Figure 3.

HGF modulates monocyte-derived DC differentiation from GC patients into a regulatory phenotype in vitro. Freshly isolated peripheral monocytes from GC patients were cultured in the presence of GM-CSF and IL-4 with HGF (=HGF condition) or without HGF (=control condition) as described in the material and method section. After 6 days, cell phenotype was assessed by flow cytometry. In both conditions, monocyte-derived cells harbored DC features such as CD11c and HLA-DR co-expression. (A) CD11c and HLA-DR expression by monocyte-derived DCs without (left panel) or with HGF (right panel). Results are indicated as percentages of CD11 HLA-DR⁺ cells. Representative staining out of 45 independent experiments with similar results is shown. (B) Expression of CD80, CD83, HLA-DR, and CD86 in DCs generated without (black dashed curve) or with HGF (black full curve). Markers were set on their proper isotype control (light grey curve). A representative staining out of respectively 23, 45 and 29 independent experiments is shown (C) Median Fluorescence Intensity (MFI) of CD83 (n = 23), HLA-DR (n = 45) and CD86 (n = 29) by DCs generated in control condition or in HGF condition. * indicates a p-value ≤ 0.05 according to the Wilcoxon test. *** indicates a p-value ≤ 0.001 according to the Wilcoxon test. NS: non significant. After 6 days of culture, DCs were maturated with LPS 1 μg/mL or control PBS. After 24 h, cells were collected, and their phenotype was studied by flow cytometry. (D) Mean expression (MFI) of CD83 (upper panel, n = 6), HLA-DR (center panel, n = 6) and CD86 (lower panel, n = 6) by DCs generated in control or HGF condition with or without LPS. * indicates a p-value ≤ 0.05 according to the Wilcoxon test. Culture supernatants were collected and cytokine levels were assessed by ELISA. (E) TGF-β levels (upper panel, n = 18) and IL-10 levels (lower panel, n = 26) in supernatants of control or HGF-generated DCs. **** indicates a p-value ≤ 0.0001 according to the Wilcoxon test.