[Study of the predictive role of serum HBV RNA on HBeAg serological conversion in children with chronic hepatitis B].

Objective: To investigate the role of serum hepatitis B virus RNA (HBV RNA) in predicting HBeAg serological conversion in children with chronic hepatitis B. Methods: 175 children aged 1~17 years with chronic hepatitis B who received interferon α (IFNα) for 48 weeks were selected. Patients were divided into HBeAg seroconversion and non-conversion based on whether HBeAg seroconversion occurred at 48 weeks of treatment.T-test and Mann-Whitney U test were used to compare between groups; chisquare test or Fisher exact probability method was used to compare the frequency between groups of classified variables; and Pearson correlation was used to analyze the correlation between indicators. Univariate and multivariate logistic regression analyses were used to identify influencing factors associated with HBeAg serological conversion. The predictive effect of HBV RNA, HBV DNA, and HBsAg on HBeAg serological conversion was compared and analyzed by the receiver operating characteristic curve (ROC). Results: The seroconversion rate of HBeAg at 48 weeks was 36.0% (63/175). The reduction in HBVRNA levels from baseline to the 12th, 24th, 36th, and 48th weeks of antiviral therapy was significantly greater in the HBeAg serological conversion group than that in the non-conversion group, and the difference was statistically significant between the two groups (P < 0.05). Univariate and multivariate regression analyses showed that age and a decline in HBV RNA levels at week 12 were independent predictors of HBeAg serological conversion. The area under the ROC curve (AUROC) of HBV RNA decline at week 12 was 0.677(95% CI∶0.549-0.806, P = 0.012), which was significantly better than the same period of AUROC of HBV DNA (0.657, 95% CI∶0.527-0.788, P = 0.025) and HBsAg (0.660, 95% CI∶0.526-0.795, P = 0.023) decline. HBV RNA levels decreased (>1.385 log10 copies/ml) at week 12, with a positive predictive value of 53.2%, a negative predictive value of 72.2%, a sensitivity of 77.4%, and a specificity of 57.9% for HBeAg seroconversion. Conclusion: HBV RNA level lowering during the 12th week of antiviral therapy can serve as an early predictor marker for HBeAg serological conversion in children with chronic hepatitis B.

[Application of cervical lifting suture in hemostasis of placenta previa with increta and percreta].

Objective: To evaluate the effect of cervical lifting suture in treatment of placenta previa with increta and percreta. Methods: From January 2016 to June 2017, 65 cases (0.78%, 65/8 322) were diagnosed placenta previa with increta and percreta by prenatal ultrasonic score system and confirmed by intraoperative findings in the department of obstetrics and gynecology of Peking University Third Hospital. Totally 62 cases (0.75%, 62/8 322) were included, because 3 cases underwent hysterectomy with placenta in situ. According to ultrasonic score system, 62 cases were divided into two groups, score 5-9 group (n=42, 67.7%) and score≥10 group (n=20, 32.3%) , cervical lifting suture techniques were all performed in cesarean sections. Demographic and clinical data were collected and compared. Results: (1) There were no significant differences between two groups in age, gravidity, parity, cesarean section history ratio and gestational week of termination (all P>0.05) . (2) In score≥10 group, the median intraoperative bleeding volume was 4 000 ml (1 200-13 000 ml) , while in score 5-9 group, it was 1 600 ml (700-10 000 ml) , intraoperative blood transfusion volume was 2 000 ml (800-8 800 ml) in score≥10 group, while 1 200 ml (0-8 000 ml) in score 5-9 group. The median operation time was 240 minutes (108-1 200 minutes) in score≥10 group, significantly higher than that in score 5-9 group, which was 135 minutes (69-335 minutes; all P< 0.05). In 8 cases for hysterectomy (12.9%,8/62) , 3 cases in score 5-9 group, 5 cases in score≥10 group. (3) In score≥10 group, the rate of postoperative ICU registration was 80% and mean hospitalization time was (6.3±1.7) days, were significantly different, compared with those in score 5-9 group, which were 26%, (4.9±1.9) days. No serious postpartum complications were found in both groups, and there were no significant differences in Apgar score and weight of newborns (all P>0.05) . Conclusion: Cervical lifting suture in placenta previa with increta and percreta could significantly reduce postpartum hemorrhage and retain uterine.

Hepatitis B virus basal core promoter/precore mutants and association with liver cirrhosis in children with chronic hepatitis B virus infection.

We investigated 168 children and analysed the virological characterization and association with disease progression in children with hepatitis B virus (HBV) basal core promoter/precore (BCP/PC) mutants. Among 168 patients with HBV infection (aged 0.5-18 years old, mean 10.1), 86 of them had HBV-related liver cirrhosis (LC) and 82 had HBV-related chronic hepatitis B (CHB). A direct sequencing method was employed to determine the HBV genotypes and the mutations in BCP/PC regions. In all, 133 of them were infected with genotype C viruses (79.17%); only 35 patients (20.83%) were infected with genotype B viruses. Both LC patients and CHB patients had significantly higher ratios of genotype C when compared with the ratios of genotype B (83.7%-16.3% versus 74.4%-25.6%). For patients with CHB, the prevalence of BCP/PC wild-type viruses was 52.4%; but this was only 4.7% in patients with LC. The C1653T, T1753C, A1762T/G1764A and G1896A mutations had a significantly higher prevalence in patients with LC. Among all the patients with genotype B viruses, those with LC had lower HBV DNA levels and higher G1899A mutation frequency than patients with CHB. Among all the patients with genotype C viruses, the patients with LC had higher prevalence of C1653T, A1762T/G1764A and G1896A mutation frequency, higher hepatitis B e antigen (HBeAg) -negative rates, lower viral load, lower elevated alanine aminotransferase and lower anti-HBe positive rates than CHB patients. The HBV BCP/PC variants were more common in HBeAg-negative LC patients than in the CHB group (BCP, 53.4% versus 15.6%; PC, 18.6% versus 3.7%, respectively, p < 0.001). Patients with HBV genotype C viruses, high viral load and C1653T, A1762T/G1764A, G1896A mutant viruses, were more susceptible to developing LC.

Cloning, expression and identification of the gene of human single-chain variable fragment antibody against Hepatitis B virus surface antigen.

Expression of single-chain variable fragment (scFv) antibodies on the surface of bacteriophage is widely used to prepare antibodies with pre-defined specificities. A phage antibody library containing the gene for scFv antibody against Hepatitis B virus surface antigen (HBsAg) was panned with HBsAg immobilized on microtiter plate wells. After five rounds of panning 30 phage clones specific to HBsAg were obtained and one selected clone was sequenced. It was found to consist of 789 bp and its amino acid sequence and specifically detected the respective antigen in the patients but not in healthy persons.

[Screening and cloning of the down-regulation gene by recombinant interferon-B using suppression subtractive hybridization technique.].

BACKGROUND: To construct a subtractive cDNA library of target genes down-regulated in human hepatocarcinoma cell line HepG2 cells treated with IFNB, and clone genes of the down-regulation by IFNB using suppression subtractive hybridization (SSH) technology and bioinformatics techniques. METHODS: The mRNA was isolated from HepG2 cells induced by recombinant interferon-B and 0.9 percent sodium chloride, respectively, then cDNA was synthesized. After restriction enzyme Rsa I digestion, small sizes cDNAs were obtained. Then tester cDNA was divided into two portions and each was ligated to the specific cDNA adaptor 1 and adaptor 2 respectively. After tester cDNA was hybridized with driver cDNA twice and underwent two times of nested PCR, the DNA fragment was subcloned into T/A plasmid vectors to set up the subtractive library. Amplification of the library was carried out with E.coli strain DH5a. The cDNA was sequenced and analyzed in GenBank with Blast search after PCR. RESULTS: The subtractive library of genes down-regulation in HepG2 cells treated with recombination interferon-B was constructed successfully. The amplified library contained 58 positive clones. Colony PCR and sequence analysis was performed in 35 clones randomly, and the full length sequences were obtained with bioinformatics method. Altogether 12 coding sequences were obtained. CONCLUSION: A subtractive cDNA library of genes down-regulation in HepG2 cells treated with IFNB using SSH technique was constructed successfully, which brings some new clues for studying the regulation mechenism of IFNB in liver cells.

[Cloning and sequence analysis of a gene encoding amastin from Leishmania major].

OBJECTIVE: To clone a gene encoding surface protein from Leishmania major. METHODS: Using T. cruzi amastin DNA sequence as a reference, computer search was done on GenBank and dbEST databases by using BLAST path. A Leishmania major DNA library has been constructed and screened by in situ colony hybridization. RESULTS: A 309nt DNA fragment from Leishmania major was found in dbEST. Leishmania major DNA library was screened using specific primers synthesized according to 309 nt DNA sequence, and a full-length coding sequence for Leishmania major amastin was cloned. The coding sequence consisted of 552 nt, and translated into 183 amino acid residues. The homology is 23.5% at amino acid sequence level between Leishmania major and T. cruzi amastins. CONCLUSION: A full length amastin coding gene for Leishmania major has been cloned.