ABSTRACT
Binding activity and biologic effect of a novel alpha-melanocyte-stimulating hormone analogue were tested on cells transiently expressing the human melanocortin-1 (MC1), MC3, MC4, and MC5 receptors. The human MC1 and MC5 receptor genes were cloned into the expression vector pcDNA3. 1/ myc-his(-) B. The vectors were transferred to HEK-293 cells by the calcium phosphate method. Stable receptor populations were generated using G418 selection (900 microg x mL(-1)) for subsequent bioassay analysis. K(i) values of the novel alpha-MSH analogue for MC1, MC3, MC4, and MC5 receptors were obtained in competition with [125I]-NDP-MSH for binding studies. The cyclic AMP level was tested by using [3H]-cyclic AMP kit. It is showed that K(i) values of the novel alpha-MSH analogue for MC1, MC3, MC4, and MC5 receptors were (0.159 +/- 0.040), (35.430 +/- 6.743), (19.293 +/- 2.780) and (2.230 +/- 0.670) nmol L(-1), respectively. Its EC50 values for MC1, MC3, MC4, and MC5 receptors were (0.45 +/- 0.07), (7.80 +/- 0.65), (2.55 +/- 0.23) and (0.33 +/- 0.09) nmol L(-1), respectively. In these tests, the novel alpha-MSH analogue is a MC1R and MC5R selective agonist.
Subject(s)
Humans , Amino Acid Sequence , Binding, Competitive , Cell Line , Cell Line, Tumor , Cyclic AMP , Metabolism , Genetic Vectors , Iodine Radioisotopes , Kinetics , Molecular Sequence Data , Plasmids , Genetics , Radioligand Assay , Receptor, Melanocortin, Type 1 , Genetics , Metabolism , Receptors, Corticotropin , Genetics , Metabolism , Receptors, Melanocortin , Genetics , Metabolism , Transfection , Tritium , alpha-MSH , Chemistry , Metabolism , PharmacologyABSTRACT
Objective: To investigate the role of T cell in the antitumor immune responses induced by MIF gene-modified tumor vaccine. Methods: MIF gene was transferred into FBL3 erythroleukemia cel l by adenovirus carrier and a new type of tumor vaccine was prepared. The chang es of the number and the function of T cell in spleen and lymph node was observe d. Results: After the mice were immunized with MIF gene-m odified FBL3 vaccine, the number of lymphocyte in spleens and lymph nodes increa sed markedly and the specific CTL activities of splenocytes also increased great ly. FACS analysis showed that the CD3+, CD4+, CD8+ T cells and CD28 posi tive cells in draining lymph nodes of MIF-FBL3 group mice increased more marked ly than that of control groups. When the wild type FBL3 cells were injected into the mice immunized with MIF gene-modified FBL3 vaccine, the growth of tumors w ere obviously inhibited and the survival rate of the mice was increased. Conclusion: It is suggested that MIF gene-modified tumor vaccine can induce specific antitumor immune responses mediated by T cells and may be a candidate for gene therapy of tumor.
ABSTRACT
Objective: To investigate the role of T cell in the antitumor immune responses induced by MIF gene-modified tumor vaccine. Methods: MIF gene was transferred into FBL3 erythroleukemia cel l by adenovirus carrier and a new type of tumor vaccine was prepared. The chang es of the number and the function of T cell in spleen and lymph node was observe d. Results: After the mice were immunized with MIF gene-m odified FBL3 vaccine, the number of lymphocyte in spleens and lymph nodes increa sed markedly and the specific CTL activities of splenocytes also increased great ly. FACS analysis showed that the CD3+, CD4+, CD8+ T cells and CD28 posi tive cells in draining lymph nodes of MIF-FBL3 group mice increased more marked ly than that of control groups. When the wild type FBL3 cells were injected into the mice immunized with MIF gene-modified FBL3 vaccine, the growth of tumors w ere obviously inhibited and the survival rate of the mice was increased. Conclusion: It is suggested that MIF gene-modified tumor vaccine can induce specific antitumor immune responses mediated by T cells and may be a candidate for gene therapy of tumor.