Objective To investigate the clinical significance of HBV pre-core and basal core promoter mutations in chronic hepatitis B using Gene-chip technique.Methods Four spot mutations of 1896,1814 dots in pre-core region and 1762,1764 dots in basal core promoter region were detected in 95 patients with chronic hepatitis B using gene chips.Serum HBV DNA level was measured using fluorescent quantitation polymerase chain reaction(FQ-PCR).Results In the 95 patients with chronic hepatitis B, HBV pre-core or basal core promoter mutations were detected in 91 patients,and the positive rate was 95.8%.The mutation rate of G1896A,A1762T associated with G1764A,and G1896A plus A1762T plus G1764A were 36.3%,70.3% and 24.2% respectively.No A1814C mutant was detected in these patients.Compared with no mutation group,the serum HBV DNA quantity was lower in G1896A plus A1762T and G1764A mutation group,and no significant changes was found in other groups.Conclusion Several different variant sites of HBV can be detected at one time using gene chip technique.HBV pre-core and basal core promoter mutations have effect on the serum HBV DNA level in chronic hepatitis B patients to a certain extent.

ObjectiveTo identify the differences in clinicopathological features between HBeAg-positive and HBeAg-negative chronic hepatitis B (CHB). MethodsA total of 665 CHB patients who were admitted to Ningbo No. 2 Hospital during June 2002 and January 2010 were enrolled, in which 428 were HBeAg-positive and 237 were HBeAg-negative. HBV DNA loads, live histological inflammation grades and fibrosis stages were compared between two groups. SPSS 1 1. 5 was used for statistical analysis.For measurement data, t (for normal distribution) or Mann-Whitney U (for skew distribution) was performed; for enumeration data, Chi-square test was performed; and Pearson correlation analysis was conducted. ResultsLiver inflammatory grade and fibrosis staging in HBeAg-negative CHB patients were more severe than those in HBeAg-positive patients (x2 = 7.92 and 10.35, P ＜ 0. 01 ). The ratio of serum HBV DNA levels ＜ 3, ≥3- ＜ 5 log10 copies/mL in HBeAg-negative CHB patients were significant higher than those in HBeAg-positive patients (x2 = 105.16 and 36.92 ,P ＜0.01 ) ; and the ratio of HBV DNA ≥7 log10 copies/mL in HBeAg-negative group was lower than that in HBeAg-positive group (x2 = 110. 18, P ＜0. 01 ). With the rising of serum HBV DNA levels, liver inflammatory grade and fibrosis staging in HBeAg-positive patients had a descending tendency (r =-0. 287 and-0. 224, P ＜0.01 ), while those in the HBeAg-negative group were ascending (r = 0. 360 and 0. 303, P ＜ 0. 01 ). ConclusionCompared with HBeAg-positive CHB patients, liver inflammation and tissue damage in HBeAg-negative patients are more severe, which need close monitoring.

Objective To investigate the relationship between HBV polymerase gene mutations and HBV genotypes in chronic hepatitis B (CHB) patients resistant to adefovir dipivoxil (ADV).Methods Blood samples were collected from 114 ADV-resistant CHB patients during February 2010 and May 2012.The HBV polymerase regions from serum samples were amplified with real-time PCR,and the PCR products were sequenced.Normal distribution data were presented as x ± s,and non-normal distribution data were presented as M (P25-P75) ; for homogeneous data analysis of variance and LSD-t test were performed.Results There were 8 types of HBV polymerase gene mutations in 114 CHB patients; single point mutation was detected in 102 patients (89.47％) and double or triple points mutations were detected in 12 patients (10.53％).rtA181V/T/S (57.89％),rtN236T (14.91％) and rtA181V/T/S + N236T (9.65％) were the predominant mutations.For 21 patients (18.42％) with HBV genotype B,rtN236T mutation was prevalent (47.62％,10/21) ; while for those with HBV genotype C (93,81.58％),rtA181V/T/S mutation was the predominant (65.59％,61/93).The differences of rtA181V/T/S (x2 =12.269,P ＜0.01) and rtN236T (x2 =18.658,P ＜0.01) mutation rates between B and C genotypes were statistically significant.Conclusion rtA181V/T/S,rtN236T and rtA181V/T/S + rtN236T are the major HBV polymerase gene mutation types in ADV resistant CHB patients,and mutation types are related with HBV genotypes.

OBJECTIVETo explore the effect of silencing the Notch2 gene by small interfering (si)RNA on the proliferation of the HepG2 human hepatocellular carcinoma (HCC) cells.

METHODSNotch2-siRNA was transfected as a liposomal formulation into HepG2 cells. The non-HCC cell lines SG07901 (gastric cancer) and SW620 (colon cancer) were used as controls. The mRNA expression of Notch2 and Hesl were detected by RTPCR, and the protein expression of Notch2 was detected by western blotting. The proliferation of transfected HepG2 cells was assessed by the cell counting kit-8 (CCK8) colorimetric assay.

RESULTSThe untransfected HepG2 cells showed significantly upregulated transcript expression of Notch2, and not of Notch1, Notch3 or Notch4, compared to the other non-HCC cell lines. Following transfection of Noteh2-siRNA into HepG2 cells, the mRNA expression of Notch2 and Hes1 and the protein expression of Notch2 were significantly decreased. The rales of proliferation inhibition in HepG2 following transfection of Notch2-siRNA showed an increasing time-related trend, with 2.64% ± 1620% at 12 h, 38.34% ± 8.80% at 24 h, 70.05% ± 7.80% at 48 h, 70.78% ± 10.00% at 72 h, and 74.22% ± 4.80% at 96 h.The inhibition rate at 24 h of transfection was significantly different from that of the groups of control cells.

CONCLUSIONNotch2 is upregulated in the common HCC cultured cell line HepG2. siRNA-mediated silencing of Notch2 exerts inhibition effects on HepG2 proliferation, suggesting the potential for this approach as targeted therapy for treating HCC.

Carcinoma, Hepatocellular ; pathology ; Cell Proliferation ; Down-Regulation ; Hep G2 Cells ; Humans ; Liver Neoplasms ; pathology ; RNA Interference ; RNA, Small Interfering ; Receptor, Notch2 ; metabolism

Objective To comparatively analyze the immunological characteristics of patients with mild and severe influenza A (H1N1), and to provide the evidence for condition monitoring and treatment . Methods 52 cases with mild influenza A ( H1N1), 152 cases with severe influenza A ( H1N1) and 26 healthy subjects from July 1, 2009 to December 31, 2009 were enrolled in the study.Lymphocyte subsets in peripheral blood were analyzed by flow cytometry and the serum concentrations of interferon -γ( IFN-γ) and interleukin-4 (IL-4) were detected by enzyme-linked immune-sorbent assay (ELISA).Results The total lymphocyte counts were decreased obviously in patients with severe influenza A ( H1N1) than in mild pa-tients and in healthy subjects (P<0.01).The T lymphocyte, NK cells, CD4+T lymphocyte and CD8+T lym-phocyte were also decreased obviously in severe patients than in mild patients (P<0.01).The B lymphocyte and CD4+/CD8+were also decreased in severe patients than in mild patients but had no significant statistical difference (P=0.11, 0.175).The serum IFN-γlevels in patients with mild and severe influenza A (H1N1) were lower than those in control group, especially in patients with severe influenza A (H1N1) (compared with control group and mild group , P<0.01).And the changes of serum IL-4 levels were the same with the former, but there were no statistically significant differences in three groups (P>0.05).Con-clusion Immune dysfunction in patients with influenza A (H1N1) infection is associated with the severity of disease, especially cellular immunity .Therefore, monitoring of the immune system is valuable for the diag-nosis of influenza A(H1N1) infection.

Purpose: The purpose of this study was to evaluate the diagnostic value of soluble Axl (sAxl) in hepatocellular carcinoma (HCC) in comparison with serum α-fetoprotein (AFP). Materials and Methods: Eighty HCC patients, 80 liver cirrhosis patients (LC), 80 patients with hepatitis B virus (HBV) infection, and 80 healthy controls (HC) were enrolled. sAxl levels were measured by an enzyme-linked immunosorbent assay, serum AFP levelswere measured by an electrochemiluminescence immunoassay. Receiver operating characteristic (ROC) curves were used to evaluate diagnostic performances. Results: The results show that levels of sAxl were high expression in patients with HCC (p < 0.05), varied with disease state as follows: HCC > LC > HC > HBV. Logistic regression and ROC curve analysis identified the optimal cut-off for sAxl in differentiating all HCC and non-HCC patients was 1,202 pg/mL (area under the receiver operating characteristic [AUC], 0.888; 95% confidence interval [CI], 0.852 to 0.924) with sensitivity 95.0%, specificity 73.3%. Furthermore, differential diagnosis of early HCC with non-HCC patients for sAxl showed the optimal cut-off was 1,202 pg/mL (AUC, 0.881; 95% CI, 0.831 to 0.931; sensitivity, 94.1%; specificity, 73.3%). Among AFP-negative HCC patients with non-HCC patients, the cut-off was 1,301 pg/mL (AUC, 0.898; 95% CI, 0.854 to 0.942) with a sensitivity of 84.6%, a specificity of 76.3%. The optimal cut-off for sAxl in differentiating all HCC and chronic liver disease patients was 1,243 pg/mL (AUC, 0.840; 95% CI, 0.791 to 0.888) with sensitivity 93.8%, specificity 61.9%. The combination of AFP and sAxl increased diagnostic value for HCC. Conclusion sAxl outperforms AFP in detecting HCC, especially in early HCC and in AFP-negative HCC. Combination sAxl with AFP improved the specificity for early HCC diagnosis. In summary, sAxl is a candidate serum marker for diagnosing HCC.