Background & objective: Costimulator OX40 (CD134) belongs to TNFR family, and its ligand OX40L (CD134L) belongs to TNF family. OX40-OX40L signal plays an important role in immune regulation, which can increase the production of cytokines and enhance the survival of antigen-specific T cells. This research was to transfect OX40L into human breast cancer cell line MDA-MB-435 to construct OX40L-MDA-MB-435 cells, and to investigate the costimulatory effect of OX40-OX40L signal on biological activity of T cells in vitro.
Methods: cDNA fragment encoding OX40L was obtained from human mature dendritic cells by reverse transcription-polymerase chain reaction (RT-PCR), and inserted into eukaryotic expression vector pcDNA3.1. The recombinant plasmid pcDNA3.1-OX40L was transfected into MDA-MB-435 cells, and verified by indirect immunophenotyping and RT-PCR. The effect of OX40L-MDA-MB-435 cells to the biological activity of T cells was investigated by MTT, enzyme-linked immunosorbent assay (ELISA), and direct immunophenotyping.
Results: OX40L-MDA-MB-435 cells were successfully constructed. These cells efficiently promoted the proliferation of T cells, enhanced the secretion of interleukin-2 (IL-2) (973.4 ng/ml at the 7th day) and interferon-gamma (IFN-gamma) (3,689.167 pg/ml at the 3rd day), and down-regulated the expression of Fas on T cells [(68.3+/-5.6)%, P<0.05].
Conclusion: OX40L-MDA-MB-435 cells could activate T cells in vitro, promote T cell proliferation and cytokine secretion, and suppress T cell activation-induced cell death.