Neuroprotective Effects of microRNA-140-5p on Ischemic Stroke in Mice via its Regulation on the TLR4/NF-κB Axis

Abstract

Background and aim: Ischemic stroke is one of the main causes of death worldwide and permanent global disability. On the basis of existing literature data, we carried out studies in an effort to explore how miR-140-5p affects ischemic stroke and whether the mechanism relates to toll-like receptor-4 (TLR4) and nuclear factor-kappa B (NF-κB).Methods: Middle cerebral artery occlusion (MCAO) was employed to establish a mouse model of ischemic stroke in vivo, while primary neurons were exposed to oxygen-glucose deprivation (OGD) to set up an ischemic stroke model in vitro. RT-qPCR was performed to detect the miR-140-5p expression and Western blot was employed to detect the expression TLR4, NF-κB, and apoptosis-related factors. Then, based gain-function of experiments using miR-140-5p mimic and TLR4 overexpression plasmid, neurological function score, TTC staining, TUNEL staining, as well as flow cytometry were carried out to evaluate the effects of miR-140-5p and TLR4 on MCAO mice and OGD neurons. Meanwhile, dual-luciferase reporter assay was used to validate the relationship between miR-140-5p and TLR4.Results: miR-140-5p expressed at a low level and TLR4 at a high level in ischemic stroke. It was verified that miR-140-5p targeted TLR4 and downregulated its expression. miR-140-5p overexpression was observed to inhibit the apoptosis of neurons under OGD exposure and restrain the progression of ischemic stroke, while TLR4 overexpression promoted the apoptosis and disease progression. Besides, miR-140-5p overexpression led to a decrease in NF-κB protein level, which was increased by TLR4 overexpression. Conclusion: In conclusion, from our data we conclude that miR-140-5p overexpression may be instrumental for the therapeutic targeting of ischemic stroke by alleviating neuron injury with the involvement of TLR4/NF-κB axis.