Objective To construct the recombinant adeno-associated virus（rAAV）encoding HBsAg-shRNA and to observe its effect on HBsAg and HBeAg expression in HepG2215 cells in vitro. Methods pAAV-shHBs-hrGFP expressing plasmid was constructed by molecular biological techniques. The recombinants were cotransfected with p-RC and p-Helper into AAV-293 cells mediated by calcium acid phosphate. The rAAVs encoding HBsAg-shRNA（rAAV-shHBs-hrGFP）were harvested and infected HepG2215 cells. The silencing effect of this virus on HbsAg and HBeAg gene expression was assessed by ELISA. Results The plasmid of pAAV-shHBs-hrGFP was successfully constructed by identification of restrict enzyme and DNA sequencing. GFP expression was observed about 80% in AAV-293 cells. After cotransfection，the recombinant AAV-shHBs-hrGFP was harvested and the virus titer was tested. The HBsAg and HBeAg expression in HepG2.215 cells was down-regulated markedly after infection of rAAV-shHBs-hrGFP. Conclusion The recombinant AAV-shHBs-hrGFP vector is successfully constructed and can be used as a potential antivirus agent to inhibit HBV replication.

Objective To study the effects of PTD-HBcAg inducing HBV specific cytotoxic T lymphocytes in mice on inhibiting HBV replication in HepG2.2.15 cells. Methods BALB/c mice were subcutaneously immunized at the tail base with 100μl of freshly prepared emulsion containing with PTD-HBcAg,HBcAg,PTD,PBS and an equal volume of CFA. 14days later,T lymphocytes in spleen were isolated and stimulated with corresponding proteins to detect the level of IFN-γ,IL-12,IL-4 and IL-10 in the supernatants. The stimulated T lymphocytes as effective cells were cocultured with HepG2.2.15 cells to determine the levels of HBsAg and HBV DNA and to observe the cytotoxicity of effective cells to HepG2.2.15 cells. Results The cytokines in PTD-HBcAg groups were markedly higher than HBcAg groups and normal controlled groups;In comparison with normal controlled groups and HBcAg groups,HBsAg and HBV DNA decreased more obviously in PTD-HBcAg groups;The T lymphocytes stimulated by PTD-HBcAg could kill HepG2.2.15 cells more effectively than others. Conclusions PTD-HBcAg can highly promote HBV specific CTL to inhibit HBV replication in HepG2.2.15 cells.

Objective To investigate the sampling errors of two different ways of percutaneous liver biopsies. Methods The liver biopsy in 3059 patients were performed by Menghini needle and in 263 patients by automatic Tru-cut needle. Results The average length of samples by Menghini needle were 2.06±0.62cm,longer than by automatic Tru-cut（1.29±0.50cm,t=17.089,P=0.000);In 221 fragmented tissue samples obtained by Menghini,the liver cirrhosis were confirmed in 184 patients（83.26%）;In other 37 patients with fragmented samples without pathological diagnosis at first needle,the second needle by automatic Tru-cut showed liver cirrhosis in 35 patients（94.6%）. Conclusion The samples obtained by Menghini are longer than that by automatic Tru-cut. Nearly all the samples with fragmented tissues by suction needle in patients with hepatitis B are diagnosed as liver cirrhosis.

Objective To investigate the clinical value of PreS1 antigen detection in the diagnosis of chronic hepatitis B. Methods In 421 chronic hepatitis B patients,the PreS1Ag and HBV markers in serum were detected by ELISA,and HBV DNA by fluorescence quantitative PCR. Result The positive rate of PreS1Ag（51.2%） was higher than HBeAg（32.4%）in 367 HBV DNA positive chronic hepatitis B patients（P<0.01）;the positive rates of PreS1Ag in high（10⁵~10⁷ copies/ml and>10⁷ copies/ml,60.2% and 60.0%） were higher than in low serum HBV DNA load（10³~10⁵ copies/ml,41.9%） and in negative group（33.3%,P<0.01）;But in 421 patients,the positive rate of PreS1Ag（48.9%） was lower than HBV DNA（87.2%,P<0.01）. Conclusion PreS1Ag is more sensitive than HBeAg to reflect the HBV replication in chronic hepatitis B patients,but can’t replace the detection of HBV DNA.

Objective To investigate the clinical effect of α interferon combination with Yigan Qingre Jiedu granula on HBeAg positive chronic hepatitis B. Methods Totally 120 patients with HBeAg positive chronic hepatitis B and having signs of internal accumulation of damp-heat type were divided into three groups,40 patients for each group receiving α interferon,α interferon combination with Yigan Qingre Jiedu granula and Yigan Qingre Jiedu granula treatment for six months. Results 3 cases in α interferon group and none in combination group were lost. Combination group was better than herbal group in ALT normalization and herbal group was better than interferon group in symptoms improvement after 6 months’treatment. Rate of HBeAg loss in combination group was better than in the other two groups,with higher HBeAg/Ab seroconversion and HBV DNA undetectable rate but without significant difference. Conclusion α interferon combination with Yigan Qingre Jiedu granula has good therapeutic effect on chronic hepatitis B without obvious adverse effect,but complete response rate needs to be further verification.

Objective To investigate the counts of natural killer cells and T lymphocytes in patients with chronic hepatitis B. Methods The CD3⁺T cells,CD3⁺CD4⁺T cells,CD3⁺CD8⁺T cells and CD3^-CD16⁺CD56⁺NK cells in peripheral blood were counted by FCM. Results The T lymphocytes and NK cells decreased in 56 patients with hepatitis B-related hepatic failure;the CD3^-CD16⁺CD56⁺ natural killer cells in 49 patients with chronic hepatitis B increased as compared to in 41 chronic hepatitis B viral carriers. Conclusions The T lymphocytes and NK cells in patients with hepatic failure decreased while T lymphocytes in patients with chronic hepatitis B with HbeAg positive increased.

Objective To detect HBV cccDNA in serum and peripheral blood monouclear cells in patients with chronic hepatitis B. Methods 95 patients with hepatitis B and 8 with liver transplantation were investigated. HBVcccDNA was detected by using synergy PCR technique. Results We did not detect positive HBV cccDNA in peripheral blood monouclear cells in any patients of our series;the serum positive rates of HBV cccDNA in 20 HBV carriers,in 75 patients with chronic hepatitis B of mild,middle and severe degree and in 8 after liver transplantation were 10%,32%、52% and 76%,and 12.5%（P<0.01）;in patients with serum HBV DNA at<1×10⁵ copies/ml,1×10⁵~10⁸ copies/ml and>1×10⁸ copies/ml,their serum HBVcccDNA positive rates were 15.4%（2/13）,50.0%（16/32）and 76.7%（23/30,P<0.01）. Conclusions HBV cccDNA do exist in sera,but not in peripheral blood monouclear cells in patients with chronic hepatitis B.

Objective To study the changes of serum hepatitis B virus e antigen and HBV DNA load in patients with hepatitis B treated with lamivudine. Methods Emzyme chemiluminescence immunoassay and real-time fluorescent quantitative PCR were performed in detecting the level of serum HBeAg and HBV DNA in 64 patients in 12-month lamivudine therapy. Results HBV DNA turned negative in 49（76.6%）out of the 64 patients,and HBeAg reduced to the level undetectable in 17（26.6%）;11 patients were nonreponded to lamivudine treatment with their serum HbeAg and HBV DNA positive;Four patients had their HBV DNA transiently negative, but it turned to be positive again after 6 month. Conciusion Detecting serum HBeAg dynamically might be useful in evaluation of lamivadine therapeutic effect.

0bjective To evaluate the efficacy and side effects of peginterferon alfa-2a or alfa-2b（PEG-IFNα-2a/α-2b）with ribavirin for treatment of hepatitis C infection,which could be helpful for optimizing the treatment strategy and increasing clinical recovery rate of chronic hepatitis C. Methods 60 patients with chronic hepatitis C were selected and 30 patients received PEG-IFNα-2a with ribavirin therapy for 48 works（A group） and other 30 received PEG-IFNα-2b with ribavirin therapy for 48 works（B group）,and a 24-week of follow-up was performed. HCV-RNA was detected at the baseline,4-week postmanagement,12-week postmanagement,48-week postmanagement,and 24-week after completion of a full course,respectively. The rapid virological response（RVR）rate,early virological response（EVR） rate,end of treatment virological response（ETVR）rate,sustained virological response（SVR）rate,relapse rate and adverse reactions were assessed. Results The RVR,EVR,ETVR,SVR and relapse rate in A group were 46.7%,63.3%,86.7%,80.0% and 6.7%,respectively,which were similar with that in B group（40.0%,56.7%,80.0%,76.7% and 3.3%,respectively,P>0.05）. The safety profile,adverse reactions and tolerance were also similar. Serious adverse events were not observed. Conclusion The sustained virological response and tolerance on peginterferon alfa-2a or alfa-2b with ribavirin for treatment of hepatitis C infection are similar.

Objective To investigate the epidemiologic and clinical features of patients with hepatitis C in recent years. Methods 210 patients with hepatitis C were collected to analyse its epidemiologic and clinical features. Results In 210 patients with hepatitis C,there were 140 male（66.7%）. The range of age was from 5 months to 92 year-old and the average age was 50.5;The main transmission routes were blood product transfusion,injection,sexual and vertical transmission. There was 3.3% of patients co-infected with hepatitis B and HIV. There were 28.1% with cirrhosis and 10% with 2-DM. Conclusions The hepatitis C has five routes of transmission in general,but the routes are not the same in different ages and in different jobs.

ObjectiveTo evaluate clinical efficacy and safety of different doses of interferon（IFN）alfa-2b in the management of patients with chronic hepatitis C. Methods 61 patients with chronic hepatitis C were randomly divided into three groups,who received 48 week treatment and 24 week follow up with IFN alfa-2b（5MU）+ribavarin,IFN alfa-2b（3MU）+ribavarin,ribavarin alone,respectively. Results After 48 week treatment,ETVR were 68.2% and 61.9% respectively in the groups with IFN alfa-2b（5MU）+ribavarin and IFN alfa-2b（3MU）+ribavarin,which were significantly higher than that in the group with ribavarin（5.6%,P<0.01）;After 24 week follow up,SVR were 50.0% and 42.9% respectively in the groups with IFN alfa-2b（5MU）+ribavarin and IFN alfa-2b（3MU）+ribavarin,which were significantly higher than that in the group with ribavarin（0%,P<0.01）;There was no difference between two groups with different doses of IFN alfa-2b in the change of liver fibrosis indexes and side-effects. Conclusion ETVR and SVR of different doses of IFN alfa-2b were satisfactory at the end of treatment,while the effect of 5MU group was better than the group of 3MU. Both of two groups were able to reduce liver fibrosis indexes and improve hepatic histology.

Objective To observe the effect of barbata on transforming growth factor-β1 and tumor necrosis factor-α expression in rats with CCl4-induced liver fibrosis. Methods The model of CCl4 induced liver fibrosis in rats were established. The serum and liver tissue TGF-β1 and TNF-α were detected by immunohistochemistry and ELISA. Enzyme-linked immunosorbent dot technology（ELISPOT）was used to detect the single-cell secretion of TNF-α. Result TNF-α and TGF-β1 were negative in the normal control group and were obviously positive in model group;Barbata inhibited the expression of both TNF-αand TGF-β1in liver tissues and in sera. Conclusion Barbata has an antifibrotic effect by inhibition of cytokines in the liver.

Objective To study the changes of hemorheology in patients with nonalcoholic fatty liver. Methods Sera from 106 patients with nonalcoholic fatty liver and 50 normal persons were obtained for hemorheology studies. Results The blood contrast viscosity,plasma viscosity,erythrocyte aggregation index,hematocrit,body mass index,blood glucose,uric acid,serum alanine aminotransferase,gamma-glutamyl transferase,serum total cholesterol and triglyceride in patients with fatty liver were much higher than in the normal group（P<0.05）. Conclusion Obesity,diabetes,hyperlipemia are the risk factors for nonalcoholic fatty liver.

Objective To explore the relationship between nonalcoholic fatty liver disease and polycystic ovary syndrome. Methods A case control study was performed to compare 47 non-pregnant patients with polycystic ovary syndrome with 47 non-pregnant without polycystic ovary syndrome. Clinic anthropometric variables,the prevalence of nonalcoholic fatty liver disease, insulin resistance,abnormal liver function were determined. Results The occurrence of nonalcoholic fatty liver disease,insulin resistance and elevated aminotransferase was 42.6%,68.1% and 34.0%,higher than those in control group respectively（17.0%,10.6%,8.5%,P<0.01）;Compared with patients without nonalcoholic fatty liver disease,patients with nonalcoholic fatty liver disease had significantly increased body mass index,HOMA-IR,follicle-stimulating hormone,luteinizing hormone（P<0.05）,alanine aminotransferase and testosterone（P<0.01）;Patients with polycystic ovary syndrome and nonalcoholic fatty liver disease were younger than the patients only with polycystic ovary syndrome（P<0.01）;Logistic analysis showed that elevated aminotransferase and HOMA-IR had obvious relationship with polycystic ovary syndrome complicated with nonalcoholic fatty liver disease（ALT OR=1.1,P=0.01;HOMA-IR OR=1.6,P=0.01）. Conclusion Nonalcoholic fatty liver disease is common in patients with polycystic ovary syndrome,which suggests an association between those two conditions. Young women with polycystic ovary syndrome should be screened for liver disease early.

Objective The aim of this study was to investigate the influence of age on intrarenal resistance index（RI）in patients with nonalcoholic fatty liver disease（NAFLD）. Methods 178 patients with NAFLD underwent a conventional abdominal ultrasound and color Doppler flower imaging（CDFI）examination. CDFI signals were taken from arcuate arteries of the cortex in the right and left kidneys,respectively. Results Patients with NAFLD had an increased length of left kidney（P<0.01）and a decreased RI in left than right（P<0.05）;The variable age showed a significant positive correlation on both arcuate arteries. Conclusion Patients with NAFLD show age dependency on arcuate arteries RI and have an lower value than in healthy adults.

Objective To distinguish residual tumor from inflammation after radiofrequency ablation （RFA） in hepatic VX2 carcinoma in rabbits according to the comparative study between CT and pathological findings. Methods CT and pathologic examination were performed in different stages after RFA in rabbits with hepatic VX2 tumors. Results Marginal enhancement band was showed in enhanced CT scan in both residual tumor and inflammation,but the enhancement of inflammation in peripheral liver tissues around the tumors was gradually weakened and disappeared two weeks later. Conclusion The residual tumor and inflammation could not be distinguished through enhanced CT scanning within 1 week after RFA. Low intensity lesions with peripheral enhancement 2 weeks after RFA should be recognized as residual tumor.