Department of Neurology, Henan Provincial People's Hospital, No. 7 Weiwu Road, Zhengzhou, 451450, China.
Neurochem Res. 2020 Apr;45(4):741-751. doi: 10.1007/s11064-019-02947-6. Epub 2020 Jan 2.
Accumulating articles reported that berberine (Ber) played a neuroprotective role in Alzheimer's disease (AD). Long noncoding RNAs (lncRNAs) have been identified as biomarkers and therapeutic targets of AD. However, the precise mechanism by which lncRNA β-amyloid cleaving enzyme 1 antisense RNA (BACE1-AS)regulates the progression of AD remains largely unknown. HPN and SK-N-SH cells treated with amyloid β 25-35 (Aβ) were regarded as AD model in vitro. Cell survival rate was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Lactate dehydrogenase (LDH) cytotoxicity assay was conducted to detect the cytotoxicity of neuronal cells. Flow cytometry was performed to determine the intracellular concentration of Ca, reactive oxygen species (ROS) and apoptosis of neuronal cells. Western blot assay was carried out to detect the apoptosis-related proteins of neuronal cells. The abundance of lncRNA BACE1-AS and miR-132-3p was measured by quantitative real time polymerase chain reaction (qRT-PCR). The binding sites between miR-132-3p and BACE1-AS were predicted by Starbase, and the combination was confirmed by dual-luciferase reporter assay. We found that Ber alleviated Aβ induced neuronal injury in AD model, especially in high concentration Ber group. The enrichment of BACE1-AS was positively regulated by Aβ and was inversely modulated by Ber in neuronal cells. The interference of BACE1-AS alleviated the neuronal damage of AD model. miR-132-3p was a direct target of lncRNA BACE1-AS in HEK293T cells, and it was negatively regulated by BACE1-AS in neuronal cells. BACE1-AS accumulation reversed the protective effect of miR-132-3p overexpression on AD model. Ber treatment and BACE1-AS intervention recovered the viability of AD model. Ber up-regulated the level of miR-132-3p via BACE1-AS in SK-N-SH and HPN neuronal cells. in conclucsion, Ber protected neuronal cells against Aβ at least partly through BACE1-AS/miR-132-3p axis. The combined therapy of Ber treatment with BACE1-AS depletion might provide new insight into AD treatment.
越来越多的文章报道，小檗碱（Ber）在阿尔茨海默病（AD）中发挥神经保护作用。长链非编码 RNA（lncRNA）已被鉴定为 AD 的生物标志物和治疗靶点。然而，lncRNA β-淀粉样蛋白裂解酶 1 反义 RNA（BACE1-AS）调节 AD 进展的确切机制在很大程度上仍然未知。用淀粉样β 25-35（Aβ）处理 HPN 和 SK-N-SH 细胞被视为体外 AD 模型。通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四唑溴盐（MTT）测定法测量细胞存活率。进行乳酸脱氢酶（LDH）细胞毒性测定以检测神经元细胞的细胞毒性。通过流式细胞术测定神经元细胞内 Ca、活性氧（ROS）和细胞凋亡的浓度。通过 Western blot 测定法检测神经元细胞的凋亡相关蛋白。通过定量实时聚合酶链反应（qRT-PCR）测量 lncRNA BACE1-AS 和 miR-132-3p 的丰度。通过 Starbase 预测 miR-132-3p 和 BACE1-AS 之间的结合位点，并通过双荧光素酶报告基因测定法确认结合。我们发现，Ber 减轻了 AD 模型中 Aβ 诱导的神经元损伤，尤其是在高浓度 Ber 组中。在神经元细胞中，Aβ 正向调节 BACE1-AS 的富集，而 Ber 则反向调节。BACE1-AS 的干扰减轻了 AD 模型的神经元损伤。miR-132-3p 是 HEK293T 细胞中 lncRNA BACE1-AS 的直接靶标，并且在神经元细胞中受 BACE1-AS 的负调控。BACE1-AS 积累逆转了 miR-132-3p 过表达对 AD 模型的保护作用。Ber 处理和 BACE1-AS 干预恢复了 AD 模型的活力。Ber 通过 BACE1-AS 在 SK-N-SH 和 HPN 神经元细胞中上调 miR-132-3p 的水平。总之，Ber 通过 BACE1-AS/miR-132-3p 轴至少部分保护神经元细胞免受 Aβ的侵害。Ber 治疗与 BACE1-AS 耗竭的联合治疗可能为 AD 治疗提供新的见解。
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