DETERMINATION OF MESSENGER-RNA EXPRESSION FOR IFN-GAMMA AND IL-4 IN LYMPHOCYTES FROM CHILDREN WITH IDDM BY RT-PCR TECHNIQUE

Insulin-dependent diabetes mellitus (IDDM) is characterized by infiltr ation of T-lymphocytes in the islets of Langerhans. Antigens are prese nted to Th-lymphocytes which can be divided into Th1- and Th2-lymphocy tes, producing interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) r espectively. The aim of our study was to determine the messenger-RNA ( mRNA) for these cytokines by RT-PCR in antigen-stimulated lymphocytes from children with newly diagnosed IDDM. The expression of mRNA for IL -4, and to a lesser degree IFN-gamma, is increased in lymphocytes stim ulated with tetanus toroid (TT). Loss of activity after freezing and t hawing could be compensated for, by increased amplification, while the use of EDTA or sodium heparin in the blood samples did not influence the results. In a pilot application, the lymphocytes from children wit h newly diagnosed IDDM were stimulated with a peptide of glutamic acid decarboxylase (GAD) (a.a. 247-279) known to have a similar aminoacid sequence as the Coxsackie B virus (a.a. 32-47). Increased IFN-gamma mR NA could be seen in two out of four children, whereas IL-4 showed a le ss pronounced mRNA expression. No increased mRNA expression for IFN-ga mma and IL-4 could be seen in healthy HLA-matched controls. Further st udies are needed to confirm whether increased IFN-gamma, mRNA in Th1-l ike lymphocytes stimulated with this specific GAD-peptide play a role in the cell-mediated immune response seen in children early after the onset of IDDM. (C) 1998 Elsevier Science Ireland Ltd. All rights reser ved.