Development of an I-Ag7-expressing antigen-presenting cell line: intrinsic molecular defect in compact I-Ag7 dimer...

Insulin-dependent diabetes mellitus (IDDM) results from chronic, T-cell dependent, autoimmune destruction of the insulin-producing beta-cells in the Langerhans' islets of the pancreas. Non-obese diabetic (NOD) mice spontaneously develop IDDM that resembles human type I diabetes. The susceptibility to diabetes in the NOD strain is a complex polygenic trait that determines a phenotype of immune alterations. The unique MHC class II molecule expressed by NOD mice (I-Ag7) plays a major role in the development of disease. Recently, it has been reported that I-Ag7 molecules generate a lower proportion of compact alphabeta heterodimers, compared to other haplotypes. However, it is not clear whether this reflects an intrinsic defect of this molecule to bind peptide stably or is the result of abnormal processing and/or peptide loading into the I-Ag7 molecule. Our aim was to develop and characterize a suitable antigen-presenting cell (APC) that expressed I-Ag7 in the context of a non-diabetes-prone antigen processing and presentation machinery. Here, we report the generation of a mouse DAP.3 fibroblast cell line (DAP.3Ag7) that constitutively expresses high levels of I-Ag7. Using DAP.3 cells transfected with I-Ag7 or I-Ak, we show that the expression of compact dimers in the same cell type is proportionally less for I-Ag7 molecules than for I-Ak molecules, implying an intrinsic defect of the I-Ag7 molecule as the cause for the low generation of compact dimers. However, DAP.3Ag7 cells are able to process and present antigen, as indicated by I-Ag7-dependent IL-2 production by a GAD67-specific NDO T-cell hybridoma after stimulation with GAD and live, but not fixed, DAP.3Ag7 cells. The IL-2 response to GAD when presented by DAP.3Ag7 was significantly higher than the response to GAD presented by NOD splenocytes. Based on these data, we conclude that the low generations of compact dimers is an intrinsic feature of I-Ag7 molecules and not affected by other genes in the NOD background. The DAP.3Ag7 cell line should be a valuable tool with which to dissect the role of the I-Ag7 molecule in antigen presentation and T-cell activation in NOD mice, which clearly contributes to the development of IDDM.