Construction of a stably transfected cell line expressing mouse inducible costimulatory ligand and its role in T cell activation and proliferation

Objective To construct a mouse inducible costimulatory ligand (ICOSL) stably transfected cell line and explore its effect on T cell activation and proliferation.Methods RT-PCR was performed to obtain full length cDNA of ICOSL from mouse spleen RNA.The PCR product was digested with restriction endonucleases EcoR Ⅰ and BamH Ⅰ,and subcloned into an eukaryotic expression vector EGFP-pIRES2.CHO cells were stably transfected with the obtained expression vector using Lipfectamine TM2000.The positive cell clones were selected with G418.Flow cytometry and RT-PCR were used to detect the expression of mouse ICOSL in stably transfected cells.In order to explore the effect of mouse ICOSL on T cells,the CHO/ICOSL cells were co-cultured with T cells.The expression of T cell activation markers CD25 and CD69 were detected using flow cytometry.T cell proliferation was measured by the CCK8 colorimetric assay.Results The expression vector of mouse ICOSL was successfully constructed and stably expressed in CHO cells.Coculture of T cells with mouse ICOSL stably transfected cells resulted in the up-regulation of CD25 and CD69 in the activated T cells and increased T cell proliferation.Conclusion Mouse ICOSL could induce T cell activation and proliferation. Key words: Costimatory molecar;  Inducible costimulatory ligand;  Transfected gene cell