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Construction and characterization of an experimental ISCOMS-based hepatitis B polypeptide vaccine

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World Journal of Gastroenterology 2002年第2期8卷 294-297页

作者： Xiao-Ju Guan Shanghai Institute of Materia Medica,Chinese Academy of Sciences,Shanghai 200031,China,previously worked as a postdoc in Institute of Immunology,Third Military Medical University,Chongqing 400038,China Xiao-Jun Guan,Department of Science & Research,Second Military Medical University,Shanghai 200433,China Yu-Zhang Wu Zheng-Cai Jia Tong-Dong Shi Yan Tan,Institute of Immunology,Third Military Medical University,Chongqing 400038,China Shanghai Institute of Materia Medica Chinese Academy of Sciences Shanghai 200031 China.

AIM: To characterize the biochemical and immunological properties of an experimental ISCOMS vaccine prepared from a novel therapeutic polypeptide based on T cell epitopes of HBsAg, and a heptatis B-ISCOMS was prepared and investigated. METHODS: An immunostimulating complexes(ISCOMS)-based vaccine containing a novel therapeutic hepatitis B polypeptide was prepared by dialysis method, and its formation was visualized by electron microscopy and biochemically verified by SDS-polyacrylamide gel electrophoresis. Amount of the peptide within ISCOMS was determined by Bradford assay, and specific CTL response was detected by ELISPOT assay. RESULTS: Typical cage-like structures of submicroparticle with a diameter of about 40nm were observed by electron microscopy. Results from Bradford assay showed that the level of peptide incorporation was about 0.33g.L(-1). At the paralleled position close to the sixth band of the molecular weight marker(3480kDa) a clear band was shown in SDS-PAGE analysis, indicating successful incorporation of polypeptide into ISCOMS. It is suggested that ISCOMS delivery system could efficiently improve the immunogenicity of polypeptide and elicit specific immune responses in vivo by the results of ELISPOT assay, which showed that IFN-gamma producing cells(specific CTL responses) were increased(spots of ISCOMS-treated group: 47+/-5, n =3; control group: 5+/-2, n =3). CONCLUSION: ISCOMS-based hepatitis B polypeptide vaccine is successfully constructed and it induces a higher CTL response compared with short polypeptides vaccine in vivo.

关键词： Hepatitis B Vaccines ISCOMs Animals Enzyme-Linked Immunosorbent Assay Epitopes Female Hepatitis B Surface Antigens Humans interferon type ii Mice Mice, Inbred BALB C Peptides Research Support, Non-U.S. Gov't T-Lymphocytes, Cytotoxic

T helper cells in patients with chronic hepatitis B virus infection

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Chinese Medical Journal 2002年第3期115卷 422-424页

作者：姜荣龙冯筱榕郭亚兵卢桥生侯金林骆抗先富宁第一军医大学南方医院感染内科广州510515

OBJECTIVE: To investigate the compositions of Th1/Th2/Th3 cells in chronic hepatitis B virus (HBV)-infected individuals by determining the expression of interleukin-4 (IL-4), inetrferon-gamma (IFN-gamma), and transform growth factor-beta (TGF-beta) in single CD4(+) T cells isolated from peripheral blood mononuclear cells (PBMCs) and the role of polarized Th cell populations in chronic HBV-infection was discussed. METHODS: PBMCs from chronically infected HBV individuals were isolated, stimulated by PMA/Ionomycin/Monensin, and IL-4, IFN-gamma and TGF-beta production by CD4(+) T cells was determined by using fluorescence activated cell sorter (FACS) analysis. RESULTS: The percentage of IFN-gamma-producing T cells, IL-4-producing T cells and TGF-beta-producing T cells ranged from 2.3% - 18.6%, 1.1% - 8.7% and 0.7% - 7.1% respectively in CD4(+) T cells from non-infected individuals. Most of CD4(+) T cells from PBMCs in chronically infected HBV individuals were Th0 cells. The proportion of Th1 cells increased significantly with hepatic inflammatory activity, and in the active period of chronic hepatitis B infection were higher than those in the non-active period (P 0.05), but were higher than that from controls (P

关键词： CD4-Positive T-Lymphocytes Hepatitis B, Chronic Humans interferon type ii Interleukin-4 T-Lymphocytes, Helper-Inducer Th1 Cells Th2 Cells Transforming Growth Factor beta

Humoral and cellular immunogenecity of DNA vaccine based on hepatitis B core gene in rhesus monkeys

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World Journal of Gastroenterology 2001年第1期7卷 102-106页

作者： Zu Hu Huang1 Hui Zhuang2 Shan Lu3 Ren Hua Guo1 Guo Min Xu2 Jie Cai1 Wan Fu Zhu2 1Department of Infectious Diseases. The First Affiliated Hospital of Nanjing Medical University, Nenjing 210029, Jiangsu Province. China2Faculty of Microbiology, Beijing University, Beijing 100000, China3University of Massachusetts Medical Center The First Affiliated Hospital of Nanjing Medical University 300 Guangzhou Road Nanjing 210029 Jiangsu Province China.

INTRODUCTIONHepatitis B virus (HBV) is the most commonetiologic agent for infectious liver diseases. It isestimated that there are more than 250 millionchronic HBV carriersin the world today and thereis a significant ... 详细信息

关键词： Vaccines, DNA Animals Antibodies, Viral Antibody Formation Antibody Specificity Cell Division Cells, Cultured Enzyme-Linked Immunosorbent Assay Female Hepatitis B control Hepatitis B Core Antigens Immunity, Cellular Immunoglobulin G interferon type ii Interleukin-4 Leukocytes, Mononuclear Macaca mulatta Male Research Support, Non-U.S. Gov't

Effects of aminoguanidine on nitric oxide production induced by inflammatory cytokines and endotoxin in cultured rat hepatocytes

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World Journal of Gastroenterology 2001年第3期7卷 331-334页

作者： Guo Liang Zhang Ye Hong Wang Hui Ling Teng Zhi Bin Lin Department of Pharmacology,School of Basic Medical Sciences,Beijing University,Beijiog 100083,ChinaDr.Guo Liang Zhang graduated from Xinxiang Medical College in 1982,got Ph.D.at Nagoya City University Medical School,Japan in 1994,finished postdoctoral research at Beijing Medical Univcrsity in 1996,now an associate professor of pharmacology,specialized in hepatic pharmacology,having 15 papers published. Department of Pharmacology School of Basic Medical Sciences Beijing University Beijing 100083 China. yuankui@***

AIM: To study the effects of aminoguanidine (AG) and two L-arginine analogues N(omega)-nitro-L-arginine methyl ester (L-NAME) and N(omega)-nitro-L-arginine (L-NNA) on nitric oxide (NO) production induced by cytokines (TNF-alpha, IL-1 beta, and IFN-gamma) and bacterial lipopolysaccharide (LPS) mixture (CM) in the cultured rat hepatocytes, and examine their mechanisms action. METHODS: Rat hepatocytes were incubated with AG, L-NAME, L-NNA, Actinomycin D (ActD) and dexamethasone in a medium containing CM (LPS plus TNF-alpha, IL-1 beta, and IFN-gamma) for 24h. NO production in the cultured supernatant was measured with the Griess reaction. Intracellular cGMP level was detected with radioimmunoassy. RESULTS: NO production was markedly blocked by AG and L-NAME in a dose-dependent manner under inflammatory stimuli condition triggered by CM in vitro. The rate of the maximum inhibitory effects of L-NAME (38.9%) was less potent than that obtained with AG(53.7%, P < 0.05). There was no significant difference between the inhibitory effects of AG and two L-arginine analogues on intracellular cGMP accumulation in rat cultured hepatocytes. Non-specific NOS expression inhibitor dexamethasone (DEX)and iNOS mRNA transcriptional inhibitor ActD also significantly inhibited CM-induced NO production. AG(0.1 mmol x L(-1)) and ActD (0.2 ng x L(-1)) were equipotent in decreasing NO production induced by inflammatory stimuli in vitro, and both effects were more potent than that induced by non-selectivity NOS activity inhibitor L-NAME (0.1 mmol x L(-1)) under similar stimuli conditions (P<0.01). CONCLUSION: AG is a potent selective inhibitor of inducible isoform of NOS,and the mechanism of action may be not only competitive inhibition in the substrate level, but also the gene expression level in rat hepatocytes.

关键词： Animals Antineoplastic Agents Cells, Cultured Comparative Study Cyclic GMP Cytokines Dactinomycin Dexamethasone Enzyme Inhibitors Glucocorticoids Guanidines Hepatocytes interferon type ii Interleukin-1 Lipopolysaccharides Male NG-Nitroarginine Methyl Ester Nitric Oxide Nitric Oxide Synthase inhibitors Nitroarginine Protein Synthesis Inhibitors Rats Rats, Wistar Research Support, Non-U.S. Gov't Tumor Necrosis Factor-alpha

pcDNAL1 genetic immunization can induce specific cell-mediated immune responses in C57BL/6 mice

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Chinese Medical Journal 2002年第11期115卷 1697-1700,154页

作者：宋建明刘天菊孙向乐王一理司履生深圳市第二人民医院病理科深圳518035 西安交通大学生命科学与技术学院肿瘤研究所西安710061

OBJECTIVE: To investigate the specific cell-mediated immune efficacy of the an HPV16 prophylactic vaccine. METHODS: C57BL/6 mice were randomly divided into 3 groups: experimental group I (treated with pcDNA L1), control group ii (treated with pcDNA3.1) and control group iiI (treated with PBS buffer). The mice were immunized three times during a three-week interval. Ten to fourteen days after the third inoculation, a footpad swelling test was used to detect delayed-type hypersensitivity (DTH) responses. Antigen-specific splenocyte proliferation assay and quantitation of IFN-gamma cells in splenocytes were performed by FACS assay. RESULTS: In the experimental group, splenocytes actively proliferated after stimulation with HPV16 VLP, and had developed a markedly larger amount of CD8(+) IFN-gamma(+) cells, which is an index for special CTL. Also, the footpad was significantly thickened upon inoculation with HPV16 VLP. CONCLUSION: Naked DNA vaccine of HPV16 L1 can induce specific cell-mediated immune responses in mice, which should be considered for evaluation of HPV16 DNA vaccine feasibility.

关键词： Animals Hypersensitivity, Delayed Immunization interferon type ii Lymphocyte Activation Mice Mice, Inbred C57BL Papillomavirus, Human Research Support, Non-U.S. Gov't Spleen Vaccines, DNA Viral Vaccines

A clinical trial of active immunotherapy with anti-idiotypic vaccine in nasopharyngeal carcinoma patients

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Chinese Medical Journal 2002年第4期115卷 567-570页

作者：李官成谢鹭周国华孙去病符红普周建华中南大学湘雅医学院肿瘤研究所长沙410078 湖南省肿瘤医院放疗科长沙410006 中南大学湘雅医学院病理教研室长沙410078

OBJECTIVE: To investigate the effect of active immunotherapy with anti-idiotypic vaccine in patients with nasopharyngeal carcinoma (NPC). METHODS: Anti-idiotypic antibodies (2H4/5D3) bearing the internal image of the NPC antigen were used in active immunotherapy in NPC patients receiving radiotherapy. Antibodies and cytokine levels in patient sera were determined using ELISA before and after active immunotherapy. IL-2 mRNA expression in the peripheral blood mononuclear cells (PBMC) was measured by in situ hybridization. RESULTS: Nineteen patients with NPC at stage IV were treated with alum-precipitated 2H4 or 5D3. Neither hypersensitivity nor adverse side effects were observed. The levels of anti-anti-idiotypic antibodies (Ab3) and anti-NPC antibodies (Ab1') were increased. Human anti-mouse antibodies (HAMA) were seen in 19 patients of the experimental group; the levels of Ab1' did not increase in the control group. Serum IL-2, IFN-gamma and TNF-alpha levels were increased in most patients in the experimental group, while no differences were observed in Ab1' and cytokine levels between pre- and post-therapy in the control group. In addition, IL-2 mRNA expression in PBMCs from NPC patients was closely related to serum IL-2 (r = + 0.8829) levels by in situ hybridization. CONCLUSIONS: Anti-idiotype vaccine is safe for clinical active immunotherapy. Anti-idiotypic vaccine might be able to enhance humoral and/or cellular immunity in NPC patients receiving radiotherapy.

关键词： Immunotherapy, Active Adult Antibodies, Anti-Idiotypic Antibody Specificity Cancer Vaccines Enzyme-Linked Immunosorbent Assay Female Gene Expression Regulation, Neoplastic Humans interferon type ii Interleukin-2 Male Middle Aged Nasopharyngeal Neoplasms RNA, Messenger Research Support, Non-U.S. Gov't Treatment Outcome Tumor Necrosis Factor-alpha

Different roles of PKC and PKA in effect of interferon-y on proliferation and collagen synthesis of fibroblasts

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Acta Pharmacologica Sinica 2004年第10期 80-86页

作者： Xuan-fen ZHANG, Shu-zhong GUO, Kai-hua LU, Hui-yuan LI, Xiang-dong LI, Lin-xi ZHANG, Li YANGDepartment of Plastic Surgery, Xi-jing Hospital, Fourth Military Medical University, Xi’an 710032, ChinaZHANG, PhD. Now in the Second Affiliated Hospital of Lanzhou Medical College, Lanzhou 730000, China.

AIM: To study the signal roles of protein kinase C (PKC) and protein kinase A (PKA) in the influence of interferon-γ(IFN-γ) on proliferation and collagen synthesis of flbroblasts derived from hypertrophic scar (HS-FB) and normal skin (NS-FB). METHODS: HS-FB and NS-FB were cultured and passaged in Dulbecco’s modified Eagle’s medium (DMEM). Activity of PKC and PKA were assayed by transferring phosphorus (32P) into substrate after treatment with IFN-γ1000 kU/L at 10, 30, 60, and 120 min. Cell proliferation was determined with MTT assay. The collagen synthesis was measured with [3H]proline incorporation and type Ⅲ pre-collagen was determined with radioimmunoassay. RESULTS: After exposure to IFN-γ1000 kU/L for 30 min, PKC activity of HS-FB and NS-FB increased from 2.57±0.14 and 2.13±0.12 nmol·min-1·g-1 of control to 3.75±0.19 and 3.36±0.16 nmol·min-1·g-1 respectively (P<0.05). After exposure to IFN-y 1000 kU/L for 60 and 120 min, PKA activities of HS-FB increased gradually from 0.82±0.04 nmol·m

关键词： signal transduction protein kinase C protein kinase A interferon type ii hypertrophic scar fibroblasts collagen synthesis

IFN-γ REGULATES Fas/FasL EXPRESSION IN CHOLANGIO CARCINOMA CELLS

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Chinese Journal of Cancer Research 2005年第1期17卷 40-43页

作者：李子禹张连海季加孚 Department of Surgery Peking University School of Oncology Beijing Institute for Cancer Research Beijing Cancer Hospital Beijing 100036

Objective: To study the regulation of Fas receptor (Fas)/Fas ligand (FasL) expression by IFN-γ in cholangiocarcinoma cell lines. Methods: We studied the expressions of Fas and FasL gene in the human cholangiocarcinoma cell line QBC939 by RT-PCR, Western blot, and immunohistochemistry and their apoptosis effects on human T cell line Jurkat cell. At the same time, we investigated the regulative effect of IFN-γ on them. Results: Fas, FasL mRNA and protein were expressed by cholangiocarcinoma cells. We also found IFN-γ could up-regulate the expressions of these two genes. However, IFN-γ could also down-regulate the ability of them to make Jurkat cells apoptotic. With the increasing of dosage and time, the effect was enhanced. Conclusion: IFN-γ could regulate the expression of Fas and FasL in cholangiocarcinoma cells, and might influence the ability of cholangiocarcinoma to regulate immune escape. This study provides new theoretical basis for immunological therapy of cholangiocarcinoma.

关键词： Cholangiocarcinoma interferon type ii Apoptosis