Objective To study the expression of programmed death 1(PD-1) on the peripheral T lymphocytes and the serum cytokine levels in different phases of hepatitis B virus (HBV) infection (immune tolerance,immune clearance and non-replicating phases).Methods A total of 105 HBV infected patients in different phases of infection were enrolled in Wuxi No.5 People's Hospital from Apr to Sep 2011,and divided into three groups:the immune tolerance group (35 cases),the immune clearance group (35 cases) and the non-replicating group (35 cases).Meanwhile,20 healthy people were enrolled as controls.The peripheral blood was collected and PD-1 expressions on the surface of T lymphocytes were assessed by flow cytometry.The cytokine levels in different groups were analyzed by enzyme-linked immunosorbent assay (ELISA).The data were analyzed by t test and Pearson correlation analysis.Results The PD-1 expression on surface of CD8+ T lymphocytes of HBV infected patients was (12.35± 3.48)％,which was significantly higher than healthy control group [(4.65±1.21) ％] (t=9.76,P＜0.01).The PD-1 expression on CD4+ T lymphocytes was (4.95±2.87) ％,which was not significantly different from the healthy controls [(4.08±2.14) ％] (t =1.29,P＞0.05).The PD-1 expressions on CD8+ T lymphocytes of patients in the immune tolerance group,the immune clearance group and the non-replicating group were (15.87±3.18)％,(7.69±3.64)％and (10.12±2.84) ％,respectively; that in the immune tolerance group was higher than the immune clearance group (t=10.01,P＜0.01) and that in healthy control group was lower than those in the other 3 groups (t=15.12,3.61 and 8.17,respectively; all P＜0.01).The PD-1 expression on the surface of CD8+ T lymphocytes in chronic HBV infected patients was positively correlated with the serum interleukin-10 (IL-10) level (γ=0.377,P＜0.05),while that was negatively correlated with interferon-γ (IFN-γ) level (r=-0.620,P＜0.05).Conclusions PD-1 is up-regulated on the surface of CD8+ T lymphocytes from chronic HBV infected patients.And PD-1 is negatively correlated with serum IFN-γ levels and positively correlated with serum IL-10 levels.The expression of PD-1 on the surface of CD8+ T lymphocytes influences the balance of helper T cell (Th)1/Th2,which might play a role in the persistence of HBV infection.

Objective To explore the relationship between serum HBV DNA level , expression of PD-1 on CTL and liver functions in patients with hepatitis B virus cirrhosis. Methods 109 HBV hepatic cirrhosis patients with , HBV DNA positive , HBeAg positive and HLA-A2 positive were divided into two groups according to their HBV DNA levels,52 cases in group A, HBV DNA 2-4 log10 copies/mL, 57cases in group B, HBV DNA 5-7 log10 copies/mL , differences of HBV specific CTL surface PD-1 expressions , HBV specific CTL levels and liver functions were compared between two groups. Results HBV specific CTL surface PD-1 expressions of group A were lower than that of group B (t = 11.101, P < 0.01), HBV specific CTL levels of group A were higher than that of group B (t = 24.424, P < 0.01), ALT of group A were lower than that of group B (t = 2.652, P < 0.01), ALB of group A were higher than that of group B (t = 2.347, P < 0.05). Child-pugh rating, grade C of group A was lower than that of group B (χ2= 4.262,P < 0.05). Conclusions HBV specific CTL surface PD-1 expression levels were higher in those hepatic cirrhosis patients with , high level of serum HBV DNA, and HBV specific CTL levels were lower, liver functions damage were much serious. So, lower HBV DNA levels , lower HBV specific CTL surface PD-1 expression levels and higher HBV specific CTL levels can product relatively good effect of anti-viral treatment.

Objective To explore the changes of programmed death receptor-1 (PD-1) in chronic hepatitis B (CHB) patients with different baseline of hepatitis B virus (HBV) DNA levels after treatment with adefovir dipivoxil (ADV).Methods One hundred CHB patients with positive hepatitis B e antigen (HBeAg),1 × 104 copy/mL≤HBV DNA≤1 × 107 copy/mL,and positive human leukocyte antigen-A2 were divided into two groups according to the baseline HBV DNA level:47 cases in low virus load group whose HBV DNA level was ≤1 × 105 copy/mL; 53 cases in high virus load group whose HBV DNA level was＞1 × 105 copy/mL.Both groups were treated with ADV 10 mg/d.Serum HBV DNA,HBeAg seroconversion rate,alanine aminotransferase (ALT) and total bilirubin (TBil) levels of both groups before treatment and 12 months after treatment were compared.Flow cytometry was used to test peripheral blood HBV-specific cytotoxic T lymphocyte (CTL) surface PD-1 and peripheral blood HBV-specific CTL level.Categorical data were tested by x2 test; quantitative data was compared with t-test.Results Peripheral blood HBV-specific CTL surface PD-1 of CHB patients in low virus load group was 20.17 ％±1.69％,which was lower than that in high virus load group (41.38％±2.30％,t =53.02,P＜0.01) ; peripheral blood HBV specific CTL levels in two groups were 0.37％±0.02％ and 0.17％± 0.02％,respectively (t=50.47,P＜0.01) ; ALT and TBil levels in low virus load group were both lower than those of high virus load group (t=13.07,P＜0.01; t=5.06,P＜0.01).Twelve months after treatment,HBV DNA of 25 cases (53.2％) in low virus load group and 10 cases (18.9％) in high virus load group were lower than the detectable level (HBV DNA＜500 copy/mL,x2 =12.89,P＜0.01);HBeAg seroconversion was achieved in 15 cases(31.9％) and 1 case (1.9％),respectively (x2 =16.72,P＜0.01) ; peripheral blood HBV-specific CTL surface PD-1 expression levels were 9.00 ％ ±1.38 ％ and 29.40 ％ ± 3.76 ％,respectively (t =36.80,P＜ 0.01) ; peripheral blood HBV-specific CTL levels were 0.65％±0.10％ and0.48％±0.07％,respectively (t=9.61,P＜0.01).Conclusions After treatment with ADV,along with the decrease of HBV DNA load,HBV-specific CTL surface PD-1 expression decreases,while HBV-specific CTL level increases.The changes in low virus load group are much more remarkable.

Objective To investigate the association between the difference of specific cytotoxic lymphocyte (CTL) and liver function of patients with hepatitis B virus (HBV) genotype B and C infections and interleukin (IL)‐7 induced follicular helper T lymphocytes (Tfh) .Methods Sixty‐seven patients with chronic hepatitis B (CHB) hospitalized at Wuxi No .5 People′s Hospital from August 2013 to January 2015 were collected and 30 healthy blood donors were set as healthy control group .The peripheral blood IL‐7 , Tfh ,IL‐21 ,HBV specific‐CTL ,nonspecific CTL ,levels of HBV DNA ,alanine aminotransferase (ALT) and total bilirubin (TBil) were compared between patients with genotype B and C infection ,hepatitis B e antigen (HBeAg)‐positive and HBeAg‐negative CHB ,high ALT level and low ALT level .Multivariate regression analysis was performed to determine the factors associated with IL‐7 .The t test was used for quantitative data and chi‐square test was used for categorical data .Results Of the 67 CHB patients with average age of (35 .1 ± 11 .4) ,48 were male and 19 were female;32 were infected with genotype C and 35 were infected with genotype B ;40 were HBeAg‐positive CHB and 27 were HBeAg‐negative CHB ;17 were with high ALT levels and 50 were with low ALT levels .IL‐7 ,Tfh ,IL‐21 and HBV specific‐CTL levels in the peripheral blood of genotype C‐infected patients were (20 .79 ± 4 .82 ) ng/L , (3 .93 ± 0 .82)% ,(24 .77 ± 7 .52) ng/L and (0 .20 ± 0 .04)% ,respectively ,while in genotype B‐infected patients , those were (29 .13 ± 8 .17) ng/L ,(5 .92 ± 1 .92)% ,(39 .94 ± 24 .00) ng/L and (0 .40 ± 0 .06)% , respectively .Levels of IL‐7 , Tfh ,IL‐21 and HBV specific‐CTL in genotype C‐infected patients were significantly lower than those in genotype B‐infected patients (t= 5 .027 ,5 .595 ,3 .553 and 15 .133 , respectively ;all P<0 .01) .Nonspecific CTL ,HBV DNA ,ALT and TBil levels in the peripheral blood of genotype C‐infected patients were all significantly higher than those in genotype B infected‐patients (t=4 .899 ,6 .815 ,2 .763 and 4 .899 ,respectively ;all P< 0 .01) .IL‐7 ,Tfh ,IL‐21 ,HBV specific‐CTL levels in the peripheral blood of HBeAg‐positive patients were significantly lower than those in HBeAg‐negative patients (all P<0 .01) .Nonspecific CTL ,HBV DNA ,ALT and TBil levels in the peripheral blood of HBeAg‐positive patients were all significantly higher than those in HBeAg‐negative patients (all P<0 .05) .IL‐7 ,Tfh ,IL‐21 ,HBV specific‐CTL levels in the peripheral blood of patients with high ALT levels were all significantly lower than those in patients with low ALT levels (all P<0 .01) .Nonspecific CTL and HBV DNA levels in the peripheral blood of patients with high ALT levels were both significantly higher than those in patients with low ALT levels (both P<0 .01) .HBV DNA ,IL‐21 and nonspecific CTL were all correlated with IL‐7 (all P<0 .01) .Conclusion The differences of HBV specific‐CTL and liver function in CHB patients infected with genotype B and C may be correlated with interleukin‐7 induced Tfhcells.

Objective To explore the relationship between viral load and expression of CD 226 on the surface of peripheral blood T follicular helper cells (Tfh) in patients with chronic hepatitis C (CHC) . Methods One hundred and thirty-five CHC patients hospitalized at Wuxi Fifth People′s Hospital from March 2015 to April 2017 were collected ,and another 30 healthy blood donors were set as healthy control group .CHC patients were divided into two groups based on hepatitis C virus (HCV) RNA level ,with 49 cases (36 .3％ ) in low viral load group and 86 cases (63 .7％ ) in high viral load group .Expression of CD226 on the surface of peripheral blood Tfh cells , Tfh cells ,interleukin (IL )-21 and HCV specific cytotoxic lymphocyte (CTL) levels of two patient groups were compared .Categorical data were compared with chi-square test and normally distribute continuous data were compared with t test .Correlations between different factors were analyzed by Pearson correlation analysis .Results Expression of CD226 on the surface of peripheral blood Tfh cells in 135 cases of CHC patients was (77 .69 ± 5 .42)％ ,which was lower than that of healthy control ([90 .06 ± 5 .83]％ ) ,and the difference between the two groups was significant (t= 7 .541 , P < 0 .01) .The CD226 expression on the peripheral blood Tfh cells in low viral load group was (88 .75 ± 6 .68)％ ,which was higher than that of high viral load group ([69 .23 ± 5 .86]％ ) ,and the difference between the two groups was significant (t = 19 .232 , P< 0 .01) .The viral load was negatively correlated with Tfh cell surface CD 226 expression (r = - 0 .705 , P < 0 .01) .The peripheral blood Tfh cell level in 135 CHC patients was higher than that of healthy control ,and the difference between the two groups was significant (t= 13 .878 , P< 0 .01) .The peripheral blood Tfh cell level in low viral load group was higher than that in high viral load group ,and the difference between the two groups was significant (t= 26 .993 , P< 0 .01) .The IL-21 level of 135 CHC patients was lower than that of healthy control ([70 .35 ± 1 .6]％ ) ,and the difference between the two groups was significant (t=18 .322 , P< 0 .01) .The IL-21 level in peripheral blood of low viral load group was higher than that of high viral load group ,and the difference between the two groups was significant (t= 84 .54 , P< 0 .01) . HCV specific CTL level in peripheral blood of low viral load group was higher than that of high viral load group ,and the difference between the two groups was significant (t = 29 .869 , P< 0 .01) .The viral load was negatively correlated with levels of HCV specific CTL (r= -0 .734 ,P< 0 .01) .Conclusions In patients with chronic hepatitis C ,different levels of viral load can result in different levels of CD 226 expression on the peripheral blood Tfh cells .Patients with low viral load has high CD226 expression on Tfh cell surface , resulting in rise of Tfh cell level ,IL-21 level and HCV specific CTL level .