The molecular adjuvant hC3d3 improves expressions of co-stimulatory molecules in human immuno-competent cells and their responsiveness to the hCGβ antigen
Objective To find if human molecular adjuvant hC3d3 promotes expressions of co-stimulatory molecules in human immuno-competent cells and responsiveness to the hCGβ antigen when fused into the antigen. Methods Human C3d gene was cloned from human hepatic cDNA library, the eukaryotic expression plasraids of pCI-gs-signal-6His-hCGβ and pCI-gs-signal-6His-hCGβ-hC3d3 were constructed. CHO cells were transfected by the recombinant expression plasmids with aid of LipofectAMINE 2000 reagent. The recombinant proteins were expressed and purified using His. Bind cohann and Sephadex G150 column gel filtration chromatography. The isolated B cells, B cells ＋ T cells and PBMC were treated with 100 nmol/L hCGβ, hCGβ-hC3d3 or PWM for 48 h. The expression of CD80, CD86, CD154 and CD25 in the cells was analyzed by flow cytometry. The IL-2 production in the supematant was assayed by sandwich ELISA. Results The hC3d gene has been successfully cloned from human hepatic cDNA library, and the eukaryotic expression plasmids of pCI-gs-signal-6His-hCGβ and pCI-gs- signal-6His-hCGβ-hC3d3 were constructed. The correspondent recombinant proteins were secreted into the cultural supematant by CHO cells. The hCGβ-hC3d3 significantly enhanced the expressions of CD80 and CD86 molecule in B cell, especially CD86 （P 〈0.01）, and that of CD154 and CD25 molecule in T cells, hCGβ-hC3d3 promoted human PBMC producing more IL-2 than the hCGβ. Conclusion The human molecular adjuvant hC3d improves expressions of co-stimulatory molecules in human immuno-competent cells and their responsiveness to the hCGβ antigen. 
 
Key words: 
hCGβ;  hC3d3;  Immuno-competent cells;  Co-stimulatory molecules;  IL-2