Article Text

A1.7 The regulation of human in vitro TH17 cell differentiation
  1. E Baricza1,
  2. E Lajkó1,
  3. L Kőhidai1,
  4. B Molnár-Érsek1,
  5. N Marton1,
  6. E Buzás1,
  7. Gy Nagy1,2
  1. 1Department of Genetics-, Cell- and Immunobiology, Semmelweis University, Budapest, Hungary
  2. 2Department of Rheumatology, Semmelweis University, Budapest, Hungary


Background and objectives Th17 cells produce several inflammatory cytokines, such as interleukin (IL)-17A, -17F, -21, -22, and tumour necrosis factor-α and play an important role in the regulation of inflammation. We studied in vitro Th17 cell differentiation of healthy donors and patients with rheumatoid arthritis (RA).

Materials and methods CD4+ T cells were negatively separated by magnetic method from peripheral blood mononuclear cells (PBMC) then CD45RO+ and CD45RO- cells were separated. The cells were treated for 5–10 days with anti-CD3 and anti-CD28 antibodies and with TGFβ (2.5 ng/ml) and IL-6 (25 ng/ml) or with IL-6 (25 ng/ml), IL-1 (10 ng/ml) and IL-23 (10 ng/ml) cytokines and with anti-IL-4 (10 μg/ml) and anti-IFNγ (10 μg/ml) blocking antibodies. The IL-17 and IL-22 production were measured by ELISPOT and ELISA, the RORγt expression was measured by real-time PCR and by Western blot methods. Cell viability was monitored by Trypan blue staining, by Annexin V binding and by an impendance-based cell analyser.

Results Anti-CD3/CD28 activation increased the IL-17 production, but did not alter the RORγt expression. The anti-CD3/CD28 activation and TGFβ, IL-6, and IL-1 cytokine induced RORγt expression of the CD4+ T cells was further increased by the anti-IL-4 and anti-IFNγ antibody treatment. The IL-22 production of CD4+ T cells was significantly reduced in the RORγt producing CD4+ cells. The CD45RO+ cells expressed high level of RORγt and produced high amount of IL-17 without stimulation or cytokine treatment. The RORγt expression of the CD4+RO- T cells increased severalfold upon stimulation and cytokine treatment. The differentiated Th17 cells of patients with RA produced more IL-17 than those of the healthy donors’. The applied treatments did not change the viability of the cells.

Conclusions Our results suggest that the IL-17 and IL-22 production are differently regulated during Th17 differentiation. The Th17 cell differentiation is most effective from CD45RO- naive T cells.

Statistics from

Request permissions

If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.