Overexpression of miR-221-3p effecting cell proliferation, apoptosis and inflammation by targeting Toll-like receptors 4 in propofol-induced rat astrocytes

Abstract

IntroductionGrowing evidence indicated that propofol has neurotoxic effects on the brains of developing rodents, leading to neuronal cell death, neurodegeneration, and brain injury.Material and methodsEctopic miR-221-3p was transfected into rat astrocytes, and Cell Counting Kit-8 assay and flow cytometry were performed to evaluate cell growth and apoptosis. The mRNA levels of Toll-like receptors 4 (TLR4), nuclear factor kappa B, interleukin-6, interleukin-1β, myeloid differentiation primary response 88 (MyD88), caspase-3, caspase-12, STAT3, and GRP78 were detected using quantitative real-time polymerase chain reaction. The proteins of TLR4 and MyD88 were determined using Western blotting. The association between miR-221-3p and TLR4 was measured using Dual-Luciferase Reporter Assay (Promega Corporation, Wisconsin, USA). Then, siTLR4 was transfected with 293T cells to study the role of TLR4 in astrocytes with propofol treatment.ResultsThe miR-221-3p expression in rat astrocytes was markedly suppressed by propofol treatment. The miR-221-3p mimics transfection in propofol-treated astrocytes effectively reduced the suppressive effect of propofol on astrocyte growth, repressed the propofol-induced apoptosis in rat astrocytes, and decreased the cell number during the G2–M phase. The expression of MyD88 and TLR4 was induced by propofol, whereas the transfection of miR-221-3p mimics dramatically reduced these genes expression at the mRNA and protein expression. After that, TLR4 was found to be target of miR-221-3p using Dual-Luciferase Reporter Assay. Furthermore, knockdown of TLR4 could suppress the apoptosis rate in propofol-treated astrocytes.ConclusionsThis study revealed that miR-221-3p might prevent astrocytes from propofol-induced damage by targeting TLR4.