Innovation Center for Neurological Disorders and Department of Neurology, Xuanwu Hospital, Capital Medical University, National Clinical Research Center for Geriatric Diseases, Beijing, P.R. China.
Beijing Key Laboratory of Geriatric Cognitive Disorders, Beijing, P.R. China.
J Alzheimers Dis. 2021;83(1):367-377. doi: 10.3233/JAD-210411.
Imbalance between amyloid-β (Aβ) production and clearance results in Aβ accumulation. Regulating Aβ levels is still a hot point in the research of Alzheimer's disease (AD).
To identify the differential expression of ATP-binding cassette transporter A1 (ABCA1) and its upstream microRNA (miRNA) in AD models, and to explore their relationships with Aβ levels.
Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were performed to determine the expression of ABCA1 in 5xFAD mice, SH-SY5Y cells treated with Aβ oligomers and SH-SY5YAβPP695 cells (AD models). TargetScan was used to predict the upstream miRNAs for ABCA1. Dual-luciferase assay was conducted to identify the regulation of the miRNA on ABCA1. qRT-PCR was used to measure the expression of miRNA in AD models. Finally, enzyme-linked immunosorbent assays were performed to detect Aβ42 and Aβ40 levels.
The expression of ABCA1 was significantly downregulated in AD models at both mRNA and protein levels. Dual-luciferase assay showed that miR-96-5p could regulate the expression of ABCA1 through binding to the 3 untranslated region of ABCA1. The level of miR-96-5p was significantly elevated in AD models. The expression of ABCA1 was enhanced while Aβ42 levels and Aβ42/Aβ40 ratios were reduced in SH-SY5YAβPP695 cells after treated with miR-96-5p inhibitor.
The current study found that miR-96-5p is the upstream miRNA for ABCA1. Suppression of miR-96-5p in AD models could reduce Aβ42/Aβ40 ratios via upregulating the expression of ABCA1, indicating that miR-96-5p plays an important role in regulating the content of Aβ.
淀粉样蛋白-β（Aβ）的产生和清除失衡会导致 Aβ 积累。调节 Aβ 水平仍然是阿尔茨海默病（AD）研究的热点。
鉴定 AD 模型中 ATP 结合盒转运蛋白 A1（ABCA1）及其上游微小 RNA（miRNA）的差异表达，并探讨其与 Aβ 水平的关系。
采用实时定量聚合酶链反应（qRT-PCR）和 Western blot 法检测 5xFAD 小鼠、Aβ 寡聚体处理的 SH-SY5Y 细胞和 AD 模型 SH-SY5YAβPP695 细胞中 ABCA1 的表达。TargetScan 用于预测 ABCA1 的上游 miRNA。双荧光素酶报告基因实验鉴定 miRNA 对 ABCA1 的调控作用。qRT-PCR 检测 AD 模型中 miRNA 的表达。最后，采用酶联免疫吸附试验检测 Aβ42 和 Aβ40 水平。
AD 模型中 ABCA1 的 mRNA 和蛋白表达均显著下调。双荧光素酶报告基因实验表明，miR-96-5p 可通过与 ABCA1 的 3′非翻译区结合调节 ABCA1 的表达。AD 模型中 miR-96-5p 水平显著升高。在 SH-SY5YAβPP695 细胞中，用 miR-96-5p 抑制剂处理后，ABCA1 的表达增强，Aβ42 水平和 Aβ42/Aβ40 比值降低。
本研究发现 miR-96-5p 是 ABCA1 的上游 miRNA。AD 模型中 miR-96-5p 的抑制可通过上调 ABCA1 的表达降低 Aβ42/Aβ40 比值，表明 miR-96-5p 在调节 Aβ 含量中发挥重要作用。
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