The Induction Of Murine Regulatory Dendritic Cells In Vivo And In Vitro

Objective To establish the methods to induce regulatory dendritic cells in vivo and in vitro and to compare the capability of C57BL/6 and NOD/LtJ mice to generate regulatory dendritic cells, we have designed and completed the following studies. Methods Regulatory dendritic cells were induced in vivo by intraperitoneal injection of bacteria E. coli K12. The surface markers of the regulatory dendritic cells generated in the spleen were analyzed by flow cytometry. The optimal conditions required to induce regulatory dendritic cells were screened by injecting different doses of bacteria and by analyzing at different time points. Meanwhile the effects of LPS and thioglycolate on the generation of regulatory dendritic cells were investigated. The capability of C57BL/6 and NOD/LtJ mice to generate regulatory dendritic cells was compared. Finally, whether regulatory dendritic cells can promote the generation of CD4~+Foxp3~+ regulatory T cells was tested. Regulatory dendritic cells were induced in vitro by coculturing bone-marrow mononuclear cells with endothelial splenic stromal cells. And the regulatory dendritic cells were identified by flow cytometry. Results Regulatory dendritic cells can be induced in vivo by intraperitoneal injection of bacteria E. coli K12. The induction effect of the group injected with 1×10~8 bacteria is most obvious, and the generation of regulatory dendritic cells have time effect, in which it’s 5 days after injection of bacteria is particularly significant. Intraperitoneal injection of Brewer thioglycollate medium solution can’t induce the generation of regulatory dendritic cells. In contrast with C57BL/6 mice, NOD/LtJ mice is more easily to induce regulatory dendritic cells. And regulatory dendritic cells can’t promote the generation of CD4~+Foxp3~+ regulatory T cells. Meanwhile approach of in vitro can induce bone-marrow mononuclear cells differentied into regulatory dendritic cells. Conclusions Both methods of in vivo and in vitro can induce regulatory dendritic cells.