Abstract
Purpose:
Endometritis is a female reproductive disease that affects the cattle industries development and microRNAs (miRNAs) play a pivotal role and critical regulators of the innate immune response in varieties of diseases. The present study intends to investigate the regulatory role of miR-24-3p in the innate immune response involved in endometritis and evaluate its therapeutic potential.
Methods:
Whole mice uteri and bovine endometrial epithelial cells (BEECs) were separately stimulated with LPS. The BEECs were also transfected with miR-24-3p mimic and negative control; siTRAF6 and siNC; pcDNA3.1 empty and pcDNA3.1(+)TRAF6 separately with LPS stimulation. The expression levels of miR-24-3p and TRAF6 were measured via quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. LPS-induced inflammatory response assessed by inflammatory cytokines secretion and expression via ELISA and qRT-PCR. Bioinformatics analysis and luciferase reporter assay validated the interaction between miR-24-3p and TRAF6. The activation of the NF-ĸB/MAPK pathway and p65 phosphorylation was investigated by Western blot and immunofluorescence assay, respectively.
Results:
The expression of miR-24-3p was decreased, and TRAF6 was elevated with an increased level of pro-inflammatory cytokines in LPS-treated BEECs and mice uterus. The overexpression of miR-24-3p suppressed LPS-induced secretion of inflammatory cytokines (IL-1β, IL-6, IL-8 and TNF-α) and deactivation of NF-ĸB/MAPK pathways. The downregulation of TRAF6 inhibited LPS-induced inflammatory response in BEECs. TRAF6 is validated as a target of miR-24-3p, and miR-24-3p reversed the overexpression of cloned TRAF6 on inflammation response in BEECs.
Conclusion:
Our findings demonstrate that the overexpression of miR-24-3p attenuates endometrial inflammation and the expression of pro-inflammatory mediators via suppressing TRAF6. Therefore, modulating the pathogenesis of endometritis and possibly, a therapeutic potential against endometritis.
Keywords:
NF-ĸB/MAPK; TRAF6; endometritis; inflammation; miR-24-3p.