The Immunoprophylactic Effect Of Inhibiting Liver Cancer Expressed Positive FoxM1 After Dendritic Cells Vaccine Loaded With CTP-FoxM1 Fusion Protein Injected C57BL/6 Mice

Objective: To investigate the immunoprophylactic effect of inhibiting liver cancer expressed positive FoxM1 after DCs vaccine loaded with CTP-FoxM1 fusion protein injected C57BL/6 mice.Methods:(1) According to Gen Bank, we selected the key antigen epitope sequences of C57BL/6 mice FoxM1 and 4 polymers in series. It linked to CTP and CTP-Fox M1 recombinant gene was inserted into pCold-TF vector, regarded as pCold-TF-CTP-Fox M1 plasmid.CTP-FoxM1 fusion protein was expressed in E.coli. BL21 bacterium and purely obtained by Ni2+-NTA. Moreover, the localization of CTP-FoxM1 fusion protein was in DCs through cellular immunofluorescence staining and laser confocal microscopy.(2)After DCs were transfected with CTP-FoxM1, FoxM1, CTP or PBS for 48 hours, the proliferative capability of lymphocyte stimulated by DCs with antigen was measured by CCK-8 kit and the CTL response effectof Hepa1-6 liver cancer lines killed by DCs with antigen in vitro was detected by LDH kit.(3) C57BL/6 mice were subcutaneously injected with Hepa1-6 liver cancer lines, after DCs vaccine loaded with CTP-FoxM1, FoxM1 or CTP protein was injected into C57BL/6 mice. Subsequently, the immunoprophylactic effect of inhibiting liver cancer of C57BL/6 mice using DCs vaccine with antigen was observed.Results:(1) pCold-TF-CTP-FoxM1 recombinant plasmid was successfully established; The efficient expression of soluble CTP-FoxM1 fusion protein was obtained by induction of 1 mM IPTG in 37℃ for 3hours; The purity of CTP-FoxM1 fusion protein was about 90% by Ni2+-NTA and was identified by Western blot; CTP-FoxM1 fusion protein was mainly into the cytoplasm of DCs through cellular immunofluorescence staining and laser confocal microscopy.(2) DCs vaccine loaded with CTP-FoxM1 fusion protein more obviously stimulated the proliferaton of lymphocyte compared with DCs vaccine loaded with FoxM1 or CTP, moreover, the proliferation of lymphocyte was in the best appropriate status at the ratio of 10:1. The CTL respones effect of killing Hepa1-6 liver cancer lines in vitro, DCs vaccine loaded with CTP-FoxM1 fusion protein had more obvious effect on killing Hepa1-6 liver cancer lines compared than other groups.(3) In the immunoprophylactic effetct observation of aganisting livercancer of C57BL/6 mice using DCs vaccine with antigen(CTP-FoxM1,FoxM1, CTP), DCs vaccine loaded with CTP-FoxM1 fusion protein had more obvious immunoprophylactic effect on inhibiting the growth of C57BL/6 liver cancer than that in other groups at 7th, 9th, 11 th, 13 th, 15 th and17th after C57BL/6 mice were subcutaneously injected with Hepa1-6 liver cancer linesConclusion:(1) The high pure CTP-FoxM1 fusion protein was obtained by Ni2+-NTA and cytoplasmic localization of CTP-FoxM1 fusion protein in DCs was observed by laser confocal microscopy, thus it laid a foundation for production of DCs vaccine loaded with CTP-FoxM1 fusion protein.(2) DCs loaded with CTP-FoxM1 fusion protein could effectively stimulate the proliferaton of lymphocyte and played a role in arousing specific CTL response against Hepa1-6 liver cancer lines in vitro.(3) DCs vaccine loaded with CTP-FoxM1 fusion protein had immunoprophylactic effect on inhibiting the growth of C57BL/6 liver cancer, thus, it laid a theoretical foundation for the clinical application of DCs vaccine in prevention and treatment of liver cancer.