BACKGROUND Some long non-coding RNAs (lncRNAs) have been suggested to play critical roles in Parkinson's disease (PD) pathogenesis, including nuclear enriched abundant transcript 1 (NEAT1). The purpose of this study was to further elucidate the molecular mechanism of NEAT1 in PD. METHODS The expression levels of NEAT1, miR-212-5p and RAB3A-interacting protein (RAB3IP) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability and apoptosis were evaluated by Cell Counting Kit-8 (CCK-8) assay and flow cytometry analysis, respectively. Western blot analysis was applied to detect the protein expression of IL-1β, TNF-α and RAB3IP. The LDH activity, ROS generation and SOD activity were measured by Lactate LDH activity assay kit, ROS assay kit, and SOD activity assay kit, respectively. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were performed to verify the relationship between miR-212-5p and NEAT1 or mRNA of RAB3IP. 1-methyl-4-phenylpyridinium ion (MPP+)-treated SK-N-SH cells were used as an in vitro model of PD. RESULTS NEAT1 and RAB3IP were upregulated while miR-212-5p was downregulated in SK-N-SH cells treated with MPP+. NEAT1 knockdown or miR-212-5p overexpression inhibited MPP+-induced apoptosis, inflammation and cytotoxicity in SK-N-SH cells. Moreover, miR-212-5p was a direct target of NEAT1 and its downregulation reversed the effects caused by NEAT1 knockdown in MPP+-induced SK-N-SH cells. Furthermore, RAB3IP was a downstream target of miR-212-5p and its overexpression attenuated the effects of miR-212-5p restoration in MPP+-induced SK-N-SH cells. Besides, NEAT1 acted as a molecular sponge of miR-212-5p to regulate RAB3IP expression. CONCLUSION NEAT1 knockdown suppressed MPP+-induced apoptosis, inflammation and cytotoxicity in SK-N-SH cells through regulating miR-212-5p and RAB3IP expression, providing a possible therapeutic strategy for PD patients.

中文翻译：

---

长时间的非编码RNA NEAT1敲低可以通过调节miR-212-5p / RAB3IP轴来抑制MPP +诱导的SK-N-SH细胞凋亡，炎症和细胞毒性。

背景技术已经提出一些长的非编码RNA（lncRNA）在帕金森氏病（PD）发病机理中起关键作用，包括富核丰富的转录本1（NEAT1）。这项研究的目的是进一步阐明PD中NEAT1的分子机制。方法采用定量实时聚合酶链反应（qRT-PCR）测定NEAT1，miR-212-5p和RAB3A相互作用蛋白（RAB3IP）的表达水平。通过细胞计数试剂盒8（CCK-8）分析和流式细胞仪分析分别评估细胞活力和凋亡。免疫印迹分析用于检测IL-1β，TNF-α和RAB3IP的蛋白表达。通过乳酸LDH活性测定试剂盒，ROS测定试剂盒和SOD活性测定试剂盒分别测定LDH活性，ROS生成和SOD活性。进行双荧光素酶报告基因分析和RNA免疫沉淀（RIP）分析，以验证miR-212-5p与NEAT1或RAB3IP mRNA之间的关系。1-甲基-4-苯基吡啶鎓离子（MPP +）处理的SK-N-SH细胞用作PD的体外模型。结果在用MPP +处理的SK-N-SH细胞中，NEAT1和RAB3IP被上调，而miR-212-5p被下调。NEAT1敲低或miR-212-5p过表达抑制了MPP +诱导的SK-N-SH细胞凋亡，炎症和细胞毒性。而且，miR-212-5p是NEAT1的直接靶标，其下调逆转了MPP +诱导的SK-N-SH细胞中NEAT1敲低引起的效应。此外，RAB3IP是miR-212-5p的下游靶标，它的过表达减弱了MPP +诱导的SK-N-SH细胞中miR-212-5p恢复的作用。除了，NEAT1充当miR-212-5p的分子海绵来调节RAB3IP表达。结论NEAT1敲低可通过调节miR-212-5p和RAB3IP表达抑制MPP +诱导的SK-N-SH细胞凋亡，炎症和细胞毒性，为PD患者提供了可能的治疗策略。