Effects of B7-2 gene RNAi lentiviral vector on dendritic cells and T lymphocytes proliferation induced by dendritic cells in vitro
10.3969/j.issn.1000-484X.2019.02.003
- VernacularTitle:慢病毒介导B7-2基因RNA干扰对小鼠树突状细胞诱导T细胞增殖的影响
- Author:
Yong KONG
1
;
Tian-Ming YAN
;
Yu-Yu WANG
;
Li-Jun SHEN
;
Jing WANG
;
Yu-Hua QIU
Author Information
1. 苏州大学医学部免疫学系
- Keywords:
B7-2;
RNAi;
Lentivirus;
Dendritic cell;
Co-stimulatory signal
- From:
Chinese Journal of Immunology
2019;35(2):140-145
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To generate mouse B7-2 gene RNAi lentivirus and study its interference effects on B7-2 expression and T lymphocytes proliferation induced by dendritic cells. Methods: Three sequences specific targeting B7-2 gene and one non-specific sequence were respectively synthesized, and inserted into lentiviral vector, then the recombinant vectors were sequencing. 293 T cells were co-transfected with lentiviral expression plasmid and packaging plasmids to produce recombinant lentivirus which titre was checked according to the expression level of green fluorescent protein ( GFP). Bone marrow cells from C57 BL/6 mice were isolated to differentiate into DCs at the present of GM-CSF, IL-4 and LPS for 48 h, then morphology and phenotypic was identified. DCs were infected by recombinant RNAi lentivirus and then the efficiency of infection and the expression of B7-2 on the surface of DCs were detected by flow cytometry. Effects on the proliferation of T cells were detected by co-culturing with DCs which were infected by B7-2 RNAi lentivirus and murine spleen T cells in vitro. Results: DNA sequencing confirmed that three B7-2 RNAi and one non-specific recombinant lentiviral transfer plasmids were successfully constructed, the titer of recombinant lentivirus was ( 2-4) × 108 TU/ml. The recombinant lentivirus could effectively infect DC and inhibit the expression of B7-2. After the B7-2 recombinant lentivirus infection, the ability of DCs to stimulate the proliferation of T cells decreased obviously ( P<0. 05). Conclusion: The lentiviral B7-2 gene RNAi vector can effectively silence the expression of B7-2 on the surface of DCs and inhibit the proliferation effect of T cells induced by DCs.