Biomedicine & Pharmacotherapy (Mar 2022)
Endometriosis derived exosomal miR-301a-3p mediates macrophage polarization via regulating PTEN-PI3K axis
Abstract
This study aimed to explore the effects of endometriosis (EMS)-derived exosomes and miR-301a-3p on the polarization of macrophages and investigate the involved molecular mechanism. The exosomes were isolated from ectopic endometrial tissues of EMS patients and normal human serum (NHS). Results of transmission electron microscope and Nanoparticle Tracking Analysis showed that both EMS-exosomes and NHS-exosomes are about 80 nm microvesicles. Exosomal markers CD63 and TSG101 were abundantly expressed in both EMS-exosomes and NHS-exosomes. No negative marker Calnexin was detected in NHS-exosomes. A small amount of Calnexin was detected in EMS-exosomes. THP-1 cells differentiatee to macrophages by incubating with phorbol-12-myristate-13-acetate. Effects of the exosomes on the phagocytosis and polarization of macrophages were evaluated by PKH26 fluorescent labeling and flow cytometry, respectively. Compared with the NHS-exosomes group, the phagocytic capacity of macrophages was reduced and the polarization of macrophages to M2 macrophages was promoted after EMS-exosomes treatment. Results of western blot showed that compared with the NHS-exosomes group, the EMS-exosomes treatment significantly up-regulated the expression of phosphatidylinositol 3-kinase (PI3K) and down-regulated the expression of phosphatase and tensin homologue deleted on chromosome 10 (PTEN). miR-301a-3p mimic, negative control (NC) mimic, miR-301a-3p inhibitor and NC inhibitor were transfected into cells. Transfection efficiency was confirmed by RT-qPCR. Effects of the miR-301a-3p expression on the macrophages polarization and the expression of Arg-1, PTEN and PI3K in the macrophages were evaluated by flow cytometry and western blot, respectively. miR-301a-3p overexpression significantly enhanced the ability of EMS-exosomes-inducing M2 transformation of macrophages, promoted the expression of Arg-1 and PI3K, and inhibited the PTEN expression. miR-301a-3p inhibitor significantly reduced the expression of Arg-1 and PI3K and promoted the PTEN expression. In conclusion, EMS derived exosomal miR-301a-3p mediated macrophage polarization via regulating PTEN-PI3K axis.
