Fig 1: Effects of IL-10, IL-1β, IL-6 and BMP4 on HHLA2 expression in monocytes. A Monocytes from two donors (donor A; left and donor B; right) were treated with IL-10, IL-1β, IL-6 and BMP4, and HHLA2 expression was assessed over a time course using flow cytometry and measuring MFI and percentage of HHLA2 positivity in CD14+ cells (n = 2 healthy donors). B Relative HHLA2 mRNA levels were measured by qPCR at 18 h after cytokine treatment in monocytes. N = 3 replicates. C Monocytes were treated with cytokines (IL-10, IL-6, IL-1β) at 1, 2, 10, 25, 50, 100 ng/mL and BMP4 at 20, 40, 200, 500, 1000, 2000 ng/mL, and surface HHLA2 expression was measured at 48 h (n = 1 healthy donor)
Fig 2: Synergistic effects of IL-10 and IFN-γ on HHLA2 induction in monocytes. A HHLA2 expression in monocytes was measured at 48 h using flow cytometry after treatment individually or with the indicated combinations of IL-10, IL-6, IL-1β, BMP2, and BMP4. No significant difference in IL-10 alone (control) versus combination with IL-10. (n = 11 healthy donors). B Combination effect of IL-10, IL-6, IL-1β, and BMP4 with IFN-γ was evaluated using flow cytometry at 24 h. (n = 6 healthy donors). C Representative flow cytometry plots and histograms of HHLA2 expression at 24 h. Cytokine concentrations used in the combination experiments in A, B, and C are shown in Table 1. D Monocytes were treated with IL-10 (50 ng/mL) plus IFN-γ at 0, 0.2, 1, 2, 10, 20 ng/mL (top) or IFN-γ (20 ng/mL) plus IL-10 at 0, 0.5, 2.5, 5, 25, 50 ng/mL (bottom). HHLA2 expression was measured as MFI at 18 h by flow cytometry (n = 2 healthy donors)
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