Fig. 3: Zn²⁺ and Ca²⁺ induce differentiation into Th1 cells.

A PBMC were stimulated at the same time with pyrithione and thapsigargin at the indicated concentrations. After 48 h, released IL-2 was measured by ELISA. B PBMC were stimulated with thapsigargin in Zn²⁺-adequate (ZA) or Zn²⁺-deficient (ZD) medium and the IL-2 production was measured after 48 h by ELISA. C PBMC were stimulated as described in (A) and the IFN-γ mRNA expression was examined by qPCR after 3 h and normalized to PBGD expression. D PBMC were stimulated as described in (A) and after 48 h, IFN-γ in the supernatant was measured by ELISA. E PBMC were stimulated as described in (A) and the protein expression of transcription factor T-bet was examined by western blot 48 h after stimulation and normalized to β-actin. A representative western blot is shown. F PBMC were stimulated as described in (A) and the IRF-1 expression was examined by qPCR after 3 h and relative mRNA expression was determined by 2^−ΔΔCt method. Data are presented as mean + SEM with n = 8 (A), n = 12 (B), n = 10 (C), n = 9 (D), n = 14 (E) and n = 12 (F) experiments. Statistical significance was determined by one-way ANOVA with Tukey’s multiple comparisons test (A, C–F) or with Friedman test with Dunn’s multiple comparison test (B). Significantly different results (p < 0.05) have no common identification letter.