Abstract
CD40 ligand (CD40L) expressed by activated CD4+ T cells is a family member of membrane bound TNF family ligand and its interaction with CD40 expressed in APC has been shown to contribute in enhancing immune response. Exogenous stimulation through CD40 has been performed using soluble trimeric CD40L, anti-CD40 monoclonal antibody and cells expressing CD40L. Schneider 2 (S2) cells, a cell line derived from Drosophila melanogaster, was transfected with a plasmid vector, pAc5.1/V5-HisA, for the constitutive expression of CD40L (S2-CD40L). Upon incubation of S2-CD40L with B-lymphocytes for 6 days, activated B cells were examined by counting B cell numbers and for activation markers including CD86 and HLA Class II molecules. The activated B cells were tested for its efficient APC function by mixed lymphocyte reactions (MLR) and enzyme-linked Immunospot (ELISPOT) assay. S2-CD40L was cultured for a year and maintained CD40L expression (>90%). S2-CD40L induced B cell activation as demonstrated by increment of total B cells and up-regulation of CD86 and MHC Class II molecules. Activated B cells pulsed with peptide from human cytomegalovirus pp65 antigen efficiently induced both proliferation and IFN-gamma secretion of T cells. Our result suggests that S2-CD40L can efficiently and conveniently generate B cells as a functional APC and represents a potential role for B-cell mediated cancer immunotherapy.
Highlights
Cancer immunotherapy has been associated with augmentation of immune response via in vivo vaccination or ex vivo expansion of antigen specific effector cells that show cytotoxicity and secretion of inflammatory molecules
We developed non-allogenic and non-xenogenic B cell stimulating system by establishing a cell line derived from Drosophila melanogaster, Schneider 2 (S2) cells stably expressing CD40 ligand (S2-CD40L)
To verify whether CD40L expression can be maintained at which B cells are cultured, 37oC, S2CD40L transfectants were suspended in B cell media, Iscove's modified Dulbecco's medium (IMDM) (10% FBS) and incubated at 37oC
Summary
Cancer immunotherapy has been associated with augmentation of immune response via in vivo vaccination or ex vivo expansion of antigen specific effector cells that show cytotoxicity and secretion of inflammatory molecules. Preclinical studies have demonstrated the important role of APC in antigen presentation to prime T lymphocytes. For effective induction of T cell mediated immune response, efficient presentation of antigen peptides is required. Most of studies on generation of antigen specific T cells in vitro have used dendritic cells for its capacity; antigen capture and efficient presentation. In order to prime naive T cells to be antigen specific, the fact that DCs only constitute 0.1-0.5% of human PBMC has hampered generating a large numbers of matured DC (Girolomoni et al, 1997). DCs generated via this method are not fully functional in stimulating antigen-specific T cells in vitro or
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