Enhanced Expression of miR-34a Enhances Escherichia coli Lipopolysaccharide-Mediated Endometritis by Targeting LGR4 to Activate the NF-κB Pathway

Figure 11

IL-1β suppresses LGR4 expression by enhancing miR-34a. (a) BENDs were treated with recombinant IL-1β (0, 1, and 10 ng/ml) for 12 h, and then miR-34a transcription levels were analysed by qRT-PCR. (b) Cells were transfected with miR-34a inhibitor or the negative control and then stimulated with recombinant IL-1β (10 ng/ml). The relative expression of miR-34a was normalized to U6 snRNA. (c) qRT-PCR assay was used to determine the LGR4 mRNA. (d) The expression of LGR4 and p-p65 was determined using western blotting. (e) Grey values of the indicated proteins were measured by ImageJ software. (f) Translocation of the p65 subunit from the cytoplasm into the nucleus was evaluated by immunofluorescence (). Blue spots represent cell nuclei, and red spots represent p-p65 staining; . (g) The fluorescence intensity of p-p65. The integrated option density (IOD) of DAPI was used as an internal control. The IOD and area of cells were measured by IPP 6.0 software, and the fluorescence intensity of p-p65 was expressed as IOD/area. Data are expressed as the of three independent experiments (). versus the inhibitor negative control group (untreated with IL-1β); ^# compared with the inhibitor group (untreated with IL-1β); ^@ compared with the inhibitor group. Comparisons among multiple groups were analysed by one-way analysis of variance with the Bonferroni posttest.

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