Objective To observe the therapeutic effects of recombinant human granulocyte colony stimulating factor(rhG-CSF)on CCl4 induced chronic liver injury.Methods Male BALB/C mice were randomly allocated into treatment and control groups.The mice model were established by injection with daily for 7 days,while the control mice were received the same volumes of saline.The mice were sacrificed to get weight,liver mass and spleen mass.The count of CD34+ cells and Thy-1+ cells were analyzed by flow cytometry and immunohistochemical staining,respectively.Results The ratio of liver/spleen was 15.94±1.20 and 10.52±0.66 on day 8 and 15 in treatment group,respectively,while those were 7.14±1.68 and 8.31±1.71 in control group,respectively(all P value＜0.05).But there was no significant difference in body weight and liver mass between two groups(P＞0.05)The concentration of album in treatment group was raised rapidly on day 15.The concentrations of alanine aminotransferase (ALT),aspartate aminotransferase(AST),hyaluronic acid(HA)and laminin(LN)on day 30 were significantly lower in treatment group compared to control group(P＜0.05).There was significant difference in score of liver fibrosis on day 30 between two groups(treatment group:5.49±2.16,control:8.74±1.86,P＜0.05).The number of CD34+ cell and Thy-1+ in treatment group(on day 8:9.54±2.24 and 5.10±1.25 and on day 15:8.18±1.93 and 7.53±1.39,respectively)were higher than those in control group(on day 8:5.40±0.99 and 3.25±0.75;on 15 days:4.46±0.77 and 3.35±0.86,all P value＜0.05).Conclusion The rhG-CSF may improve the reparation of chronic liver injury,and may provide a novel method in treatment of liver fibrosis.

CD4+CD25+ Regulatory T cells (Treg) have been found to down-regulate immune activation in HIV-1 infection. However, whether the depletion of Treg benefits to the disease status of HIV infection remains undefined. To address this issue, we enumerated the Treg absolute counts and frequency in 75 antiviral-na(i)ve HIV-1-infected individuals in this study. It was found that HIV-infected patients displayed a significant decline in Treg absolute counts but a significant increase in Treg frequency. In addition, with disease progression indicated by CD4 T-cell absolute counts, circulating Treg frequency gradually increased; while Treg absolute counts were gradually decreased, suggesting that the alteration of Treg number closely correlated with disease progression in HIV infection.Functional analysis further showed that Treg efficiently inhibit both CD4 and CD8 T cell proliferation in vitro. Thus, our findings indicates that Treg actively participate in pathogenesis of chronic HIV infection,influencing the disease progression.

ObjectiveTo investigate the correlation of serum high-mobility group box 1 (HMGB1) level with hepatic inflammatory activity, liver fibrosis degree, and liver function parameters in chronic hepatitis B patients. MethodsA total of 73 patients with chronic hepatitis B and liver cirrhosis were enrolled as subjects. Liver biopsy was performed to determine inflammatory activity and liver fibrosis degree, liver function parameters and serum HMGB1 level were measured, and the correlation between HMGB1 level and other indices was analyzed. The t-test was used for comparison between two groups, and a linear correlation analysis was performed to investigate the correlation between two indices. ResultsThe patients with chronic hepatitis B and liver cirrhosis had a significantly higher HMGB1 level than the healthy control group (29.46±7.54 ng/ml vs 16.86±3.48 ng/ml, t=5.668, P＜0.01). The G3-G4 group had a significantly higher HMGB1 level than the G1-G2 group (t=4.441, P＜0.01), while there was no significant difference in HMGB1 level between the S1-S2 group and the S3-S4 group (t=0.658, P＞0.05). Serum HMGB1 level was positively correlated with alanine aminotransferase (r=0.256 6, P=0028 4) and aspartate aminotransferase (r=0.471 9, P＜0.000 1), while it was not correlated with albumin or total bilirubin (both P＞005). ConclusionSerum HMGB1 level is closely correlated with hepatic inflammatory activity.

Objective To investigate whether the co-culture of Huh7.93 cells and peripheral blood mononucleated cells from chronic hepatitis B patients (cPBMCs) can simulate the replication features of hepatitis B virus (HBV) and immune function in chronic hepatitis B (CHB) patients,and to provide an in vitro cell co culture system for the research on immune clearance in chronic HBV infection.Methods Huh7.93 cells were cultured alone or co-cultured with peripheral blood mononucleated cells from healthy people who underwent physical examination (nPBMCs) or cPBMCs for 7 days.The CCK8 assay was performed to measure the proliferative activity of Huh7.93 cells,and quantitative real-time PCR and Southern blot were used to measure HBV replication in cPBMCs and co-cultured cells.The independent samples t-test was used for comparison between two groups.Results When Huh7.93 cells were co-cultured with peripheral blood mononucleated cells (PBMCs) at a certain ratio,Huh7.93 cells had a high proliferative activity and good cell growth.HBV could not infect or replicate in cPBMCs.HBV DNA in the supematant of Huh7.93 cells co-cultured with cPBMCs showed significant increases and significantly higher than that in the supernatant of Huh7.93 cells cultured alone on day 4 (6.01 ± 0.20 log10copies/ml vs 4.99 ± 0.08 lOg10copies/ml,P =0.000) and day 7 (7.82 ± 0.24 log10copies/ml vs 6.96±0.09 log10copies/ml,P =0.000).On day 7 of culture,the cell lysis buffer of Huh7.93 cells co-cultured with cPBMCs had a significant increase in the level of HBV replicative intermediate compared with that of Huh7.93 cells cultured alone.After HepG2.2.15 cells were co-cultured with cPBMCs,there was no significant increase in HBV replication.Conclusion The co-culture of Huh7.93 cells and cPBMCs produces similar viral replication as human body infected with HBV and can well simulate the liver microenvironnent and immune function in CHB patients,which provides a new method for the research on immune clearance in chronic HBV infection.

Objective: To investigate the curative effect of local application of CpG-oligodeoxynucleotide (CpG-ODN) combined with 4-1BB monoclonal antibody in hepatoma-bearing mice, and to evaluate the effect of 4-1BB monoclonal antibody on CpG-ODN immunotherapy. Methods: H22 single cell suspension was injected subcutaneously into the axilla and four limbs of the BALB/c male mice to establish a tumor-bearing mice model. After 7 days, 30 mice with corresponding tumor-bearing volume were screened and randomly divided into model control group, CpG group and CpG+4-1BB group, and the drug was injected into the tumors of left lower extremity. The same batch of normal mice was selected as normal control group. Survival of mice was recorded. Tumor-bearing volume and organ index were calculated. Serum levels of interleukin (IL) - 12 and interferon (IFN) gamma and spleen CD8⁺T lymphocyte ratio were measured. The measurement data were analyzed by analysis of variance. The survival rate of each group of mice was analyzed by log-rank test. Results: Mice in the model control group with tumor-bearing volume had a sustained growth before the execution. CpG group and the CpG+4-1BB group [(976.08 ± 29.55) mm³, (47.25 ± 0.93) mm³)] tumor-bearing volume was decreased than model group [(1 336.52 ± 39.40) mm3] (F = 5 329.273, P < 0.05). CpG+4-1BB group distant tumor-bearing volume [(611.83 ± 113.02) mm3] was decreased than model group and CpG group [(1 406.62 ± 51.09) mm³, (1 380.01 ± 51.44) mm3] (F = 247.160, P < 0.05), but there was no significant difference between the CpG group and the model group (P > 0.05). Serum IL-12 concentration (23.90 ± 2.33 pg/ml), IFN-γ concentration (103.02 ± 6.10 pg/ml) and spleen CD8⁺T cell ratio (4.54 ± 0.62%) in the model group were lower than those in the normal group (P < 0.05). Serum IL-12 concentration in CpG group and CpG+4-1BB group (29.21 ± 2.23 pg/ml, 37.04 ± 1.49 pg/ml), IFN-γ concentration (116.12 ± 4.08 pg/ml, 138.65 ± 1.72 pg/ml), CD8⁺T cell ratio (6.65 ± 0.64%, 12.73 ± 0.88%) were higher than the model group, while CpG+4-1BB group was higher than the CpG group (P < 0.05). The survival rate of CpG+4-1BB group was higher than that of model group and CpG group (χ² = 25.544, P < 0.05), but there was no significant difference between CpG group and model group (P > 0.05). There was no significant difference in organ index between the four groups (P > 0.05). Conclusion 4-1BB monoclonal antibody combined with CpG-ODN therapy can shrink hepatoma-bearing capacity, inhibit the growth of distant tumors and significantly prolong the survival time of mice.