| Objective: To detect the levels of T lymphote subsets,CD4+CD25+ Tre- gs,IL-10 and IFN-γin peripheral blood in patients with chronic hepatitis B(CHB) and investigate the relevancy and significance between the effective- eness after lamivudine therapy and the change of immune index.Methods: 90 chronic hepatitis B patients which included 66 men and 34 women in our hospital and Shijiazhuang Fifth Hospital from January 2009 to February 2010 were enrolled in this study. They all were treated with lamivudine 100mg/d. And no one had received anti-HBV agent or immunore- gulant 6 months before sampling. The diagnoses and the anti-virus standards were complied with the diagnostic criteria of the 2000 Xi'an Viral Hepatitis Management Scheme issued by the Chinese Society of Infectious Diseases and Parasitology.Serum ALT was measured using an Olympus AU2700 auto-biochemical analyzer. HBV markers: HBsAg,HBsAb,HBeAg,HBeAb,HBcAb, anti-HAV, anti-HCV, anti-HDV, anti-HEV and anti-HIV were detected by commercial enzyme immunoassay kits respectively.Peripheral heparinized blood samples were collected from all the enrolled individuals. For staining of CD4+CD25+Tregs, FITC-anti-CD4 and PE-anti-C- D25 were used. After all the red blood cells were dissolved, 0.02M phosphate buffer saline (PBS) was used to wash the blood samples, then the remaining cells were fixed in paraformaldehyde-PBS and the frequency of CD4+ CD25+ T cells were detected by flow cytometric analysis, gated from lymphocytes.Peripheral heparinized blood samples were collected from all the enrolled individuals. For staining of CD4+ CD8+ T cells, FITC-anti-CD4 and PE-anti-C D8 were used. After all the red blood cells were dissolved, 0.02M phosphate buffer saline (PBS) was used to wash the blood samples, then the remaining cells were fixed in paraformaldehyde-PBS and the frequency of CD4+ CD8+ T cells were detected by flow cytometric analysis, gated from lymphocytes.Peripheral heparinized blood samples were collected from all the enrolled individuals. To detect the levels of IL-10 and IFN-γby ELISA. The detection was did strictly by direction.All data were analyzed by SPSS version 17.0 for Windows software. All data were showed by mean plus or minus standard deviation. Analysis of variance or Dunnett's test was used for comparison in one group. Paired-sam- ple t test was used for comparison between before and after therapy. Spearman correlation analysis was performed between the frequency of CD4+CD25+ Tregs,ALT and HBV DNA load.Results:1. Efficacy of chronic hepatis B patients after lamivudine therapy: 90 chronic hepatitis B patients with 52 weeks'lamivudine therapy were divided into the following three groups according to anti-viral effects: complete-respo- nd group: HBeAg and HBV DNA were negative, the levels of ALT were normal; partial-respond group: One item or two items among HBeAg,HBV DNA and ALT didn't meet the standard of complete-respond. non-respond group: The levels of HBeAg, HBV DNA and ALT failed to meet the standard of complete-respond. There were 32 complete-responders,45 partial-res- ponders and 15 non-responders all of the 90 patients.2. Changes of immune index in each group: Complete-respond group: The levels of CD4+T cells,CD8+T cells and CD4+/CD8+ were increased significantly after 52 weeks'lamivudine therapy. With the extension of the anti-virus time, the freq uencies of CD4+CD25+Tregs in peripheral blood were decreased, the levels of IFN-γ, IFN-γ/IL-10 were increased, the levels of IL-10 were decreased. Partial-respond group: The levels of CD4+T cells and CD4+/CD8+ were increased after 52 weeks'amivudine therapy. The levels of CD8+T cells were no significant change before and after treatment. With the extension of the anti-virus time, the frequencies of CD4+CD25+Tregs in peripheral blood were decreased, the levels of IFN-γ, IFN-γ/IL-10 were increased, the levels of IL-10 were decreased. Non-respond group: The levels of CD4+T cells, CD8+T cells and CD4+/CD8+ were no significant change after 52 weeks'lamivudine therapy. With the extension of the anti-virus time, the frequencies of CD4+CD25+Tregs, the levels of IFN-γ, IFN-γ/IL-10 and IL-10 were no significant change before and after treatment.3. Changes of HBV DNA levels during lamivudine therapy: When treated with 12 weeks, 32 complete-respond patients'HBV DNA levels were less than 5×102 copies/ml. 12 of 43(27.9%)partial-respond patients'HBV DNA levels were less than 5×102 copies/ml. the remaining patients'HBVDNA levels were 1.26×103~8.23×104 copies/ml. 15 non-respond patients'HBV DNA levels were 7.26×105~ 1.23×107 copies/ml. When treated with 24weeks, 14 of 43(32.6%) partial-respond patients'HBV DNA levels were less than 5×102 copies/ml. When treated with 36weeks, 16 of 43(37.2%) partial-respond patients'HBV DNA levels were less than 5×102 copies/ml. When treated with 52 weeks, 15 of 43(34.8%) partial-respond patients'HBV DNA levels were less than 5×102 copies/ml.4. Changes of ALT levels during lamivudine therapy: When treated with 12 weeks, 30 of 32 (94.6%)complete-respond patients'ALT levels were less than 40U/L. 14 of 43(32.5%)partial-respond patients'ALT levels were less than 40U/L. When treated with 24weeks, 32 complete- respond patients'ALT levels were less than 40U/L. 18 of 43(41.9%)partial-respond patients'ALT levels were less than 40U/L. When treated with 36weeks, 19 of 43(44.2%) partial-respond patients'ALT levels were less than 40U/L. When treated with 52 weeks, 18 of 43(41.9%) partial-respond patients'ALT levels were less than 40U/L.Conclusions: During the process of lamivudine therapy, the complete-respond chronic hepatitis B patients'CD4+CD25+Tregs frequencies,IL-10 levels decrease significantly than without treatment, the levels of IFN-γ, CD4+T cells, CD8+T cells, CD4+/CD8+, IFN-γ/IL-10 increase significantly than without treatment. Above immune indexes are no significant changes after 52 weeks'amivudine therapy in non-respond chronic hepatitis B patients. After lamivudine therapy, the replication is inhibited and the changes of above immune indexes may be related to the improvement of secondary immune function which may be caused by the relief of hepatis B virus'inhibition on immune function.
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