miR-23a-3p involved in drug resistance through directly targeting influx drug transporter Organic anion transporting polypeptide 2
miR-23a-3p involved in drug resistance through directly targeting influx drug transporter Organic anion transporting polypeptide 2
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miR-23a-3p通过直接靶向流入药物转运蛋白参与耐药性有机阴离子转运多肽2
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通讯作者：
郭艺
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作者：
郭艺
摘要：目的：药物转运蛋白通过影响抗癫痫药物（AEDs）的脑内转运参与耐药性癫痫患者的耐药性。药物转运蛋白的表达与microRNA相关。我们前期研究发现顽固性癫痫患者血液中miR-23a-3p水平升高。另外，流入的药物转运蛋白有机阴离子转运多肽2(Oatp2)参与最常用的AEDs丙戊酸(VPA)的脑内转运，反复癫痫发作导致Oatp2表达下降。 miR-23a-3p在Oatp2表达和耐药性发展中的作用尚未确定。在此，我们旨在通过体内和体外研究确定miR-23a-3p在VPA抵抗性癫痫中的潜在作用。方法-在成年Wistar大鼠中锂-毛果芸香碱诱导癫痫持续状态（SE）后引发癫痫，然后进行VPA治疗，选择大鼠耐 VPA。采用免疫组化和逆转录聚合酶链反应（RT-PCR）检测miR-23a-3p的表达。将miR-23a-3p抑制剂注射到VPA耐药大鼠脑内，进行组织学染色和Morris水迷宫实验，以确定这些大鼠的脑损伤和学习记忆功能。随后，在脑微血管内皮细胞（BMEC）中进行双荧光素酶报告基因测定和 VPA 摄取测定，以研究 miR-23a-3p 的潜在作用机制。结果-我们的结果表明，与对照大鼠相比，VPA 耐药组的 miR-23a-3p 升高。脑内注射 miR-23a-3p 抑制剂可减轻 VPA 耐药大鼠的脑损伤以及相关的学习和记忆功能缺陷。进一步研究表明，Oatp2是miR-23a-3p的直接靶标，并且在大脑和BMEC中受到miR-23a-3p的反向调节。此外，我们证明 miR-23a-3p 通过调节 Oatp2 表达来减少 BMEC 中 VPA 的摄取。结论-miR-23a-3p通过直接靶向流入药物转运蛋白Oatp2参与癫痫的VPA抵抗，表明miR-23a-3p可能成为难治性癫痫的潜在治疗靶点。
Abstract.Objective-Drug transporters involve in drug resistance of individuals with drug-resistant epilepsy , by influencing intracerebral transport of antiepileptic drugs(AEDs) . The expression of drug transporters are associated with microRNAs.We previously researches revealed miR-23a-3p level elevated in the blood of patients with intractable epilepsy.Additionally，the influx drug transporter organic anion transport polypeptide 2(Oatp2) involve in the intracerebral transport of valproic acid (VPA),the most common used AEDs;repeated seizures lead to the decreased expression of Oatp2.However,the role for miR-23a-3p in the expression of Oatp2 and in the development of drug resistance has not been established.Herein, we aimed to determine the potential role of miR-23a-3p in VPA-resistant epilepsy through in vivo and in vitro study..Methods-Epilepsy was elicited after status epilepticus (SE) induced by lithium-pilocarpine in adult Wistar rats,followed by a VPA treatment for selecting the rats with VPA-resistant. Expression of miR-23a-3p was detected by immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR).A miR-23a-3p inhibitor was intracerebrally injected into the VPA-resistant rats, and histological staining and Morris Water Maze test were performed to determine the brain damage and learning/memory function in these rats. Subsequently, a dual-luciferase reporter assay and a VPA-uptake assay were performed in the brain microvascular endothelial cells (BMECs)to investigate the underlying mechanism of action of miR-23a-3p. .Results-Our results indicated that compared to control rats, miR-23a-3p elevated in the VPA-resistant group. The intracerebral injection of a miR-23a-3p inhibitor reduced brain damage and the associated deficits in learning and memory function in rats with VPA-resistant rats. Further investigations indicated that the Oatp2 was the direct target of miR-23a-3p, and it was inversely regulated by miR-23a-3p in the brain and BMECs. Furthermore, we demonstrated that miR-23a-3p reduced VPA uptake in the BMECs by regulating Oatp2 expression. .Conclusions-miR-23a-3p involved in VPA resistance of epilepsy through directly targeting influx drug transporter Oatp2,indicating that miR-23a-3p could be a potential therapy target for intractable epilepsy.