Effects of cell-mediated immunity induced by intramuscular chitosan-pJME/ GM-CSF nano-DNA vaccine in BAlb/c mice / 病毒学报

This study aimed to investigate the immune adjuvant effect and mechanism induced by chitosan nanoparticles carrying pJME/GM-CSF. In this study, plasmid DNA (pJME/GM-CSF) was encapsulated in chitosan to prepare chitosan-pJME/GM-CSF nanoparticles using a complex coacervation process. Immunohistochemistry was used to detect the type of infiltrating cells at the site of intramuscular injection. The phenotype and functional changes of splenic DCs were measured by flow cytometry after different immunogens were injected intramuscularly. The killing activity of CTLs was assessed using the lactate dehydrogenase (LDH) release assay. The preparation of chitosan-pJME/GM-CSF nanoparticles matched the expected theoretical results. Our results also found that, after pJME/GM-CSF injection, the incoming cells were a mixture of macrophages, neutrophils, and immature DCs. Meanwhile, pJME/GM-CSF increased the expression of MHC class II molecules on splenic DCs, and enhanced their Ag capture and presentation functions. Cell-mediated immunity was induced by the vaccine. Furthermore, chitosan-pJME/GM-CSF nanoparticles outperformed the administration of standard pJME/GM-CSF in terms of DC recruitment, antigen processing and presentation, and vaccine enhancement. These findings reveal that chitosan could be used as delivery vector for DNA vaccine intramuscular immunizations, and enhance pJME/GM-CSF-induced cellular immune responses.

Analysis of the clinical features of and responsive factors on the prognosis in patients with fulminant hepatic failure / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To judge the prognosis in the patients with fulminant hepatic failure and to provide the evidences of correct therapy.</p><p><b>METHODS</b>The clinical features and the indexes which may affect the prognosis of the patients with fulminant hepatic failure were analyzed. Indexes including prothrombin time (PT), the routine biochemical analysis of liver and kidney functions, the plasma levels of glucose and ammonia, cortisol, lipases, amylase, age, gender and complications were analyzed using the software Statistical Product and Service Solutions (SPSS)15.0. The differences between the died and living patients were compared.</p><p><b>RESULTS</b>The mortality of the patients was 65% and the highest was 80% for those with HBV and HEV coinfection. The age and gender had no influence on mortality (P value was 0.423 and 0.728 respectively). HBV infection was the main factor which caused fulminant hepatic failure (52%), The next was hepatitis E virus infection (39%). Among the indexes analyzed, the plasma levels of total bilirubin, usea nitrogen, creatinine, glucose, cholesterol and prothrombin time had positive correlations with the prognosis of the patients (P value was 0.005, 0.001, 0.001, 0.005, 0.010 and 0.049 respectively). The incidence rate of hepatic coma, hepatorenal syndrome, and adrenal insufficiency were higher in the died group than that in the living group (P value was 0.005, 0.012 and 0.025 respectively). But prothrombin time was the only factor which had correlation with the prognosis (P=0.035) analyzed by multivariate logistic regression analysis. The scores of MELD were higher in the died group than that in living group (t=18.236, P<0.01) and especially in the patients with hepatic coma and hepatorenal syndrome. The scores of MELD also had positive correlation with the plasma level of TNFa (r=0.585, P<0.01).</p><p><b>CONCLUSIONS</b>The HBV infection was the main cause of fulminant hepatic failure and HBV and HEV coinfection had the highest mortality. The plasma levels of total bilirubin, cholesterol, glucose , prothrombin time and some complications including hepatic coma, hepatorenal syndrome, and adrenal insufficiency maybe had positive correlations with the prognosis of fulminant hepatic failure. The scores of MELD may predict the prognosis of these patients.</p>

Selection and application of serotypical synthetic peptides derived from hepatitis C virus NS5A region / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Numerous studies have reported a relationship between hepatitis C virus (HCV) genotype and the response to interferon therapy. Despite high sensitivity and specificity, genotyping methods can be performed only on HCV RNA positive samples. Serotyping might be a rapid and cost effective method for determining HCV genotypes, especially in patients with previously undetectable HCV RNA. In this study, an enzyme linked immunosorbent assay (ELISA) method for HCV serotyping with the genotype specific, synthetic peptides derived from HCV nonstructural 5a (NS5A) region was developed.</p><p><b>METHODS</b>Based on 45 sequences, representing HCV genotypes 1 - 6 from Genebank, we synthesised 305 overlapping 30-mer peptides within NS5A region at positions 2182 - 2343 of HCV. All peptides for antigenic reactivity were tested by enzyme immunoassay with 69 human sera with antiHCV positive representing genotype 1 - 6. Forty hepatitis C patient sera were serotyped using serotype specific, synthetic peptides and genotyped by sequencing analysis.</p><p><b>RESULTS</b>The correspondence of amino acids in HCV NS5A region with amino acids in positions 2182 - 2343 was very low among different genotype peptides. The highly conserved sequences were residues 2182 - 2211 (R1), 2272 - 2301 (R7) and 2302 - 2331 (R9): the highly variable 2212 - 2241 (R3) and 2257 - 2286 (R6). Using 305 peptides, antigenic regions were located in R3, R7 and R9. Eighteen peptides from highly conserved region representing genotypes 1 to 6 showed broad immunoreactivity with sera containing antibody to all HCV genotypes. Immunoreactivity of the peptides from highly variable region was stronger with similar genotype sera. Twelve unique peptides showed highly, genotype specific, reactivity with types 1 and 3 sera. Type 2 genotype specific peptides had cross reaction with type 3 serum. No type 4, 5 or 6 specific peptides were selected. The serotyping results showed high agreement with sequencing analysis.</p><p><b>CONCLUSIONS</b>The major antigenic regions in HCV NS5A region were at 2212 - 2241 (R3), 2272 - 2301 (R7) and 2302 - 2331 (R9). Eighteen peptides from highly conserved region show genotype independent, immunoreactivity, useful for antiHCV antibody test. Twelve peptides from highly variable region show genotype 1 and 3 dependent immunoreactivity, useful for determining HCV serotype, especially for patients with previously undetectable HCV RNA.</p>

Immunoreactivity studies on synthetic peptides deriving from immunodominant region of hepatitis C virus NS5a gene / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To identify the location of major immunodominant antigenic region and study the relationship between the gene heterogeneity and immunoreactivity via detecting antigenic reactivity of synthetic peptides deriving from immunodominant region in different genotypes of hepatitis C virus (HCV) NS5a gene.</p><p><b>METHODS</b>In total, 305 non-identical 30-mer long and overlapping by 15 aa peptides derived from HCV NS5a region from codon 2,182 to 2,343 among 45 unique published HCV sequences in GenBank corresponding to different genotype were designed and synthesized. The amino acid sequences of all peptides were compared with DNA Star software. The antigenic reactivity of those peptides was detected with indirect ELISA with both anti-HCV and anti-NS5 positive serum.</p><p><b>RESULTS</b>The sequences showed highly conserved among HCV genotype in regions 2,272-2,301 and 2,302-2,331 as compared to regions 2,212-2,241 and 2,257-2,286. The peptides basing on amino acid residues among 2,212-2,241, 2,272-2,301 and 2,302-2,331 showed stronger immunoreactivity than any other peptides. Eighteen peptides derived from this region showed a broad immunoreactivity, 3 of them could react with 96% of anti-HCV positive sera. Whereas the immunoreactivity of the peptides derived from the region showing highly variable among HCV genotype was found to react more strongly with homologous-genotype sera.</p><p><b>CONCLUSION</b>The major linear antigenic region was located at amino acid residues among 2,212-2,241, 2,272-2,301 and 2,302-2,331; the short synthetic peptide derived from NS5a region at position 2,212-2,241, 2,272-2,301 and 2,302-2,331 can be used for efficient detection of HCV antibody; some peptides showed genotype specific immuunoreactivity.</p>