Objective To construct DNA vaccine expressing HIV-1 AE2f gp145-tat-rev-nef fusion gene( AE-Gp145TRN) and to compare the immunogenicities of DNA vaccines expressing Tat, Rev and Nef in gene fusion formulations of tat-rev-integrase(c-half)-vif-nef( AE-TRIVN) and AE-Gpl45TRN. Methods DNA vaccine was constructed by inserting the codon optimized HIV-1 AE2( gp145-tat-rev-nef fusion gene into mammalian expression DNA vector. In vitro expression efficiency of the constructed DNA vaccine was determined by Western blot and the immunogenicities of AE-Gpl45TRN and AE-TRIVN were compared by immunizing female BALB/c mice. IFN-r ELISPOT assay was used to read out the specific T cell immunity. Results Western blot assay showed the constructed DNA vaccine could be expressed efficiently in vitro. After vaccination, AE-TRIVN mounted significantly higher T cell responses against Tat, Rev and Nef[(148±91)SFCs/106 splenocytes]than Gpl45TRN[(55±28) SFCs/106 splenocytes]. Specific T cell responses elicited by AE-TRIVN predominantly targeting Rev, whereas Gpl45TRN could significantly enhance T cell responses against Nef. Conclusion AE-TRIVN and Gpl45TRN induced distinct T cell response modalities, which implied different gene fusion formulations may affect the immunogenicity of specific DNA vaccines.

Objective To investigate the clinical value of mean channel of neutrophil volume (MNV), mean channel of neutrophil conductivity (MNC) and mean channel of neutrophil scatter (MNS) in predicting acute bacterial infection.Methods Peripheral blood samples from 112 patients with positive blood cultures for bacteria,70 healthy subjects and 45 non-infectious subjects with high white blood cell count(WBC) were studied using the Coulter LH 750 hematology analyzer.MNV, MNC, MNS and neutrophil volume distribution width (NDW) were retrospectively analyzed and compared with total WBC, percentage of neutrophils,neutrophil left-shift and CRP.112 blood bacterial infections were grouped according to WBC count (A:WBC ＜11.0×109 /L;B:11.0×109/L≤WBC＜15.0×109 /L;C:WBC≥15.0×109 /L) and neutrophil rate (NE ＜ 0.85 and NE ≥ 0.85 ).Results MNV and NDW increased significantly in septic patients (154.17 ± 10.08,24.36 ± 4.14 ) compared with those of healthy control group (142.09 ± 4.13,19.04 ± 1.97) and non-infectious patients with high WBC group ( 150.63 ± 8.14,20.19 ± 4.73 ).There was statistically significant difference (F value were 20.738 and 28.190 respectively,P ＜ 0.01 ). On the contrary, MNS decreased significantly in septic patients (137.15 ± 7.61 ) compared with that of healthy group (144.51±4.36) and nonspetic patients with high WBC group (142.45±7.11) ,there was significant statistical difference (F=5.217,P＜0.01).The MNV, NDW and MNS of A group were 148.09±5.76,22.39±1.97,140.07±6.11 respectively.The MNV, NDW and MNS of B group were 152.83±5.75,24.14±1.35,141.44±5.35 respectively.The MNV, NDW and MNS of C group were 164.28±6.49,29.42±5.93,134.27±9.61 respectively. There was statistically significant difference compared with healthy group (F value were 24.720,31.642,7.931, P ＜ 0.01).The MNV, NDW and MNS in the group with NE ＜0.85 were 149.17±9.06,22.59±2.73,141.19±4.34 respectively.The MNV, NDW and MNS in the group with NE≥0.85 group were 159.03±10.23,27.64±4.51,135.62 ± 8.95 respectively.There was statistically significant difference compared with healthy group ( F value was 23.970,51.309,19.792,P＜0.01).With a cut-off of 150 for the MNV, a specificity of 90% and sensitivity of 70% were achieved.NDW was associated with neutrophil left-shift (r=0.33,P＜0.01).With a cut-off of 23 for the NDW, a specificity of 100% and a sensitivity of 72% were achieved.The sensitivity of the MNV and NDW was better than total white blood cells count (with a cut-off≥ 11.0×109 /L, the sensitivity was 57% ), percentage of neutrophils( with a cut-off≥0.85, the sensitivity was 44% ) and neutrophil shift to left ( with a cut-off ＞5%, the sensitivity was 66% ) and CRP (with a cut-off ≥10 mg/L, the sensitivity was 65% ).Conclusions The MNV and NDW of the neutrophil can reflect the morphologic change of neutrophil sensitively and specificialy in acute infection. As quantitative, objective and more sensitive parameters, MNV and NDW may have a potential role for predicting the acute bacterial infection.

Objective To compare the 2-year efficacy of de novo combination therapy with lamivudine (LAM) and adefovir dipivoxil (ADV) to that of entecavir (ETV) monotherapy in treatment of patients with hepatitis B virus ( HBV )-related decompensated cirrhosis.Methods A total of 120 naive patients with HBV-related decompensated cirrhosis admitted to Shangyu People's Hospital and the First Affiliated Hospital of Zhejiang University from January 2007 to April 2008 were enrolled,in which 60 were treated with LAM and ADV combination therapy,and other 60 patients were treated with ETV monotherapy.Tests for liver and kidney function,alpha-fetoprotein,HBV serum markers,HBV DNA load,prothrombin time (PT),and ultrasonography or CT scan of liver were performed every 1-3 months.Repeated measure ANOVA and x2test were used to compare the efficacy,side effects and accumulated survival rates at 12 and 24 month in two groups.Results Forty-five patients in each group were followed-up for 24 months.There was no significant difference in HBV DNA negative rates and ALT normalization rates at month 12 (x2 =2.12 and 2.88,P ＞0.05 ) and month 24 between two groups (x2 =3.21 and 3.24,P ＞ 0.05); while HBeAg seroconversion rate in LAM + ADV group at month 24 was significantly higher than that in ETV group (43.5％ vs.36.4％,x2 =4.09,P＜0.05).Viral breakthrough occurred in 2 cases (4.4％) by month 12 and 3 cases (6.7％) by month 24 in LAM + ADV group,and no viral mutation was observed; while in ETV group,viral breakthrough occurred in 1 case ( 2.2％ ) by month 12 and 2 cases (4.4％) by month 24,and viral mutation was observed in 1 case (2.2％) by month 24.At the end of month 24,increase of AIb (F=18.9 and 17.3,P＜0.05),decrease of TBil and ALT (F=16.5,17.1 and 23.7,24.8,P ＜0.05 ),shortening of PT ( F =22.7 and 24.5,P ＜ 0.05 ),and the improvements of CTP and MELD scores (F=18.5,17.8 and 24.2,23.8,P＜0.05) were observed in both groups.The accumulative rates of mortality or liver transplantation were 16.7％ ( 10/60 ) and 18.3％ ( 11/60 ) in LAM + ADV and ETV groups,respectively.No blood creatinine increased above the normal upper limit was observed in both groups.Conclusion Both LAM + ADV combination therapy and ETV monotherapy can effectively inhibit HBV replication,improve liver function,decrease mortality and viral resistance,but the 24-month HBeAg seroconversion rate in combination therapy group is higher than that in monotherapy group.

Objective To analyze CD127 expression on the memory CD8+ lymphocytes from hepatitis B e antigen (HBeAg) positive chronic hepatitis B (CHB) patients treated with peginterferon α-2a (Pegasys). Methods Thirty HBeAg positive CHB patients were treated with peginterferon α-2a 180 μg once a week for 48 weeks and followed up for 24 weeks. The memory CD8+ lymphocytes were characterized by expressing CD45RA and CD27 markers. CD127 expression on cell surface was measured by four-colour flow cytometry. The difference of mean values between groups was evaluated by Mann-Whitney test. Results The CD127 expression on CD8+ T lymphocytes was significantly lower in HBeAg positive CHB patients compared to healthy controls (Z=2.889, P＜0.05), which was negatively correlated with serum hepatitis B virus (HBV) DNA level and HBeAg titers. The CD127 expression increased along with the decrease of HBV DNA and HBeAg after 24-week, 48-week and 72-week treatment in patients showing good response to peginterferon α-2a, while CD127 expression didn't change markedly in non responders (Z24w = 1.954, Z48w = 2.789, Z72w = 2. 989; all P＜0. 05). Conclusion CD127 expression on memory CD8+ lymphocytes increases along with effective anti-HBV treatment in CHB patients, which can be used as a marker for evaluating the effectiveness of anti-viral treatment.

Objective To compare the efficacy of add-on adefovir dipivoxil (ADV) therapy and switch-to entecavir (ETV) monotherapy in chronic hepatitis B (CHB) patients with suboptimal response to lamivudine (LAM).Methods A prospective study was performed in 120 CHB patients from Zhuji People' s Hospital and the First Affiliated Hospital of Zhejiang University School of Medicine during June 2010 and June 2011.All patients previously received more than 24 weeks LAM treatment,but HBV DNA was still positive.Patients were randomized assigned to two groups:60 patients received add-on ADV therapy and another 60 switched to ETV monotherapy.Both groups were treated for 48 weeks.Liver and kidney function,alpha-fetal protein (AFP),HBV serum markers,HBV DNA and prothrombin time (PT) were examined,and ultrasonography or CT scan of liver was performed every 1-3 months.x2 test was used to compare the HBV DNA negative rates,HBeAg seroconversion rates,resistance rates and adverse reaction at week 48 between two groups.Results Thirty-three out of 38 patients (86.8％) with baseline HBV DNA 103-105 copies/mL became HBV DNA negative after add-on ADV treatment for 48 weeks,twenty-seven out of 39 patients (69.2％) with baseline HBV DNA 103-105 copies/ml became HBV DNA negative after switch-to ETV treatment.There was a statistical difference between two groups (x2 =4.578,P ＜ 0.05).Sixteen out of 22 patients (72.7％) with baseline HBV DNA ＞ 105 copies/mL became HBV DNA negative after add-on ADV treatment for 48 weeks,while only 52.4％ (11/21) patients achieved HBV DNA negative in the switch-to ETV group.There was also a statistical difference between two groups (x2 =4.865,P ＜0.05).None of patients in add-on group developed virological breakthrough and resistance,while 5 patients in switch-to ETV group developed virogical breakthrough and 3 patients developed genetic mutation.Among them,rtM204V + rtL180M + rtS202G mutation was detected in 2 patients,and rtM204V + rtL180M +rtT184A mutation was detected in 1 patient; all mutations happened in the baseline HBV DNA ＞ 105 copies/mL group.Conclusion The add-on ADV therapy is better in viral inhibition than switch-to ETV therapy for CHB patients with suboptimal response to LAM,and it can reduce the occurrence of drug resistance.