113. Studies on the mechanism of the dysregulation of t-cell interferon- γ production in ANCA-vasculitis patients

Background: T cells play an important role in pathogenesis of ANCA vasculitis. Predominant presence of CD4+ T cell subsets and proinflammatory cytokines such as IFN-γ in inflammatory lesions implies that T cells during remission contribute to autoantibody generation and subsequent disseminated vasculitis. Previously, we have shown that B cells are unable to down regulate IFN-γ production by activated CD3+ T-cells from AAV patients during remission, in contrast to healthy control. The aim of this study is to investigate the mechanisfor this dysregulation of IFN-γ production. Methods: To reveal mechanisfor B-cell inhibition of T-cell IFN-γ production, inhibitors of the following molecules were added to co-cultures of cells from healthy controls (HC): PDL1, CTLA4, IL-10, B- and T- lymphocyte attenuator (BTLA) protein. Sandwich ELISA were used to measure IFN-γ and other cytokines in culture supernatants. Intracellular staining for Foxp3 was used to enumerate Treg cell generation. The effect of recombinant IL-10 on IFN-γ production of isolated T cells was assessed in both HC and AAV patients in remission. This effect was studied in different T cell subsets (CD3+, CD4+, CD8+,CD45RO+, CD45RO-, Th1, Th2, Th17) after sorting by FACS. Results: The inhibition of IFN-γ in HC (n=4) was attenuated by blocking PDL1, CTLA4, BTLA or IL-10 on activated CD3+ T cells in co-culture with B cells. However, this attenuation was markedly higher by blocking the IL-10 receptor compared (343 pg/ml), with the other blockers (298 pg/ml, 287 pg/ml, 265 pg/ml, p=0.028, p=0.027, p=0.057 respectively). Addition of recombinant IL-10 to activated CD3+ T-cells increased numbers of Treg cells in both AAV patients (p=0.001, n=10) and HC (p=0.004, n=4), but supressed IFN-γ secretion only in HC; activated T-cells alone vs activated T cell + recombinant IL-10 (395 pg/ml vs 297 pg/ml, p=0.013), not in AAV patients; activated T-cells alone vs activated T cell + recombinant IL-10 (322 pg/ml vs 327 pg/ml, p=0.484). Preliminary data from the T-cell subsets indicate that IFN-γ production is elevated and unresponsive to regulation primarily in the Th1 and Th17 subpopulations in cells from AAV patients (n=3) (regardless of whether they are in remission or active disease). Conclusions: Suppression of T-cell’s IFN-γ production is mainly mediated by IL-10 in HC, but in AAV patients this mechanism seeto be absent. The importance of these preliminary findings is unclear but signalling pathways downstream of IFN-γ are possible to target pharmacologically. Disclosures: Non