胆红素通过调控miR-27a-3p/Rgs1轴对脑缺血/再灌注损伤起保护作用。
Biliverdin Protects Against Cerebral Ischemia/Reperfusion Injury by Regulating the miR-27a-3p/Rgs1 Axis.
作者信息
Li Junjie, Peng Lijia, Bai Wenya, Peng Peihua, Chen Wendong, Yang Wei, Shao Jianlin
机构信息
Department of Anesthesiology, First Affiliated Hospital, Kunming Medical University, Kunming City, 650032, People's Republic of China.
出版信息
Neuropsychiatr Dis Treat. 2021 Apr 22;17:1165-1181. doi: 10.2147/NDT.S300773. eCollection 2021.
BACKGROUND
We have previously demonstrated that biliverdin has neuroprotective effects that ameliorate cerebral ischemia/reperfusion (I/R) injury in rats. However, the underlying mechanism is unknown. This study aimed at elucidating on the modulatory role of miR-27a-3p on Rgs1 as a mechanism by which biliverdin affects cerebral I/R injury.
METHODS
Middle cerebral artery occlusion/reperfusion (MCAO/R) was used to establish I/R rat models while oxygen glucose deprivation/reoxygenation (OGD/R) was used to induce hippocampal neurons to establish I/R models in vitro. Infarct volume was assessed by TTC staining. Apoptotic analyses of ischemic cortical neurons and cells were performed by TUNEL staining and flow cytometry, respectively. Cell viability was assessed by the CCK-8 assay while the target of miR-27a-3p was determined by double luciferase reporter assay. Relative expression levels of miR-27a-3p and Rgs1 (in vivo and in vitro) as well as markers of inflammation and apoptosis (in vitro) were detected by RT-qPCR. Then, Elisa and western blot were used to assess protein expression levels of inflammatory and apoptotic markers in vitro.
RESULTS
Biliverdin suppressed inflammation and apoptosis in hippocampal neurons upon OGD/R, and reduced cerebral infarction volume as well as apoptosis in the MCAO/R rat model. Furthermore, biliverdin upregulated miR-27a-3p and downregulated hippocampal neuron Rgs1 after OGD/R as well as in rat brain tissues after cerebral I/R. Bioinformatic analysis revealed an miR-27a-3p docking site in the 3'-UTR region of Rgs1. Luciferase reporter assays showed that Rgs1 is an miR-27a-3p target. Moreover, miR-27a-3p upregulation inhibited OGD/R-triggered inflammation and suppressed neuronal apoptosis. Rgs1 knockdown suppressed OGD/R-triggered inflammation and decreased neuronal apoptosis while miR-27a-3p downregulation reversed the protective effect of Rgs1 knockdown. Moreover, miR-27a-3p overexpression and Rgs1 silencing suppressed NF-κB (p65) expression.
CONCLUSION
Biliverdin protects against cerebral I/R injury by regulating the miR-27a-3p/Rgs1 axis, thereby inhibiting inflammation and apoptosis.
背景
我们之前已经证明，胆红素具有神经保护作用，可改善大鼠脑缺血/再灌注（I/R）损伤。然而，其潜在机制尚不清楚。本研究旨在阐明miR-27a-3p对Rgs1的调节作用，作为胆红素影响脑I/R损伤的一种机制。
方法
采用大脑中动脉闭塞/再灌注（MCAO/R）建立I/R大鼠模型，同时采用氧糖剥夺/复氧（OGD/R）诱导海马神经元建立体外I/R模型。通过TTC染色评估梗死体积。分别通过TUNEL染色和流式细胞术对缺血皮质神经元和细胞进行凋亡分析。通过CCK-8法评估细胞活力，同时通过双荧光素酶报告基因检测确定miR-27a-3p的靶标。通过RT-qPCR检测miR-27a-3p和Rgs1（体内和体外）的相对表达水平以及炎症和凋亡标志物（体外）。然后，使用酶联免疫吸附测定（ELISA）和蛋白质印迹法评估体外炎症和凋亡标志物的蛋白质表达水平。
结果
胆红素抑制OGD/R后海马神经元的炎症和凋亡，并减少MCAO/R大鼠模型中的脑梗死体积和凋亡。此外，胆红素在OGD/R后以及脑I/R后的大鼠脑组织中上调miR-27a-3p并下调海马神经元Rgs1。生物信息学分析揭示了Rgs1的3'-UTR区域中的miR-27a-3p对接位点。荧光素酶报告基因检测表明Rgs1是miR-27a-3p的靶标。此外，miR-27a-3p上调抑制OGD/R引发的炎症并抑制神经元凋亡。Rgs1敲低抑制OGD/R引发的炎症并减少神经元凋亡，而miR-27a-3p下调逆转了Rgs1敲低的保护作用。此外，miR-27a-3p过表达和Rgs1沉默抑制NF-κB（p65）表达。
结论
胆红素通过调节miR-27a-3p/Rgs1轴来预防脑I/R损伤，从而抑制炎症和凋亡。
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