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. 2022 Dec 1;11(23):3878. doi: 10.3390/cells11233878

Figure 4.

Figure 4

Enhancement of HIF-1α-related proteins after normoxic HIF-1α stabilization. (a) Fold change of the VEGF-A protein level in the cell culture medium of HCAECs incubated with cell culture medium (control group), 50 ng/mL each of TNF-α, INF-γ, IL-1β, and cocktail 1 (TNF-α, INF-γ, and IL-1β) for 12 h or 50 ng/mL each of IL-8, M-CSF, IGF-I, and cocktail 2 (IL-8, M-CSF, and IGF-I) for 18. Normoxic HIF-1α stabilization only led to an increase in the VEGF-A protein level in the cell culture medium of HCAECs treated with TNF-α, INF-γ, IL-1β, and cocktail 1 (TNF-α, INF-γ, and IL-1β). The data are represented as the mean ± SEM of two independent experiments (N = 2), ns = not significant, * p value ≤ 0.05 and ** p value ≤ 0.01 vs. medium. (b) Immunoblots and graphical representation of the fold change of the VEGF-A protein level in the total cell lysates of HCAECs incubated with cell culture medium (control group), 50 ng/mL each of TNF-α, INF-γ, and IL-1β for 12 h or 50 ng/mL each of IL-8, M-CSF, and IGF-I for 18 h. The data show an increase in the VEGF-A protein level inside of HCAECs upon all treatments. The data are represented as the mean ± SEM of three independent experiments (N = 3), ns = not significant, * p value ≤ 0.05, ** p value ≤ 0.01, and*** p value ≤ 0.001 vs. medium. (c–j) After the incubation of HCAECs with the medium (control group), 50 ng/mL each of TNF-α, INF-γ, IL-1β, and cocktail 1 (TNF-α, INF-γ, and IL-1β) for 12 h or 50 ng/mL each of IL-8, M-CSF, IGF-I, and cocktail 2 (IL-8, M-CSF, and IGF-I) for 18, the cell culture medium of HCAECs was analyzed to measure the protein fold change of (c) IL-6, (d) IL-9, (e) G-CSF, (f) CCL5, (g) CCL2, (h) PDGF-BB, (i) IL-4, and (j) CXCL10. The data are represented as the mean ± SEM of two independent experiments (N = 2), ns = not significant, * p value ≤ 0.05, ** p value ≤ 0.01, *** p value ≤ 0.001, and **** p value ≤ 0.0001 vs. medium.