Aim: To establish a method to expand dentritic cells (DC) in vivo by combined Flt3-L-expressing plasmid with GM-CSF-expressing plasmid and to analyze their function in antigen presenting.
Methods: Flt3-L and GM-CSF expressing DNAs were injected into the tail veins of BALB/c mice on day 0 and day 10 respectively. The spleens were harvested on day 15 and the splenocytes were prepared to analyze the percentage of CD11c, CD11b, B220, CD8, and NK1. 1 positive cells by FACS. The splenocytes from HBsAg-vaccinated mice were simulated in vitro with DC pre-incubated with HBsAg and then IFN-gamma in supernatant after simulation was detected by ELISA.
Results: Combined injection of Flt3-L-expressing plasmid with GM-CSF-expressing plasmid resulted in dramatic plednomegaly in mice. Averagely 4 x 10(8) splenocytes were harvested from the expanded spleen of each mouse, more than 20% of which were CD11c(+) DC. Among CD11C(+) splenocytes, 17% of CD11B(+) cells, 10% of CD11b(-) cells, 26% of B220(+) cells, 16% of CD8(+) cells and 7% of NK1.1(+) cells were detected respectively. When loaded with HBsAg, Flt3-L/GM-GSF DC could stimulate HBsAg-primed splenocytes to secrete much higher IFN-gamma than DC or HBsAg alone.
Conclusion: Combined intravenous injection of Flt3-L-expressing plasmid with GM-CSF-expressing plasmid could in vivo expand dentritic cells which seemed to have many subsets and were able to present antigens to antigen special T cells in vitro.