Figure 4.

Effect of 1,25(OH)₂D₃ on IL-1β or TNF-α or IFN-γ induced indolemaine-2,3-dioxygenase-1 (IDO-1) expression in hPDLSCs. Primary hPDLSCs were treated with 10 ng/mL TNF-α or 5 ng/mL IL-1β or 100 ng/mL IFN-γ in the absence or presence of 1,25(OH)₂D₃ (0.01–100 nM). IDO-1 gene expression was analyzed by calculating the n-fold expression compared to unstimulated cells (n-fold expression = 1) using qPCR. GAPDH served as endogenous reference. qPCR data were received from six independent experiments with hPDLSCs from six different individuals (a). Intracellular IDO-1 staining and flow cytometry analysis were performed to determine m.f.i. of IDO-1 positive cells. Data were received from five independent experiments with hPDLSCs from five different individuals (b). IDO-1 enzymatic activity was verified by measuring L-kynurenine concentrations photometrically in conditioned media. L-kynurenine concentrations were normalized to total protein amounts followed by subtraction of the L-kynurenine concentrations from control group from each sample. Data were received from five independent experiments with hPDLSCs isolated from five different individuals (c). (d) shows the extent of the 1,25(OH)₂D₃ effect regarding differently-induced IDO-1 production in hPDLSCs (expressed in % of corresponding cytokine treatment). All data are presented as mean ± S.E.M. (a–c): * significantly higher (p < 0.05) compared to unstimulated hPDLSCs. # significantly lower (p < 0.05) compared to appropriate cytokine treated hPDLSCs treated in the absence of 1,25(OH)₂D₃. (d): * significantly different (p < 0.05) compared between groups as indicated.