CircN4bp1 Facilitates Sepsis-Induced Acute Respiratory Distress Syndrome through Mediating Macrophage Polarization via the miR-138-5p/EZH2 Axis

Figure 6

CircN4bp1-miR-138-5p ceRNA modulates macrophage differentiation via targeted-regulating EZH2. (a) Schematic showing the 3UTR of EZH2 recognized by miR-138-5p. (b) miR-138-5p inhibited the expression of EZH2 mRNA in RAW264.7 macrophages as evidenced by qRT-PCR. (c) Luciferase reporter assay demonstrated that EZH2 was a target of miR-138-5p, but the rescue test by use of miR-138-5p inhibitors reversed its effect on EZH2. ( vs. miRNA inhibitor control group) (d) RAW264.7 cells was transfected with miR-138-5p mimic with/withoutcircN4bp1 lentivirus plasmids (circN4bp1-OE) or scrambled control and then exposed to either LPS (50 ng/ml) or IL-4 (10 ng/ml) for an additional 24 h. Representative western blot depicting raw264.7 cell lysates probed for EZH2, p-STAT1, PPAR-γ, and GAPDH. Expression levels of iNOS and Arg-1 were quantified by densitometry and normalized using GAPDH. (e) MH-S cells were transfected with miR-138-5p mimic with/withoutcircN4bp1 lentivirus plasmids (circN4bp1-OE) or scrambled control and then exposed to either LPS (50 ng/ml) or IL-4 (10 ng/ml) for an additional 24 h. Representative western blot depicting raw264.7 cell lysates probed for EZH2, p-STAT1, PPAR-γ, and GAPDH. Expression levels of iNOS and Arg-1 were quantified by densitometry and normalized using GAPDH. All data are expressed as . ( vs. NC group, # vs. miRNC + LPS stimulated group, ^@ vs. circN4bp1-OE + LPS group, & vs. miRNC +IL-4 stimulated group, and % vs. circN4bp1-OE + IL-4 group determined by one-way ANOVA for multiple group comparisons).

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