miR-196a-5p-Rich Extracellular Vesicles from Trophoblasts Induce M1 Polarization of Macrophages in Recurrent Miscarriage

Figure 4

hnRNPA1 regulates miR-196a-5p packaging into EVs from trophoblasts. (a) The mRNA and protein expression levels of hnRNPA1 were evaluated by qPCR and western blotting in HTR-8 cells transfected with si-hnRNPA1 as well as the control siRNA. (b) qPCR analysis of miR-196a-5p expression levels in HTR-8 cell-derived EVs and the HTR-8 cells after transfection with si-hnRNPA1. (c) miRNA pull-down assays were performed using the nuclear, cytoplasmic, or EV lysates of HTR-8 cells. The expression of hnRNPA1 pulled by biotin-labeled miR-196a or mutant miR-196a was analyzed using western blotting. (d) RIP assays with hnRNPA1 antibody or IgG were performed using the lysates of HTR-8 cells or the EVs. (e) Cy3-miR-196a-5p and si-hnRNPA1 were cotransfected into HTR-8 cells for 48 h. Then, EVs were isolated and added to the PMA-pretreated THP-1 cells. Fluorescence microscopy was used to evaluate the red fluorescent signals in THP-1 cells (; ; ; ns: not significant). EV: extracellular vesicle; RIP: RNA immunoprecipitation.

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