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AB0023 The Regulation of Human In Vitro Th17 Cell Differentiation by Cytokines
  1. E. Baricza,
  2. E. Lajkό,
  3. L. Kőhidai,
  4. B. Molnár-Έrsek,
  5. N. Marton,
  6. E.I. Buzás,
  7. G. Nagy
  1. Department of Geneticts-, Cell- and Immunolbiology, Semmelweis University, Budapest, Hungary

Abstract

Background Th17 cells have a central role in inflammation via their pro-inflammatory cytokine production, such as interleukin (IL)-17A, -17F, -21, -22, tumor necrosis factor-α and the expression of master regulator transcription factor RORc. The Th17 cells have essential role in many autoimmune diseases via induction of the development of inflammation.

Objectives We studied the in vitro Th17 cell differentiation of healthy donors and patients with rheumatoid arthritis (RA).

Methods CD45RO- and CD45RO+ cells were isolated from peripheral blood mononuclear cells with a two-step negative magnetic separation. The cells were activated with anti-CD3 and anti-CD28 antibodies and treated with TGFβ (2.5 ng/ml), IL-6 (25 ng/ml), IL-1β (10 ng/ml) and IL-23 (10 ng/ml) cytokines and with an anti-IL-4 (10 μg/ml) blocking antibody. After 5 and 10 days IL-17 and IL-22 productions were measured by ELISPOT and ELISA; the RORc and Tbet expression were measured by quantitative real-time PCR methods. Cell viability was monitored by impendance-based cell analyzer (CASY-TT).

Results Although anti-CD3/CD28 activation increased the IL-17 and IL-22 productions, it did not alter the RORc expression of the CD45RO- cells of healthy donors. The IL-22 production and the Tbet expression of the RORc producing cells were inhibited by TGFβ and stimulated by IL-23 treatment. The CD45RO+ cells of healthy donors expressed higher level of RORc, T-bet and IL17 without stimulation or cytokine treatment compared to the CD45RO- cells. However, there was no difference between the RORc and Tbet expression of the freshly separated CD45RO+ and CD45RO- cells of RA patients. IL-23 treatment increased both of IL-17 and IL-22 cytokine production of the differentiated CD45RO- cells.

Conclusions IL-17 and IL-22 production are differently regulated during human Th17 differentiation. Our data suggest that the increased baseline RORc expression of CD45RO- cells may contribute to the accelerated Th17 differentiation in RA.

Disclosure of Interest None declared

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