Hepatitis B, Chronic ; Humans

Liver sinusoid endothelial cells are the first defense mechanism that protects the liver against various injuries,and they also play a significant role in the development of liver fibrosis and liver cirrhosis caused by chronic liver injury.They are involved in liver fibrosis by mediating liver inflammation,participating in sinusoid capillarization and revascularization,activating hepatic stellate cells,secreting many proinflammatory cytokines,participating in extracellular matrix formation,and mediating liver microcirculation disturbance.Clarification of these mechanisms helps to identify new targets and develop new regimens for the treatment of liver fibrosis.

ObjectiveTo investigate the effect of pravastatin on serum levels of interleukin-6(IL-6) and tu- mor necrosis faetor-α(TNF-α) in patients with acute coronary syndromes(ACS). Methods50 patients with ACSwere randomly divided into pravastatin group( n = 25) and routine therapy group( n = 25). Serum TNF-α and IL-6levels were measured before and four weeks after the two treatment options respectively. ResultsThe level of TNF- α and IL-6 were higher than routine therapy group before therapy(P < 0.01 ). The level of TNF-α and IL-6 weredecreased significantly after pravastatin therapy,and higher thancontrols(P < 0.05). The levels of TNF-α and IL-6changed only slightly after the routine therapy(P > 0.05). ConclusionThe level of TNF-α and IL-6 becominghigher in ACS patients may be related to the pathogenesis of ACS. Pravastatin can reduce serum level of TNF-α and IL-6 contributing to treatment of ACS.

Objective: To study the effects of soluble multi-peptide agent,marked “Gu Kang Tai Ling”,derived from bone tissue on rat bone fracture healing.Methods 80 right tibia defects of rats were sawed.The “Gu Kang Tai Ling”were intramuscularly injected to the rats.The quality of the defect healing was investigated continuously and respectively by the defect bone histomorphometry,biomechanics,X-ray film,and bone mineral density(BMD).Results: Under the treatment of the agent,at early stage trabecular bone volume (TBV)ratio increased 24.7%,and osteoblast surface(OBS)26.7%,at late stage mean trabecular healing surface rate(MTHSR)was increased 38.8%,tibia anti-fracture strength 0.4527～1.4350N，and BMD 7.65% compared with the control group.Conclusion: “Gu Kang Tai Ling”can accelerate rat fracture healing and improve the quality of healing.

Background and purpose:Regular physical exam became the main means to discover the population with early stage RCC. So far, there have been no reports with large sample that have been published about the biomarkers as predictor for the metastasis of the postoperative patients with stage T_(1-3)N_0M_0 (RCC) in China. Furthermore, the underlying mechanism of metastasis of RCC is not clear. This research was carried out in order to study the correlation between delayed metastasis of early stage renal cell carcinoma after surgical operation and expressions of KAI-1、Ki-67 and HER-2/neu on RCC. Methods:Two hundred and forty-one patients with RCC underwent surgical operations and were pathologically diagnosed as T_(1-3)N_0M_0 stage. Twenty-four patients were found to have metastases after long-term follow-up and their clinical data were reviewed (metastasis group). One hundred and ninety-four patients without postoperative metastases were taken as control group. Twenty three patients were excluded from this study because they were lost to follow-up. The expressions of KAI1、 Ki-67 and HER-2/neu in the samples of the two groups were tested with immunohistochemical staining by PowerVision or EnVision two-step procedure. Significant difference was calculated with Chi square test.Results:The positive expression rates of KAI1、HER-2/neu and Ki-67 in 218 RCC were 82.6%、27.5% and 83.5%, respectively. Both mild (20.8%) and strong positive rates (4.2%) of KAI1 in metastasis group were dramatically less than that in control group (90.2%, 73.7%, P

Objective To investigate the characteristics of HBV replication in HepG2.2. 15 cells treated with LAM alone or sequentially treated with LAM and IFN-α, and to further explore the different suppressive effect on HBV replication by LAM and IFN-α in vitro. Methods Untreated HepG2. 2. 15 cells were used as control group, HepG2. 2. 15 cells treated with 1 000 IU/ml of IFN-α alone for 10 d were served as IFN-α group. The HepG2.2. 15 cells treated with 0.2,1,5,20,100 μmol/L of LAM were used as LAM groups. HepG2. 2. 15 cells treated with 0. 04,0. 2,5,25,125,200 μmol/L of LAM for 7 d, then combined with 1 000 IU/ml of IFN-α for another 3 d. Afterwards, the cells were treated with 1 000 IU/ml of IFN-α only for another 10 d. These cells were served as LAM/IFN-α sequential treatment group. The ELISA was used for analyzing the secreted HBV antigens, while the Dot blot, Southern blot were applied for analyzing the extracellular HBV DNA and intracellular HBV replicative intermediate DNA in HepG2. 2. 15 cells of different treatment groups. Results The secreted HBsAg in the LAM group were 1. 77 ± 0. 22, 1.65 ±0.25, 1.95 ±0. 19, 1.34 ±0. 11, 1.07 ±0.05, respectively, and the secretion of HBeAg were 1.41 ±0. 13, 1.37 ± 0. 09, 1.63 ± 0. 07, 1.26 ± 0. 12, 1.05 ± 0. 09. The secreted HBsAg and HBeAg in control group were 3. 34 ± 0. 15 and 3.33 ± 0. 05. Statistical analysis showed that HBsAg and HBeAg secretion in the LAM group were significantly reduced by treatment with LAM. The t values of HBsAg were 10. 21,10.04, 9.94, 18.62, 24.86, and the t values of HBeAg were 23.87, 32.97, 34.22, 27.57, 38.35,respectively, all P values were ＜ 0.05. Dot blot, Northern blot hybridization analysis indicated that the extracellular HBV DNA and intracellular HBV replicative intermediate DNA could not be detected in the LAM group after cells treated by LAM for 10 days. When LAM was replaced with treatment of 1 000 IU/ml of IFN-α alone, it could not suppress the HBV replication effectively. Moreover, the intracellular HBV replicative intermediate DNA still existed in almost all groups, accompanied with the recovered expression of HBV antigens as well as extracellular HBV DNA, which suggested that the HBV particles restored replication again and secreted extracellular in HepG2. 2. 15 cells, although the sequential treatment lasted for 10 days.Conclusion The effect of viral suppression by LAM and IFN-α in vitro were different, which attributed to the different HBV replicative characteristcs in HepG2. 2. 15 cells.

Objective:To prepare and test tetrameric sH-2K~d-HBc complex for the further measurement of the specific CTL response.Methods:PE labled streptavidin with 4 biotinylated binding sites can bind to 4 biotinylated monomer to form the corresponding tetramer.Mice were immunized via different methods of genetic immunization by use of the construted pcDNA3-C plasmid to get the specific CTLs.Then our prepared tetramer was applied to stain the specific CTLs by the analysis of flow cytometry.Results:We applied our prepared tetramer to stain the cells from the experimental groups and control group.The results showed the tetramer was able to discriminate the frequencies of specific CTL induced by the three immunol methods(0.24%,0.26%,0.36% vs 0.07%,P≤0.05).This demonstrated that the prepared tetramer could bind its targets specifically and efficiently.The three immunol methods induced different levels of immune responses.Compared with the traditional muscle injection,gene gun induced weaker humoral immune response and stronger cellular immune response,and hydrodynamic injection induced the strongest humoral and cellular immune responses.Conclusion:Have successfully constructed the sH-2K~d-HBc tetramer.The techniques and methods can be used for preparation of tetramers of other types of MHCⅠ molecules.

Objective:To clarify the differences of host immune responses at different stages of HBV infection. Methods:We constructed three HLA-A*0201/HBV tetramers with immunodominant epitopes of core18-27,polymerase 575-583 and envelope 335-343,and analyzed antigen specific CD8+ T cells and the expression of CD127 in peripheral blood mononuclear cells ( PBMCs) from patients infected with HBV using these HLA-A*0201/HBV tetramers. Results: The frequencies and expansion ability of antigen specific CD8+ T cells in most self-limited HBV infected individuals were higher than that in chronically HBV infected patients. In low copy period the frequencies of antigen specific CD8+ T cells were similar to those in immune clearance phase at a high viral load and liver damage and in immune clearance phase, which had no significant correlation with virus quantitation and ALT level. In chronic infection the ability of antigen specific CD8+ T cells proliferation was inversely proportional to the viral titer. In most self-limited HBV infected individuals the IFN-γsecretion functions of antigen specific CD8+ T cells were higher than in chronic infection,but in immune tolerance phase these cells lost the ability. HBsAg level was different at different stages after HBV infection:it was highest in immune tolerance phase,but in immune clearance phase,activity period and low copy period the correlation with HBV DNA replication gradually declined. The frequency of CD8+ CD127+ T cells in chronic HBV infection was lower than the control group and self-limited infection group,especially in immune tolerance with HBeAg+ and immune clearance phase. Conclusion: The frequencies of antigen specific CD8+ T cells are not the main determinant of immune-mediated protection in chronic HBV infection,memory antigen specific CD8+ T cells are not clear or missing,which provides the possibility for therapeutic vaccines and immunization therapy.

In order to establish stable high expression cell lines, the eukaryotic expression vector pIRES2EGFP and recombinant plasmid pIRES2EGFP-TIM-3 were transfected into mammalian cells CHO by Lipofectamine. The transfected cells were cultivated under selective growth medium including G418 and green fluorescent protein (GFP) positive cells were sorted by FACS. Simultaneously, growing transfectants were selected only by G418 in the medium. The GFP expression in stably transfected cells was detected by FACS. Under selective growth conditions with G418, the percentage of GFP positive cells was reduced rapidly and GFP induction was low. In contrast, the percentages of GFP positive cells were increased gradually after FACS. By 3 rounds of GFP selection, the stable high expression cell lines were established. Furthermore, using FACS analysis GFP and the target protein TIM-3 co-expression in the stable transfectants cultured in nonselective medium was detected. Theses results demonstrated that the stably transfected cell lines that express high titer of recombinant protein can be simply and fleetly obtained by using GFP and selective growth medium.

To observe the expression of CD40/CD40L on peripheral blood mononuclear cells (PBMC) in patients with condyloma acuminatum (CA), flow cytometry was employed to examine the expression of CD40 and CD40L on PMBC in 36 patients with CA and 20 healthy controls. Our results showed that mean level of CD40 expression in CA patients was significantly lower than that in the controls (6.58% +/- 2.74% vs 14.81% +/- 6.12%, t = 5.703, P < 0.05); the average level of CD40L in CA patients was also significantly lower than that in the controls (0.73% +/- 0.54% vs 2.67% +/- 2.43%, t = 3.532, P < 0.05). Our results suggest that the reduced costimulatory interaction of CD40 and CD40L in CA patients may be one of the important factors responsible for the low cellular immunity.