Fig. 2

HERA-GITRL enhances human T cell activation following stimulation in vitro. a, b Naïve CD4+ T cells were isolated from the peripheral blood of healthy volunteers, labeled with CFSE and stimulated with anti-CD3 antibody (or medium) in the presence of HERA-GITRL or vehicle control (PBS), as indicated. On day five, T cells were harvested and examined by FCM. Representative histograms (a) (HERA-GITRL at 100 ng/mL) and quantified data (b) (mean ± S.D.) are shown. Although not labeled, all anti-CD3 treated groups were significantly different from the medium alone and HERA-GITRL alone groups. c Naïve CD4+ T cells were stimulated with anti-CD3 antibody (or medium) in the presence of HERA-GITRL (100 ng/mL) or vehicle control (PBS), as indicated. On day five, T cells were harvested, stained for surface expression of CD45RO and CD45RA and examined by FCM. d Naïve CD4+ T cells were stimulated with anti-CD3 antibody (or medium) in the presence of HERA-GITRL (100 ng/mL) or vehicle control (PBS), as indicated. On day six, T cells were stained for intracellular expression of IFNγ and TNFα and examined by FCM. e Treg cells and Teff cells isolated from human PBMCs were cultured together at different ratios (Treg:Teff) and stimulated with anti-CD3 and anti-CD28 antibodies in the presence of IL-2 and ATRA or left unstimulated (resting). HERA-GITRL (10 or 100 ng/mL) or vehicle control (PBS) was added immediately. On day five, T cells were harvested and CTV expression by Teff cells was examined by FCM. Numbers indicate the percentage of cells within the defined region. Representative plots, gated on live single cells, from the median sample of triplicates from at least three independent experiments are shown (a, c, d). The p values represent comparisons between samples using a one-way (b) or two-way (e) ANOVA plus post hoc Bonferroni multiple comparisons test. *p < 0.05, **p < 0.01, ****p < 0.0001