Temporal relationship between immune cell influx and the expression of inducible nitric oxide synthase, interleukin-4 and interferon-gamma in pancreatic islets of NOD mice following adoptive transfer of diabetic spleen cells

Beta cell destruction in NOD mice can be accelerated by adoptive transfer o f diabetic spleen cells into irradiated adult NOD mice. Here mice receiving diabetic spleen cells were examined at days 0, 7, 14, 21 and at onset of d iabetes for the resulting insulitis and the number of intra-islet CD4 and C D8 cells and macrophages. The progression of insulitis and the number of in tra-islet CD4 and CD8 cells and macrophages were correlated with the expres sion and co-localization of inducible nitric oxide synthase, interferon-gam ma and interleukin-4 by dual-label light and confocal immunofluorescence mi croscopy. Diabetes developed in 7/8 mice by 27 days following cell transfer . The insulitis score increased slightly by day 7 but rose sharply at day 1 4 (p=0.001) and was maintained until diabetes. The mean number of intra-isl et CD4 and CD8 cells and macrophages showed a similar trend to the insuliti s scores and were present in almost equal numbers within the islets. Immuno labelling for inducible nitric oxide synthase was observed at day 7 in only some cells of a few islets but increased sharply from day 14. It was restr icted to islets with insulitis and was co-localized in selective macrophage s. Weak intra-islet interleukin-4 labelling was observed at days 7 and 14 b ut became more pronounced at day 21 and at onset of diabetes, being present in selective CD4 cells. Intra-islet labelling for interferon-gamma was fir st observed at day 21, but became more intense at onset of diabetes and was co-localized in a proportion of macrophages. Both cytokines were expressed in islets with advanced insulitis. Interferon-gamma staining was also obse rved within endothelial cells located in the exocrine pancreas. We conclude that transfer of diabetic spleen cells results in a rapid influx of CD4 an d CD8 cells and macrophages within the pancreas of recipient mice. During t he period of heightened insulitis, selective immune cells begin to express inducible nitric oxide synthase and the opposing cytokines, interferon-gamm a and interleukin-4. Expression of these molecules becomes more pronounced immediately prior to and during the onset of diabetes.