CYTOKINE GENE-EXPRESSION IN THE BE RAT PANCREAS - NATURAL COURSE AND IMPACT OF BACTERIAL VACCINES

In diabetes prone BE rat pancreas the Th1/Th2 cytokine balance and the expression of inducible nitric oxide synthase (iNOS) was determined b y mRNA analysis before and after the onset of insulitis. Specific mRNA was amplified by reverse transcriptase polymerase chain reaction, qua ntitated with radiolabelled probes by phosphoimaging and calibrated wi th the amount of co-amplified beta-actin mRNA. At 50 days of age, prio r to recognizeable insulitis, there was already significantly enhanced expression of both, Th1 and Th2 cytokines, and of iNOS mRNA, when com pared to Wistar rat pancreas (p < 0.001). This supports the concept of an inconspicuous early phase of islet infiltration by single immunocy tes, called single eel insulitis. At 70 days of age mononuclear infilt ration of islets had begun and was associated with upregulation of int erferon gamma (IFN gamma) and iNOS, but downregulation of interleukin- 10 and transforming growth factor beta mRNA (p < 0.001). These finding s correlate the onset of insulitis with a shift of the Th1/Th2 cytokin e balance towards Th1 cell reactivity. Indeed there was a close correl ation of the Th1/Th2 cytokine ratio but not of absolute IFN gamma mRNA levels with the insulitis score. Vaccination at day 50 with tetanus t oroid did not affect cytokine gene expression while diphtheria toroid and even more strongly BCG administration induced a shift towards Th2 reactivity (p < 0.001) while iNOS mRNA was decreased (p < 0.01). Oral dosing with immunostimulatory components of Escherichia coli also chan ged the quality of inflammation. Oral lipopolysaccharide (LPS) from E. coli and OM-89, an endotoxin free extract containing immunostimulator y glycolipopeptides and heat shock protein (hsp) 65, both downregulate d IFN gamma mRNA while only OM-89 in addition suppressed INOS mRNA and enhanced Th2 cytokine gene expression (p < 0.001). We conclude that t he onset of insulitis is associated with a shift towards Th1 cytokine and iNOS gene expression. Diphtheria toroid and BCG vaccination stimul ates Th2 reactivity but does not: downregulate Th1. The latter can be achieved through oral administration of LPS or a glycopeptide fraction (OM-89) from E. coli.