动物造模 z-bio 2022-03-08 495

　　Objective To investigate the genetic and immunological mechanisms of guinea pigs resistance to foot-and-mouth disease virus (FMDV). To elucidate the feasibility of FMDV pseudovirus-infected cells for studying FMDV pathogenesis.

　　Methods Guinea pig primary kidney cells were infected by lentiviral vector containing FMDV-VP1 fragment to construct FMDV pseudovirus-infected target cells. Two FMDV-susceptible (Zmu-1:DHP strains) and resistant (British strains) guinea pig peripheral blood lymphocytes (PBLCs, not treated with antigen) were isolated and co-cultured with pseudovirus-infected target cells. Toxicity experiments were used to determine the killing rate of PBLC, and ELISA was used to detect the content of cytokines in the cell culture supernatant. The ratios of CD3+/CD4+ and CD3+/CD8+ cells in guinea pig PBLC of two strains were determined by flow cytometry.

　　Results For both strains of guinea pigs, the cytotoxicity of PBLC in the experimental group was significantly greater than that in the respective control groups (P < 0.01); the cytotoxicity of PBLC in the resistant strain on target cells was significantly greater than that in the susceptible strain (P < 0. 01); the expression levels of cell killing factors PF, GRA, GRB and cell surface Fas secreted by PBLC of resistant strains were significantly higher than those of susceptible strains ( P < 0. 01), but there was no significant difference in the expression of secreted FasL ( P < 0. 01). The contents of cytokines IFN-γ, TNF-α, IL-2 and IL-12 secreted by PBLC of resistant strains were significantly higher than those of susceptible strains (P < 0.05). There was no significant difference in MHC II (P > 0.05); the lentiviral vector containing the FMDV-VP1 fragment infects primary guinea pig kidney cells, which can express the FMDV-VP1 protein on the cell surface; the number of CD4+ cells in the PBLC of the resistant strains was significant Less than the susceptible strain (P < 0. 05), the number of CD8+ cells in the resistant strain was significantly higher than that in the susceptible strain (P < 0. 05).

　　Conclusion FMDV-VP1 fragment significantly stimulates the immune response of effector cells and enhances the cytotoxicity of effector cells to target cells; resistant guinea pig PBLC has strong natural cytotoxicity to FMDV-infected cells; the process of effector cells producing cytotoxicity to target cells The cellular immune factors secreted by effector cells are involved in the related process; guinea pig primary kidney cells were infected by lentiviral vector containing FMDV-VP1 fragment, and the target cells infected with FMDV pseudovirus were constructed to effectively simulate the infection of target cells by FMDV. Improve the safety of follow-up research work. The genetic characteristics of different strains of guinea pigs determine their differences in susceptibility to FMDV.