Department of Paediatrics, Taizhou People's Hospital, Taizhou 225300, Jiangsu, China.
Department of Paediatrics, Taizhou People's Hospital, Taizhou 225300, Jiangsu, China.
Neurosci Res. 2021 Oct;171:103-113. doi: 10.1016/j.neures.2021.03.003. Epub 2021 Mar 17.
Neonatal hypoxic-ischemia encephalopathy (HIE) refers to hypoxic-ischemic brain damage caused by perinatal asphyxia. Increasing evidence has revealed the crucial roles of microRNAs (miRNAs) in neonatal HIE. In the current research, we aimed to explore the biological role of miR-363-3p in neonatal HIE. For this purpose, we established in vitro models of PC-12 and SH-SY5Y cells subjected to oxygen-glucose deprivation and reperfusion (OGD/R) and an in vivo rat model subjected to middle cerebral artery occlusion/reperfusion (MCAO/R) treatment. First, using H&E staining, TTC staining, and western blot analysis, we observed that DUSP5 knockdown suppressed HIE in vivo. Then, by performing flow cytometric analysis, western blotting, RT-qPCR, and MTT assays, we observed that DUSP5 silencing suppressed OGD/R-induced cell injury in vitro. Subsequently, we explored the potential regulatory mechanism of DUSP5 in OGD/R-treated cells with luciferase reporter assays and RT-qPCR analysis. The results demonstrated that DUSP5 was targeted by miR-363-3p. Next, functional assays, including flow cytometric analysis, MTT assays, western blotting and RT-qPCR, were conducted to explore the biological functions of miR-363-3p in SH-SY5Y and PC-12 cells. Our data showed that miR-363-3p overexpression suppressed OGD/R-induced cell injury. Finally, the results from rescue experiments showed that enhanced DUSP5 expression counteracted the effect of miR-363-3p overexpression. In conclusion, our data suggested that miR-363-3p attenuates neonatal HIE by targeting DUSP5.
新生儿缺氧缺血性脑病 (HIE) 是指围产期窒息引起的缺氧缺血性脑损伤。越来越多的证据表明 microRNAs (miRNAs) 在新生儿 HIE 中发挥着关键作用。在目前的研究中,我们旨在探索 miR-363-3p 在新生儿 HIE 中的生物学作用。为此,我们建立了 PC-12 和 SH-SY5Y 细胞体外氧葡萄糖剥夺再灌注 (OGD/R) 模型和体内大脑中动脉阻塞再灌注 (MCAO/R) 大鼠模型。首先,通过 H&E 染色、TTC 染色和 Western blot 分析,我们观察到 DUSP5 敲低抑制了体内 HIE 的发生。然后,通过流式细胞术分析、Western blot、RT-qPCR 和 MTT 测定,我们观察到 DUSP5 沉默抑制了体外 OGD/R 诱导的细胞损伤。随后,我们通过荧光素酶报告基因检测和 RT-qPCR 分析探索了 DUSP5 在 OGD/R 处理细胞中的潜在调节机制。结果表明 DUSP5 是 miR-363-3p 的靶基因。接下来,进行了功能测定,包括流式细胞术分析、MTT 测定、Western blot 和 RT-qPCR,以探索 miR-363-3p 在 SH-SY5Y 和 PC-12 细胞中的生物学功能。我们的数据表明 miR-363-3p 过表达抑制了 OGD/R 诱导的细胞损伤。最后,通过挽救实验的结果表明,增强的 DUSP5 表达抵消了 miR-363-3p 过表达的作用。总之,我们的数据表明,miR-363-3p 通过靶向 DUSP5 来减轻新生儿 HIE。