Changes of Treg and Th17 cells balance in the development of acute and chronic hepatitis B virus infection.

BACKGROUND: Many studies suggest that in chronic hepatitis B virus (HBV) infection regulate T (Treg) cells and interlukin-17-producing T help cells (Th17) are mutually antagonistic in the immune response. This study is aimed to reveal the cell differentiation environment and the significance of Treg and Th17 balance in the development of acute and chronic HBV infection. METHODS: Ten patients with acute HBV infection (AHB) and forty-eight patients with chronic HBV infection, including 12 asymptomatic HBV carriers (HBV carriers), 18 chronic hepatitis B patients (CHB) and 18 acute-on-chronic HBV-related liver failure (ACHBLF) were enrolled. Treg and Th17 cells differentiation related cytokine levels were detected by using ELISA. Flow cytometry was employed to count the Treg and Th17 frequency in peripheral blood. RESULTS: Compared to health controls both AHB and ACHBLF patients favoured Th17 cell differentiation, accompanied by a higher proportion of peripheral Th17 cells (P < 0.01) and high level of interleukin-17A (IL-17A) (P < 0.01). However, asymptomatic HBV carriers and CHB were conducive to Treg cell differentiation. In AHB and ACHBLF, peripheral blood IL-17A + CD4 + T cell frequency increased significantly compared with healthy controls. Changes of Treg and Th17 cell frequency were not completely consistent. Both CHB and ACHBLF had lower level of Treg/Th17 ratio than in health control (P < 0.05). Both plasm IL-17A levels (r = -0.72, p<0.001) and Th17 frequency(r = -0.49, p = 0.0003) negatively correlated with plasma HBV DNA load in patients with chronic HBV infection. In addition, both Th17 frequency and plasm IL-17A levels positively correlated with ALT (r = 0.33,p = 0.01 Vs r = 0.29, p = 0.04) and total bilirubin levels (r = 0.72,p<0.0001 Vs r = 0.53, p = 0.0001) in these chronic HBV-infected subjects. However, for AHB there were positive correlation between both Th17 frequency (r = 0.64, p = 0.04) and plasm IL-17A levels (r = 0.69, p = 0.02) with serum ALT levels, but no significant correlation between both HBV DNA level and total bilirubin level with Th17 frequency or plasm IL-17A levels were found. Furthermore, Treg/Th17 ratio was negatively correlated with total bilirubin levels (r = -0.41, p = 0.004) in chronic HBV-infected patients, especially in patients with ACHBLF (r = -0.69, p = 0.001) and positively correlated with viral load in these chronic HBV-infected subjects (r = 0.55, p<0.0001). CONCLUSIONS: Th17 cells are involved in acute and chronic HBV infection, especially in AHB and ACHBLF. CHB and ACHBLF patients manifested obvious Treg/Th17 ratio imbalance, which might be linked to disease progression and the continuous HBV infection.

The study on frequency distribution of regulatory T cells and its functional markers in peripheral blood of chronic hepatitis B / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To explore the role of treg cells and its functional markers in pathogenesis of chronic hepatitis B, and the correlation with disease progression.</p><p><b>METHODS</b>20 cases of healthy control people,53 cases of chronic hepatitis B patients and 24 cases of liver cirrhosis patients were enrolled into the groups. Detecting the frequencies of CD4+ CD25+ Fox3+ cells, CD4+ CD25+ CD127(low) cells, CD39+ treg cells and CTLA-4+ treg cells in treg cells by flow cytometry. Clinical parameters were investigated in the same time.</p><p><b>RESULTS</b>The frequencies of treg cells, CD4+ CD25+ CD127(low) cells and CD39+ treg cells were significant different among healthy control group, CHB group and LC group (P < 0.01). The frequencies of treg cells, CD4+ CD25+ CD127(low) cells and CD39+ treg cells were significantly different in moderate-severe CHB group compared with mild CHB group (P < 0.05, P < 0.05, P < 0.01). In CHB group the frequencies of CD4+ CD25+ Foxp3+ cells were positively correlated with ALT (r = 0. 289, P < 0.05) and AST (r = 0.302, P < 0.05), the frequencies of CD4+ CD25+ Foxp3+ cells had a significant positive correlation with the frequencies of CD4+ CD25+ CD127(low) cells (r = 0.478, P < 0.01).</p><p><b>CONCLUSION</b>The frequencies of treg cells and its functional markers probably had a dynamic tendency in the process of chronic hepatitis B and were closely related with the change of liver function parameters. CD39+ treg cells may be a group of functional treg cells, which indicated that CD39 be a sensitive marker to react treg cells function. In some sense, CD4+ CD25+ CD127(low) cells frequency could represent treg cell frequency.</p>

The degree of HBV suppression with 24 week telbivudine- or lamivudine-treatment in hepatitis B patients predicts the efficacy of the treatment at week 52 / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To investigate the possibilities of an association between the degrees of HBV suppression with nucleoside treatments at week 24 and week 52 in hepatitis B patients and to find a useful predictor for treatment efficacy.</p><p><b>METHODS</b>In this phase III, double-blind, multicenter trial, we compared the efficacy of telbivudine treatment with lamivudine treatment in 332 Chinese compensated chronic hepatitis B patients. The patients were randomly assigned to a daily 600 mg telbivudine treatment group or daily 100 mg lamivudine group for 24 weeks. They were then categorized into 4 groups according to their serum HBV DNA levels (copies/ml) at week 24: a PCR-undetectable group (< 300 copies/ml); a QL- < 10(3) copies/ml group; a 10(3)-<10(4) copies/ml group; and a > or = 10(4) copies/ml group. The treatments were continued as they previously had been for another 28 weeks and the patients serum HBV DNA levels were examined again.</p><p><b>RESULTS</b>At week 52, mean reductions of serum HBV DNA were significantly greater in the telbivudine-treated patients than in the lamivudine-treated group (6.2 log10 vs 5.4 log10, t = 3.6, P < 0.01). Viral resistance was twice as common in lamivudine-treated patients compared to those receiving telbivudine. Telbivudine was well-tolerated with an adverse event profile similar to that of lamivudine. The lower the HBV DNA level achieved at week 24, the higher HBV DNA non-detectable by PCR. ALT normalization and HBeAg seroconversion achieved at week 52, and viral resistance at week 48 decreased parallel to the degree of HBV DNA inhibition.</p><p><b>CONCLUSION</b>HBV DNA PCR-undetectable at week 24 in nucleoside-treated hepatitis B patients suggests a better efficacy at week 52 and lower viral resistance at week 48. The degree of suppression of HBV at week 24 may be used as a predictor of 1-year outcome.</p>

Comparison of immune responses induced by recombinant attenuated Salmonella typhi carrying eukaryotic expression plasmid or prokaryotic expression plasmid of HCV core protein / 生物工程学报

Hepatitis C virus (HCV) core protein is considered to be an attractive candidate for development of protective HCV vaccines. However, this protein may attenuate the induction of systemic immune responses due to its immunomodulatory properties. In this study, we constructed a HCV core gene-containing eukaryotic expression plamid pCI-C, and an in vivo-inducible prokaryotic expression plasmid pZW-C, and transformed the recombinant plasmids into an attenuated Salmonella typhimurium aroA strain SL7207. The resulting bacterial strains SL7207/pCI-C and SL7207/pZW-C were used to orally immunize BALB/c mice, and the immune responses specific to HCV core protein were assessed. Immunization with the recombinant bacteria SL7207/pCI-C led to a persistent drop in percentage of CD3 CD4 T cells, and induced a weak anti-core IgG production. Splenocytes from SL7207/pCI-C immunized mice developed a relatively weak proliferation response and inferior cytotoxic activity compared to those from the mice immunized with bacteria SL7207/pZW-C. Boost immunization with SL7207/ pCI-C yielded limited improvement in immune strength, while the boost with bacteria SL7207/pZW-C significantly enhanced the immune response. These results suggest that de novo synthesis of native HCV core protein may blunt the induction of immune responses. Attenuated S. typhimurium carrying HCV core protein could efficiently activate systemic cellular and humoral responses, and may be a promising strategy for the development of core-based HCV vaccines.

A multicenter, randomized, open-label study of the safety and effectiveness of pegylated interferon alpha 2b and interferon alpha 2b in treating HBeAg positive chronic hepatitis B patients / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To compare the efficacy and safety of PEG-IFNalpha-2b (Peg-Intron) with IFNalpha-2b (Intron A) in treating HBeAg positive chronic hepatitis B patients.</p><p><b>METHODS</b>Two hundred thirty chronic hepatitis B (CHB) patients eligible to the following criteria were enrolled into this study: HBsAg and HBeAg(Abbott kit) positive for at least 6 months, serum HBV DNA > or =10(5) copies/ml (real time PCR, LLQ <10(3) copies/ml) and ALT > or =2 x ULN. After 1:1 randomization, the patients received PegIntron (group A: 1.0 microg/kg body weight, SC, once a week) or Intron A (group B: 3 MIU SC, three times a week) for 24 weeks, and followed up for 24 weeks.</p><p><b>RESULTS</b>(1) In groups A and B, respectively, 80.87% and 83.48% were males; their median ages were 31.0 and 32.0 years old; their median body weights were 65.6 and 65.5 kg; mean serum HBV DNA loads were 8.06 log10 and 7.99 log10; their mean ALT values were 4.17 x ULN and 3.77 x ULN. All of the above parameters between the two groups had no statistically significance differences. (2) At the end of treatment and after follow-up, compared to the Intron A group, the PegIntron group showed better response (including complete and partial response rate, HBV DNA undetectable rate, HBeAg seroconversion rate), but the differences of all of them had no statistical significance. The rate of HBeAg loss was higher in patients receiving PegIntron after follow-up (P = 0.0424). (Table 2) (3) PegIntron and Intron A reduced serum HBV DAN persistently during the therapy. Mean reduction at the end of the treatment was much higher in the PegIntron group than in the Intron group (2.22 log10 copies/ml vs 1.66 log10 copies/ml, P = 0.0283). (4) The overall incidence of adverse events (AEs) in the PegIntron group was similar to that of the Intron A group (94.78% vs 95.65%). The AEs associated with PegIntron administration were similar in nature to those with Interon A, such as influenza-like symptoms, fever, fatigue, headache, nausea, etc and the differences of their incidences had no statistical significance.</p><p><b>CONCLUSIONS</b>The efficacy and safety of PEG-IFNalpha-2b treatment for CHB patients seems to be better than that of IFNalpha-2b; however, further studies are needed to confirm it.</p>

Oxymatrine in the treatment of chronic hepatitis B for one year: a multicenter random double-blind placebo-controlled trial / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To evaluate the efficacy and safety of oxymatrine in the treatment of chronic hepatitis B.</p><p><b>METHODS</b>A multicenter randomized double-blind placebo-controlled trial was conducted. A total of 144 patients with chronic hepatitis B entered the study for 52 weeks; of them 72 received oxymatrine, and 72 received a placebo. Before and after the treatment, clinical symptoms, liver function, serum hepatitis B virus markers, and adverse drug reactions were observed.</p><p><b>RESULTS</b>In 144 patients, 14 were dropped and excluded due to inconsistencies in the included standard. Therefore, the efficacy and safety of 130 patients were analyzed. After being treated for 52 weeks, 70.77% of the patients in the study group had a normal ALT level, and in 43.08% and 33.33% their HBV DNA and HBeAg became negative. In the placebo group, 39.68% had normal ALT level, and 12.31% and 3.33% had their HBV DNA and HBeAg become negative. The rates of complete response and partial response in the oxymatrine group were 23.08% and 58.46%, and in the placebo group they were 3.08% and 44.62%. They were significantly higher in the oxymatrine group than in the placebo group. In the oxymatrine treated patients, 12 weeks after its withdrawal, 60.00% had a normal ALT level, 41.54% and 23.33% had both HBV DNA and HBeAg negative. In the placebo group, 31.75% had a normal ALT level, 3.08% and 1.67% had both HBV DNA and HBeAg negative. The rates of complete response and partial response in the oxymatrine group were 21.54% and 47.69%, and in the placebo group they were 0 and 41.54%. They were significantly higher in the study group than in the placebo group. The adverse reaction rates of oxymatrine in the study and the placebo group were 7.69% and 6.15%, respectively, but there was no statistical significant difference between them.</p><p><b>CONCLUSION</b>Oxymatrine is an effective and safe agent for the treatment of chronic hepatitis B.</p>

Coding single nucleotide polymorphism is an ideal marker for detecting gene imprinting by 5' nuclease assay / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To establish a novel approach for quick and high throughput verification of human gene imprinting.</p><p><b>METHODS</b>By use of a pair of dye-labeled probes, 5' nuclease assay was combined with reverse transcriptase-PCR(RT-PCR) to genotype a coding single nucleotide polymorphism (cSNP), rs705(C/T) of a known imprinted gene, small nuclear ribonucleotide protein N (SNRPN), on both genomic DNA and cDNA of human lymphoblast cell lines.</p><p><b>RESULTS</b>Allele discrimination showed a clear monoallelic expression pattern of SNRPN, which was confirmed by RT-PCR based restriction fragment length polymorphisms. Pedigree analysis verified the paternal origin of expressed allele, which is in consistency with previous report.</p><p><b>CONCLUSION</b>Coding SNP is an ideal marker for detecting gene imprinting by 5' nuclease assay. This approach has also a potentiality to discover differential allele expression of non-imprinted genes in order to find gene cis-acting functional polymorphism.</p>

Levofloxacin distribution in serum and ascites in patients with cirrhosis and evaluation of its efficacy in treatment of spontaneous bacterial peritonitis / 第二军医大学学报

Objective: To study the distribution of levofloxacin in the serum and ascites in patients with cirrhosis and to evaluate its efficacy in treatment of patients with spontaneous bacterial peritonitis(SBP). Methods:（1）Concentration of levofloxacin in the serum and ascites was detected with HPLC in 7 patients with cirrhosis at different time (in the serum: 0.5, 1, 1.5, 2 and 12 h;in the ascites:2, 4, 6 and 12 h). （2）The effects of levofloxacin were observed in treatment of 30 patients with SBP. Results:（1） Levofloxacin was determined in serum and ascites of patients with cirrhosis, whose concentration depended on the duration after oral administration. In serum: tmax was 1.5 h and cmax was (3.913±1.388) μg/ml. In ascites: tmax was 6.0 h and cmax was (2.520±1.213) μg/ml. The levels decreased gradually after reaching peak concentration, then stabilized from 12 h.（2）The symptoms and signs were significantly improved in patients with SBP treated with the levofloxacin. Conclusion: After the oral administration, levofloxacin can both distribute in serum and ascites, and it is efficient in the treatment of the patients with SBP.

Levofloxacin distribution in serum and ascites in patients with cirrhosis and evaluation of its efficacy in treatment of spontaneous bacterial peritonitis / 第二军医大学学报

Objective: To study the distribution of levofloxacin in the serum and ascites in patients with cirrhosis and to evaluate its efficacy in treatment of patients with spontaneous bacterial peritonitis(SBP). Methods:（1）Concentration of levofloxacin in the serum and ascites was detected with HPLC in 7 patients with cirrhosis at different time (in the serum: 0.5, 1, 1.5, 2 and 12 h;in the ascites:2, 4, 6 and 12 h). （2）The effects of levofloxacin were observed in treatment of 30 patients with SBP. Results:（1） Levofloxacin was determined in serum and ascites of patients with cirrhosis, whose concentration depended on the duration after oral administration. In serum: tmax was 1.5 h and cmax was (3.913±1.388) μg/ml. In ascites: tmax was 6.0 h and cmax was (2.520±1.213) μg/ml. The levels decreased gradually after reaching peak concentration, then stabilized from 12 h.（2）The symptoms and signs were significantly improved in patients with SBP treated with the levofloxacin. Conclusion: After the oral administration, levofloxacin can both distribute in serum and ascites, and it is efficient in the treatment of the patients with SBP.

Delineation of chromosome translocations by fluorescence in situ hybridization / 第二军医大学学报

Objective:To delineate the G-banding-sug gested chromosome translocations by fluorescence in situ hybridization (FISH ) technique. Methods: Locus-specific probes, generated by degen erate oligonucleotide-primed PCR (DOP-PCR) technique from yeast artificial chr omosomes (YACs) mapping the regions in question, were used for FISH tests. Results: Among the 2 cases unresolved by G-banding, FISH confirm ed that one had a balanced translocation between chromosome 11 and chromosome 13 , the other had an unbalanced translocation between chromosome 6 and chromosome X.Conclusion: Because of its high sensitivity and specificity, FISH technique is a powerful adjunct to chromosome banding techniques, particula rly for the delineation of subtle chromosome rearrangement(s) and the origin of segment(s).

Delineation of chromosome translocations by fluorescence in situ hybridization / 第二军医大学学报

Objective:To delineate the G-banding-sug gested chromosome translocations by fluorescence in situ hybridization (FISH ) technique. Methods: Locus-specific probes, generated by degen erate oligonucleotide-primed PCR (DOP-PCR) technique from yeast artificial chr omosomes (YACs) mapping the regions in question, were used for FISH tests. Results: Among the 2 cases unresolved by G-banding, FISH confirm ed that one had a balanced translocation between chromosome 11 and chromosome 13 , the other had an unbalanced translocation between chromosome 6 and chromosome X.Conclusion: Because of its high sensitivity and specificity, FISH technique is a powerful adjunct to chromosome banding techniques, particula rly for the delineation of subtle chromosome rearrangement(s) and the origin of segment(s).