virus, the relapse rate was significantly higher than those patients with non-genotype 1 virus (54.5% vs 32.1%, P=0.039). The relapse rate after PEG-IFN alpha -2a treatment was lower than that of Roferon-A treatment (47.0% vs. 52.8%), but not significantly. CONCLUSION: The viral relapse of chronic hepatitis C patients after IFN treatment was significantly associated with the genotypes of the HCV.
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Antivirales/uso terapéutico , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/virología , Interferón-alfa/uso terapéutico , Femenino , Genotipo , Hepacivirus/genética , Humanos , Interferón alfa-2 , Masculino , Persona de Mediana Edad , ARN Viral/sangre , Proteínas Recombinantes , Recurrencia , Resultado del Tratamiento , Carga ViralRESUMEN
OBJECTIVE: To evaluate the efficacy and investigate the influencing factors of the interferon (IFN) retreatment for patients with chronic hepatitis C relapsed after a previous IFN treatment. METHODS: A retrospective study was designed to analyze the retreatment with IFN of 60 relapsed chronic hepatitis C patients. All patients were from a randomized, opened and multi-center clinical trial about the efficacy and security of PEG-IFNalpha-2a compared to CIFNalpha-2a in the treatment of chronic hepatitis C in China. There were 35 patients treated with PEG-IFNalpha-2a and 25 with CIFNalpha-2a. The main parameter to evaluate the efficacy was sustained viral response (SVR) rate. The influence of viral concentration in serum, genotype and drug categories on the responses to IFN were analyzed. RESULTS: For all the patients, the end of treatment virus response (ETVR) and SVR rates were 55.00% and 35.00% respectively. ETVR rate of PEG-IFNalpha-2a was significantly higher than that of CIFNalpha-2a (74.29% and 28.00% respectively, P < 0.01). SVR rate of PEG-IFNalpha-2a was also markedly higher than that of CIFNalpha-2a (45.71% and 20.00% respectively, P < 0.05). However, there was no significant difference between the high and low viral load groups. Among the patients with genotype 1, ETVR and SVR rates of PEG-IFNalpha-2a (75.00%, 45.83%) were significantly higher than those of CIFNalpha-2a (22.22%, 11.11%), (P < 0.01, P < 0.05 respectively), but in patients with genotype non-1, there were no such differences between the two groups. CONCLUSION: Some relapsed patients were not responsive to the IFN retreatment. The efficacy of PEG-IFNalpha-2a was superior to CIFNalpha-2a. The conventional IFN was not suggested to be used in the relapsed cases with genotype 1. The viral load was not associated with the efficacy of IFN retreatment.
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Antivirales/uso terapéutico , Hepatitis C Crónica/terapia , Interferón-alfa/uso terapéutico , Interferones/uso terapéutico , Polietilenglicoles/uso terapéutico , Adulto , Femenino , Humanos , Interferón alfa-2 , Interferón beta , Masculino , Persona de Mediana Edad , Proteínas Recombinantes , Recurrencia , Estudios RetrospectivosRESUMEN
BACKGROUND: This study was undertaken to investigate the predictive factors of sustained viral response in roferon-A or pegasys treated chronic hepatitis C patients after logistic regression analysis of the factors that might be associated with the therapeutic effects of interferon (IFN). METHODS: All patients enrolled into this randomized, open and multi-center controlled trial were divided into two groups randomly and treated with pegasys and roferon-A for 24 weeks, then followed up for another 24 weeks. Before treatment, hepatitis C virus (HCV) genotype was determined, and HCV-RNA in serum was detected before and at the end of treatment and follow-up. HCV-RNA turning negative was considered the major index for evaluating the therapeutic effect. The clinical characteristics including gender, age, infection route of HCV, treatment with IFN, platelet count, AST/ALT ratio and treatment drugs were analyzed by logistic regression. RESULTS: Intention to treat (ITT) and per-protocol (PP) population groups have 208 and 197 patients respectively. In the PP group, after treatment for 24 weeks, the response rates of female patients aged less than 50 years, infected through non-transfusion, relapsed after IFN treatment, and presented with a AST/ALT ratio/=1, virus load equal or more than 8 x 10(5) IU/ml, and genotype 1 infection, and treated finally with roferon-A. But, at the end of follow-up, the patients with a AST/ALT ratio>/=1 and virus load more than 8 x 10(5) IU/ml had a higher rate of sustained response than did those with a AST/ALT ratio
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Farmacorresistencia Viral , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/patología , Interferón-alfa/uso terapéutico , Adolescente , Adulto , Relación Dosis-Respuesta a Droga , Esquema de Medicación , Femenino , Estudios de Seguimiento , Genotipo , Hepacivirus/genética , Anticuerpos contra la Hepatitis C/efectos de los fármacos , Anticuerpos contra la Hepatitis C/inmunología , Humanos , Interferón alfa-2 , Modelos Logísticos , Masculino , Persona de Mediana Edad , Farmacogenética , Polietilenglicoles , Valor Predictivo de las Pruebas , ARN Viral/análisis , Proteínas Recombinantes , Valores de Referencia , Medición de Riesgo , Método Simple Ciego , Resultado del Tratamiento , Carga ViralRESUMEN
BACKGROUND: Some factors have been reported to be associated with a greater likelihood of sustained viral response (SVR) in the interferon (IFN) treatment of chronic hepatitis C. The factors include HCV genotype, HCV RNA level in serum, state of liver disease, baseline body weight, age, sex, and race. The aim of this trial was to investigate the influence of HCV genotype on the IFN treatment of patients with chronic hepatitis C. METHODS: The genotypes of HCV virus were determined in the patients with chronic hepatitis C from several hospitals of China enrolled into the randomized, opened and controlled trial of Peg-IFN alpha-2a (pegasys) treatment, controlled with IFN-alpha-2a (roferon-A). The serum ALT levels and HCV RNA concentrations of the patients were detected before and at the end of treatment and during the follow-up. The influence of HCV genotype on the IFN treatment of patients with chronic hepatitis C was analyzed in intention-to-treat (ITT) population. RESULTS: The HCV genotypes of 202 patients were determined. Of these patients, 158(78.22%) were infected with genotype 1 HCV and 44(21.78%) with genotype non-1. The viral response at the end of treatment (ETVR) and sustained viral response (SVR) rates were 53.80% and 25.32% respectively in patients with genotype 1 HCV, but they were 61.36% and 43.18% in patients with genotype non-1. The difference of SVR between patients with genotype 1 HCV and those with genotype non-1 was significant (P=0.021). After being grouped by the used drugs, the ETVR rates of patients infected with genotype 1 and non-1 HCV were 76.83% and 80.95% in the patients treated with pegasys (P=0.686); but their SVR rates were 35.37% and 66.67% (P=0.01). The viral relapse rate of genotype 1 HCV (55.56%) was significantly higher than that of genotype non-1 HCV (23.53%) (P=0.02). In roferon-A group, the ETVR and SVR rates of patients with genotype 1 HCV were 28.95% and 14.47% respectively, which were lower but not more significant than those of patients with genotype non-1 HCV (43.48% and 21.74%). Moreover, the viral relapse rate of genotype 1 HCV (72.73%) was higher but not more significant than that of genotype non-1 HCV (50.00%) (P=0.21). CONCLUSION: HCV genotype could affect the efficacies, mainly sustained responses, of IFN treatment in patients with chronic hepatitis C, and the effects of IFN are related to drugs and therapeutic course.
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Antivirales/administración & dosificación , Hepacivirus/genética , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/virología , Interferón-alfa/administración & dosificación , Polietilenglicoles/administración & dosificación , Adolescente , Adulto , Anciano , Farmacorresistencia Viral , Femenino , Estudios de Seguimiento , Genotipo , Hepacivirus/efectos de los fármacos , Humanos , Interferón alfa-2 , Masculino , Persona de Mediana Edad , Proteínas RecombinantesRESUMEN
OBJECTIVE: To investigate the predictors of IFN therapy in patients with chronic hepatitis C through making the multivariate logistic regression analysis. METHODS: The patients in the opened, randomized and controlled trial were enrolled into two group, pegasys and Roferon-A group, and were given 24 weeks of pegasys (injection of 180 microg a week), and Roferon-A (injection three times of Roferon-A 3 MU a week) therapy, and followed 24 weeks. The HCV RNA content was determined at the time before, end of treatment and at the followed-up. The association of the response to the treatment with the clinical characteristics including age, gender, way of HCV infection, history of IFN treatment, planet count, AST/ALT ratio, HCV RNA level, HCV genotype and treatment drugs was made trough multivariate logistic regression analysis. RESULTS: The PP population containing 197 cases was analyzed. After controlling for age, gender, way of HCV infection, history of IFN treatment, planet count, AST/ALT ratio, HCV RNA level and treatment, the HCV genotype was not predictor of the end of treatment viral response (ETVR) to IFN therapy (OR 0.604, 95% CI 0.271-1.349, P = 0.219), but was the independent predictor of sustained viral response (SVR) (OR 0.408, 95% CI 0.189-0.881, P = 0.023). After controlling for other characteristics, the treatment drug was the predictors of ETVR (OR 0.105, 95% CI 0.052-0.212, P < 0.001) and SVR (OR 0.255, 95% CI 0.123-0.529, P < 0.001). CONCLUSION: The pegasys using and HCV genotype were the independent predictors of the response to antiviral therapy in chronic hepatitis C.
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Antivirales/administración & dosificación , Hepacivirus/genética , Hepatitis C Crónica/tratamiento farmacológico , Interferón-alfa/administración & dosificación , Polietilenglicoles/administración & dosificación , Adolescente , Adulto , Anciano , Femenino , Estudios de Seguimiento , Genotipo , Hepacivirus/efectos de los fármacos , Hepatitis C Crónica/virología , Humanos , Interferón alfa-2 , Modelos Logísticos , Masculino , Persona de Mediana Edad , ARN Viral/sangre , Proteínas RecombinantesRESUMEN
OBJECTIVE: To investigate the influence of HCV genotype on the IFN treatment of patients with chronic hepatitis C. METHODS: The genotypes of HCV virus were determined in the patients enrolled into the Randomized, opened and controlled trial of Peg-IFN alpha-2a (Pegasys) treatment, controlled with IFN-alpha-2a (Roferon-A), on chronic hepatitis C patients in China. The serum ALT levels and HCV RNA concentration of the patients were detected in the time of before treatment, the end of therapy and follow-up. The influence of HCV genotype on the IFN treatment of patients with chronic hepatitis C was analyzed in intention to treat (ITT) population. RESULTS: The HCV genotypes of 202 cases were determined. 158 (78.2%) cases infected with genotype 1 HCV and 44 (21.8%) cases with genotype non-1. For overall patients, the viral response at the end of treatment (ETVR) and sustained viral response (SVR) rates were 53.8% and 25.3% respectively in patients with genotype 1 HCV, but in genotype non-1 patients those was 61.4% and 43.2%, and the difference of SVR between genotype 1 and non-1 was significant (P=0.021). After grouped by the used drugs, in the patients given Pegasys treatment, the ETVR rates of patients with genotype 1 and non-1 HCV infection were 76.8% and 81.0%, the difference was not significant (P=0.686), but the difference of SVR rates, which were 35.4% and 66.7%, of the patients was significant (P=0.01). The viral relapse rate of genotype 1 was 55.6%; it was significant higher than that of genotype non-1 (23.5%) (P=0.02). In Roferon-A group, the ETVR and SVR rates of patients with genotype 1 HCV were 29.0% and 14.5%, which were lower, but not significant, than those of patients with genotype non-1 (43.5% and 21.7%). The viral relapse rate of genotype 1 was 72.7% and higher, but not significant, than that of genotype non-1 also (50.0%) (P=0.21). CONCLUSION: HCV genotype could affects the efficacies, mainly the sustained responses, of IFN treatment of patients with chronic hepatitis C, and the effects of IFN were related to the kinds of drugs and therapeutic course.
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Antivirales/uso terapéutico , Hepatitis C Crónica/tratamiento farmacológico , Interferón-alfa/uso terapéutico , Polietilenglicoles/uso terapéutico , Genotipo , Hepacivirus/clasificación , Hepacivirus/genética , Hepatitis C Crónica/virología , Humanos , Interferón alfa-2 , Proteínas Recombinantes , RecurrenciaRESUMEN
OBJECTIVE: To study the role of tumor necrosis factor-alpha (TNFalpha) and caspase-3 expression on hepatocyte apoptosis in experimental model of fulminant hepatic failure (FHF). METHODS: Mouse experimental model of FHF was induced by lipopolysaccharide (LPS) and D-galactosamine (D-GalN). Serum TNFalpha level and TNFalpha mRNA expression in liver were tested by ELISA and reverse transcriptase PCR (RT-PCR) method, respectively. The expression of caspase-3 in liver tissue was determined by in situ hybridization. Hepatocyte apoptosis was examined by DNA agarose gel electrophoresis and TUNEL method. TNFalpha, caspase-3 and hepatocyte apoptosis were observed in different stages after drug administration. In addition, we observed the changes of above variables after pretreatment with anti-TNFalpha IgG1. RESULTS: TNFalpha mRNA expression in liver increased significantly (0.91 +/- 0.75, that of normal control was 0.32 +/- 0.10) 2 to 4 hours after administration of LPS and D-GalN. The level of serum TNFalpha increased [(320.50 +/- 86.57) ng/L; that of normal control was (16.66 +/- 7.01) ng/L] and caspase-3 expressed to a slight extent, too. Typical manifestation of hepatocyte apoptosis appeared 8h after drug administration. 8 hours after drug administration the level of serum ALT and total bilirubin (TBil) remarkably increased [(560.66 +/- 60.20) U/L and (163.66 +/- 34.51) micro mol/L, respectively, that of normal control was (23.56 +/- 8.03) U/L and (14.90 +/- 4.80) micro mol/L, respectively] and the expression of caspase-3 reached the peak. 12 hours after drug administration, hepatocyte apoptosis and necrosis coexisted, and the level of serum ALT and TBil reached a peak [(668.30 +/- 78.69) U/L and (203.17 +/- 19.92) micro mol/L, respectively] whereas the expression of caspase-3 already decreased. Hepatocyte apoptosis and liver injury and the expression of caspase-3 could be blocked by antagonizing TNFalpha. CONCLUSIONS: TNFalpha played an important role in hepatocyte apoptosis and liver injury in fulminant hepatic failure. The hepatocyte apoptosis induced by TNFalpha is correlated with the activation of caspase-3. Hepatocyte apoptosis occurs earlier than necrosis.