C-Type Natriuretic Peptide (CNP) Inhibition of Interferon-γ-Mediated Gene Expression in Human Endothelial Cells In Vitro

Figures

• 

  Figure 1. ICAM-1 expression in HUVEC after incubation with IFN-γ alone or in combinations with CNP for up to 72 h. (A) Representative flow cytometry histograms for ICAM-1 after incubation of HUVEC alone (Red) or with IFN-γ (Blue), CNP (Orange) or IFN-γ and CNP (Green) for the indicated times. (B–D) Mean Fluorescence Intensity (expressed as fold increase over MFI of untreated cells, which range from 820 to 3089; mean ± SEM) for ICAM-1 on untreated HUVEC or after 24 h (B), 48 h (C) or 72 h (D) treatment with IFN-γ alone or in combination with CNP; n = 3 HUVEC isolates (* p < 0.05, ** p < 0.01, significantly different from IFN-γ alone).

• 

  Figure 2. MHC class I expression in HUVEC after incubation with IFN-γ alone or in combinations with CNP for up to 72 h. (A) Representative flow cytometry histograms for MHC-I after incubation of HUVEC alone (Red) or with IFN-γ (Blue), CNP (Orange) or IFN-γ and CNP (Green) for the indicated times. (B–D) Mean Fluorescence Intensity (expressed as fold increase over MFI of untreated cells, which ranged from 2079 to 6549; mean ± SEM) for MHC-I on untreated HUVEC or after 24 h (B), 48 h (C) or 72 h (D) treatment with IFN-γ alone or in combination with CNP; n = 3 HUVEC isolates (* p < 0.05, ** p < 0.01, significantly different from IFN-γ alone).

• 

  Figure 3. MHC class II expression in HUVEC after incubation with IFN-γ alone or in combinations with CNP for up to 72 h. (A) Representative flow cytometry histograms for MHC-II after incubation of HUVEC alone (Red) or with IFN-γ (Blue), CNP (Orange) or IFN-γ and CNP (Green) for the indicated times. (B–D) Mean Fluorescence Intensity (expressed as fold increase over MFI of untreated cells, which ranged from 187 to 216; mean ± SEM) for MHC-II on untreated HUVEC or after 24 h (B), 48 h (C) or 72 h (D) treatment with IFN-γ alone or in combination with CNP; n = 3 HUVEC isolates.

• 

  Figure 4. cGMP mediated effects on cell surface receptor expression in HUVEC. (A) CNP-stimulated cGMP accumulation in HUVEC isolates, after 30 min stimulation with 100 nm CNP in the presence of 1 mm IBMX. Data shown are means ± SEM pooled from 6 independent isolates (n = 6), each performed in duplicate (* p = 0.015, significantly different from Basal). (B–D) Effect of 8-bromo-cGMP on IFN-γ mediated ICAM-1 (B), MHC-I (C), and MHC-II (D) expressed as fold increase over MFI of untreated cells expression after 48hr treatment (which ranged from 572 to 819, 1313 to 2012, 176 to 230, for ICAM-1, MHC-I and MHC-II, respectively). n = 3 HUVEC isolates (* p < 0.05, *** p < 0.001, significantly different from IFN-γ alone).

• 

  Figure 5. CNP downregulates IFN-γ-mediated tryptophan metabolism in HUVEC. (A) Representative chromatogram of kynurenine production following IFN-γ treatment. (B) Kynurenine production from untreated HUVEC or after 24 h and 48 h treatment with IFN-γ alone or in combination with CNP. Data shown are representative from a single HUVEC isolate, expressed as % IFN-γ -stimulated kyunurenine production), and performed in duplicate. (** p < 0.01, significantly different from IFN-γ alone).