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. 2012 Jun;136(2):218–230. doi: 10.1111/j.1365-2567.2012.03573.x

Figure 3.

Figure 3

Reverse transcription-PCR analysis of GARP and cytokine gene expression in expanded regulatory T cells (Tregs) and stimulation assay to analyse T-cell anergy. Adult peripheral blood (APB) and cord blood (CB) Tregs were expanded according to the protocol described in Fig. 1(a). Total RNA was isolated, reverse-transcribed and used for quantitative analysis. (a) Gene expression of GARP, transforming growth factor-β (TGF-β) and interleukin-10 (IL-10) of freshly isolated APB CD4+ CD25− T cells and Tregs from APB and CB was displayed as relative expression normalized to β-actin. Gene expression of GARP (b), IL-2 (c), interferon-γ (IFN-γ) (d), IL-10 (e) and TGF-β (f) of activated CD4+ CD25− T cells and expanded Tregs from APB and CB was displayed as relative expression normalized to β-actin. Values are given as the mean ± standard deviation (SD) of three to five independent experiments. **P < 0·01 compared with activated CD4+ CD25− T cells. (g) CFSE-labelled CD4+ CD25− T cells and expanded Tregs from APB and CB were stimulated with anti-CD3/anti-CD28 antibodies in the absence or presence of exogenous IL-2. Proliferation was determined by CFSE dilution on Day 4.