微小 RNA-542-3p 通过 Toll 样受体 4/核因子-κB 信号通路调节大鼠癫痫中的 P-糖蛋白表达。

MicroRNA-542-3p Regulates P-glycoprotein Expression in Rat Epilepsy via the Toll-like Receptor 4/Nuclear Factor-kappaB Signaling Pathway.

机构信息

Department of Neurology, Huzhou Central Hospital, Huzhou City, Zhejiang Province, 313000, China.

College of Basic Medicine, Nanjing Medical University, Nanjing City, Jiangsu Province, 211166, China.

出版信息

Curr Neurovasc Res. 2019;16(5):433-440. doi: 10.2174/1567202616666191023160201.

DOI:10.2174/1567202616666191023160201

PMID:31702493

Abstract

BACKGROUND

The function of microRNA-542-3p (miR-542-3p) in rat epilepsy is still unclear.

METHODS

The levels of miR-542-3p and toll-like receptor 4 (TLR4) were determined through quantitative real-time PCR. The protein levels were examined via the western blot analysis. The relationship between miR-542-3p and TLR4 was confirmed through luciferase assay. Pathological changes were analyzed via Hematoxylin-eosin (HE) and Nissl staining.

RESULTS

The rats and hippocampal cells were treated with kainic acid (KA) in vivo and in vitro. miR-542-3p was low in KA-treated rats, hippocampal cells and cerebrospinal fluid of patients with epilepsy. Further functional analysis showed that miR-542-3p overexpression inhibited KAinduced average seizure frequency, damage of hippocampal neuron and cell apoptosis, leading to the alleviation of the brain injury in epilepsy rats. miR-542-3p was determined to downregulate TLR4 expression. The relationship between miR-542-3p and TLR4 was confirmed. TLR4 knockdown reduced KA-induced nuclear factor-kappa B p65 (NF-κB p65), multidrug resistance 1 (MDR1), P-glycoprotein (P-gp) and apoptosis-associated protein levels. Further, for NF-κB p65, MDR1, P-gp and apoptosis-associated protein levels detection, miR-542-3p mimic showed a suppressive effect on these KA-induced protein levels, whereas TLR4 overexpression ameliorated the miR-542-3p-induced these protein levels in KA-treated epilepsy rats.

CONCLUSION

We identified that miR-542-3p attenuated seizure-induced brain injury and the expression of P-gp in epilepsy rats through inhibiting TLR4/NF-κB signaling pathway, which might contribute to improved epilepsy therapy.

摘要

背景

微小 RNA-542-3p（miR-542-3p）在大鼠癫痫中的作用尚不清楚。

方法

通过实时定量 PCR 测定 miR-542-3p 和 Toll 样受体 4（TLR4）的水平。通过 Western blot 分析检测蛋白水平。通过荧光素酶测定法证实 miR-542-3p 和 TLR4 之间的关系。通过苏木精-伊红（HE）和尼氏染色分析病理变化。

结果

在体内和体外使用海人酸（KA）处理大鼠和海马细胞。KA 处理的大鼠、海马细胞和癫痫患者脑脊液中的 miR-542-3p 水平较低。进一步的功能分析表明，miR-542-3p 过表达抑制 KA 诱导的平均发作频率、海马神经元损伤和细胞凋亡，从而减轻癫痫大鼠的脑损伤。miR-542-3p 被确定下调 TLR4 表达。证实了 miR-542-3p 和 TLR4 之间的关系。TLR4 敲低降低了 KA 诱导的核因子-κB p65（NF-κB p65）、多药耐药 1（MDR1）、P-糖蛋白（P-gp）和凋亡相关蛋白水平。此外，对于 NF-κB p65、MDR1、P-gp 和凋亡相关蛋白水平的检测，miR-542-3p 模拟物对这些 KA 诱导的蛋白水平具有抑制作用，而 TLR4 过表达改善了 KA 处理的癫痫大鼠中 miR-542-3p 诱导的这些蛋白水平。