The effects of JNK1 on the responses of cytolytic T cells against viral infection / 中华微生物学和免疫学杂志

Objective To examine the role of c-Jun NH2-terminal kinase(JNK) 1 in cytolytic T lymphocyte (CTL) responses against virus infection.Methods Wild-type (WT) and JNK1-knockout (JNK1-/-) mice were infected with ectromelia virus(ECTV) through hind footpads.Survival and virus titers in the target organs (liver and spleen) were analyzed.Effector T cells in the spleen and popliteal lymph nodes were determined on day 3 and 7 post-infection.Proliferation and INF-γ production of CTL were also detected.Results JNK1 deficiency caused an increased susceptibility to ECTV infection in mice,indicated by higher case fatality and viral burden in target organs.The decrease in CTL response correlated with a defect in CTL proliferation and INF-γproduction.Conclusion The data suggest that JNK1 is involved in expansion of activated CTL during ECTV infection,and plays an important antiviral role in regulating the proliferation and effector function of CTL.

Genetic construction of HLA-A2 restricted multi-epitopes gene of hepatitis C virus,expression and purification in E.coli for antigenic analysis / 中国免疫学杂志

Objective:To construct the recombinant prokaryotic plasmid to express HCV HLA-A2 restricted multi-CTL epitopes and to purify the fused protein for antigenic analysis.Methods:The human ubiquitin gene and multi-CTL epitopes gene was synthesized respectively,and digested by restrict enzyme before being cloned into pRSET-A.Then it was transformed into E.coli DH5α and the positive recombinant plasmid named pRSET-Ub-Mep was sequenced.Target protein was distinctly expressed after transformed into E.coli BL21 and induced with IPTG.Thus the protein was scanned and purified on Ni~(2+)-NTA column as well as Western blot performed after solubility analysis.Results:The recombinant plasmid pRSET-Ub-Mep was successfully constructed and it could efficiently express the target gene.Protein production was mainly in inclusion body and could be purified through Ni~(2+)-NTA column.The purified protein kept the antigen activity.Conclusion:The gene encoding for HCV HLA-A2-restricted multi-CTL epitopes is efficiently expressed and the target protein is purified,which establishes a foundation of further research to evaluate the cellular immune response induced by the target gene.

Role of adenovirus mediated CTLA4-Ig gene in the immune cells infiltration and cell apoptosis in murine liver transplantation / 中国普通外科杂志

Objective To investigate the role of B7/CD28 costimulation pathway blockade with adenovirus-mediated CTLA4-Ig gene in macrophage and CD8~+T cell infiltration and cell apoptosis in murine liver transplantation. Methods Rat pairs were divided into three groups: SD-to-Wistar transplantation control group, CsA-treated group and CTLA4-Ig-treated group. IHC and TUNEL were used to analyze the expression of CTLA4-Ig gene in liver and immune cells infiltrate and cell apoptosis in liver grafts. Pathology was done on all harvested grafts. ResultsCTLA4-Ig gene expression was positive in the donor liver on day 7 after administering adenovirus-mediated CTLA4-Ig gene via vein, and remained positive until day 60 after liver transplantation. Infiltration of immune cells in CTLA4-Ig-treated group was less than that in rejection control group. the apoptotic index of rejection group on day 3,5,7 was significantly higher than those of CTLA4-Ig-treated. Conclusions CTLA4-Ig gene was constantly expressed in the donor liver after single intravenousely injection into rats using adenovirus as vector. Adenovirus-mediated CTLA4-Ig gene therapy can inhibit infiltration of immune cells and apoptosis in grafts, thus prolonging the survival of recipients.

Induction of specific cytotoxic T lymphocytes using hepatoma antigenic peptide mixed with HSP70 in vitro / 中国免疫学杂志

Objective:To explore the possibility of inducing cell mediated immune response with HSP70 antigenic peptide complex in vitro.Methods:HSP70 peptide complex was reconstituted in vitro.Granulocyte/macrophage colony stimulating factors and interleukin 4 were used to cultivate DC from peripheral blood of HLA A2 positive healthy donors.HSP70,HSP70 peptide complex or peptide was used to activate the DC individually,which will initiate homogenize T lymphocyte to form cytotoxic T lymphocyte(CTL).The cytotxicity of the CTL was detected by MTT assay.Results:It was found that peptide specific CD8 + CTL responses were readily elicited by HSP70 peptide complex or peptide.The CTL response primed by HSP70 peptide complex was more potene than peptide alone.Conclusion:The results suggest that HSP70 peptide complex is immunogenic and HSP70 can lead to great efficient CTL response,antigenic peptides and HSP70 complex may be used as peptide vaccines for cancer immunotherapy.