Astrocytes release exosomes that regulate neuronal cell function. 1-methyl-4-phenylpyridinium (MPP+) is a well-known neurotoxin used to induce cell death in in vitro Parkinson's disease models, and microRNA (miRNA) transferred by released exosomes can regulate its mechanisms. Here, we demonstrated that exosomes released from normal astrocytes (ADEXs), but not exosomes derived from MPP+-stimulated astrocytes (MPP+-ADEXs), significantly attenuate MPP+-induced cell death in SH-SY5Y cells and primary mesencephalic dopaminergic neuron cultures, and reduce expression of mitogen-activated protein kinase kinase 4 (MKK4), an important upstream kinase in the c-Jun N-terminal kinase cell death pathway. Similar neuroprotective results were obtained from primary hippocampal neuron cultures, an in vitro glutamate excitotoxicity model. Through small-RNA sequencing of exosomal miRNA, we identified miR-200a-3p as the most down-regulated miRNA expressed in MPP+-ADEXs. miRNA target analysis and reporter assay confirmed that miR-200a-3p targets MKK4 through binding to two independent sites on the 3'-UTR of Map2k4/MKK4 mRNA. Treatment with miR-200a-3p mimic suppressed both MKK4 mRNA and protein expressions, and attenuated cell death in MPP+-treated SH-SY5Y cells and glutamate-treated hippocampal neuron cultures. Our results suggest that normal astrocytes release miR-200a-3p which exhibits a neuroprotective effect through down-regulation of MKK4.

中文翻译：

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星形胶质细胞来源的外泌体微RNA miR-200a-3p通过下调MKK4防止MPP +诱导的凋亡细胞死亡。

星形胶质细胞释放调节神经元细胞功能的外来体。1-甲基-4-苯基吡啶鎓（MPP +）是一种众所周知的神经毒素，用于在体外帕金森氏病模型中诱导细胞死亡，释放的外泌体转移的microRNA（miRNA）可以调节其机制。在这里，我们证明了正常星形胶质细胞（ADEXs）释放的外泌体，而不是MPP +刺激的星形胶质细胞（MPP + -ADEXs）释放的外泌体显着减弱了SHP-SY5Y细胞和原发性中脑多巴胺能神经元培养物中MPP +诱导的细胞死亡，并减少了有丝分裂原活化的蛋白激酶激酶4（MKK4）的表达，这是c-Jun N端激酶细胞死亡途径中的重要上游激酶。从原代海马神经元培养物（一种体外谷氨酸兴奋性毒性模型）获得了类似的神经保护结果。通过外泌体miRNA的小RNA测序，我们确定miR-200a-3p是MPP + -ADEXs中表达最下调的miRNA。miRNA靶标分析和报告基因测定证实，miR-200a-3p通过与Map2k4 / MKK4 mRNA 3'-UTR上的两个独立位点结合而靶向MKK4。用miR-200a-3p模拟物处理可抑制MKK4 mRNA和蛋白质表达，并减少MPP +处理的SH-SY5Y细胞和谷氨酸处理的海马神经元培养物中的细胞死亡。我们的结果表明正常的星形胶质细胞释放miR-200a-3p，它通过下调MKK4表现出神经保护作用。miRNA靶标分析和报告基因测定证实，miR-200a-3p通过与Map2k4 / MKK4 mRNA 3'-UTR上的两个独立位点结合而靶向MKK4。用miR-200a-3p模拟物处理可抑制MKK4 mRNA和蛋白质表达，并减少MPP +处理的SH-SY5Y细胞和谷氨酸处理的海马神经元培养物中的细胞死亡。我们的结果表明正常的星形胶质细胞释放miR-200a-3p，它通过下调MKK4表现出神经保护作用。miRNA靶标分析和报告基因测定证实，miR-200a-3p通过与Map2k4 / MKK4 mRNA 3'-UTR上的两个独立位点结合而靶向MKK4。用miR-200a-3p模拟物处理可抑制MKK4 mRNA和蛋白质表达，并减少MPP +处理的SH-SY5Y细胞和谷氨酸处理的海马神经元培养物中的细胞死亡。我们的结果表明正常的星形胶质细胞释放miR-200a-3p，它通过下调MKK4表现出神经保护作用。