Min Zhuo, Tang Ying, Hu Xiao-Tong, Zhu Bing-Lin, Ma Yuan-Lin, Zha Jing-Si, Deng Xiao-Juan, Yan Zhen, Chen Guo-Jun
Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing Key Laboratory of Neurology, Chongqing, China.
Department of Neurology, The Ninth People's Hospital of Chongqing, Chongqing, China.
Front Mol Neurosci. 2018 Jun 11;11:198. doi: 10.3389/fnmol.2018.00198. eCollection 2018.
The α-secretase "a disintegrin and metalloproteinase domain-containing protein" (ADAM10) is involved in the processing of amyloid precursor protein (APP). Upregulation of ADAM10 precludes the generation of neurotoxic β-amyloid protein (Aβ) and represents a plausible therapeutic strategy for Alzheimer's disease (AD). In this study, we explored compounds that can potentially promote the expression of ADAM10. Therefore, we performed high-throughput small-molecule screening in SH-SY5Y (human neuroblastoma) cells that stably express a luciferase reporter gene driven by the ADAM10 promoter, including a portion of its 5'-untranslated region (5'UTR). This has led to the discovery of cosmosiin (apigenin 7-O-β-glucoside). Here, we report that in human cell lines (SH-SY5Y and HEK293), cosmosiin proportionally increased the levels of the immature and mature forms of the ADAM10 protein without altering its mRNA level. This effect was attenuated by translation inhibitors or by deleting the 5'UTR of ADAM10, suggesting that a translational mechanism was responsible for the increased levels of ADAM10. Luciferase deletion assays revealed that the first 144 nucleotides of the 5'UTR were necessary for mediating the cosmosiin-induced enhancement of ADAM10 expression in SH-SY5Y cells. Cosmosiin failed to increase the levels of the ADAM10 protein in murine cells, which lack native expression of the ADAM10 transcript containing the identified 5'UTR element. The potential signaling pathway may involve phosphatidylinositide 3-kinase (PI3K) because pharmacological inhibition of PI3K attenuated the effect of cosmosiin on the expression of the ADAM10 protein. Finally, cosmosiin attenuated Aβ generation because the levels of Aβ40/42 in HEK-APP cells were significantly reduced after cosmosiin treatment. Collectively, we found that the first 144 nucleotides of the ADAM10 5'UTR, and PI3K signaling, are involved in cosmosiin-induced enhancement of the expression of ADAM10 protein. These results suggest that cosmosiin may be a potential therapeutic agent in the treatment of AD.
α-分泌酶“含解整合素和金属蛋白酶结构域蛋白”（ADAM10）参与淀粉样前体蛋白（APP）的加工过程。ADAM10的上调可防止神经毒性β-淀粉样蛋白（Aβ）的产生，是治疗阿尔茨海默病（AD）的一种可行治疗策略。在本研究中，我们探索了可能促进ADAM10表达的化合物。因此，我们在稳定表达由ADAM10启动子驱动的荧光素酶报告基因（包括其部分5'-非翻译区（5'UTR））的SH-SY5Y（人神经母细胞瘤）细胞中进行了高通量小分子筛选。这导致了紫薇苷（芹菜素7-O-β-葡萄糖苷）的发现。在此，我们报告在人细胞系（SH-SY5Y和HEK293）中，紫薇苷可成比例增加ADAM10蛋白未成熟和成熟形式的水平，而不改变其mRNA水平。翻译抑制剂或删除ADAM10的5'UTR可减弱这种作用，表明翻译机制是ADAM10水平升高的原因。荧光素酶缺失分析表明，5'UTR的前144个核苷酸是介导紫薇苷诱导的SH-SY5Y细胞中ADAM10表达增强所必需的。紫薇苷未能增加缺乏含有已鉴定5'UTR元件的ADAM10转录本天然表达的鼠细胞中ADAM10蛋白的水平。潜在的信号通路可能涉及磷脂酰肌醇3-激酶（PI3K），因为PI3K的药理学抑制减弱了紫薇苷对ADAM10蛋白表达的影响。最后，紫薇苷减弱了Aβ的产生，因为紫薇苷处理后HEK-APP细胞中Aβ40/42的水平显著降低。总体而言，我们发现ADAM10 5'UTR的前144个核苷酸和PI3K信号传导参与了紫薇苷诱导的ADAM10蛋白表达增强。这些结果表明，紫薇苷可能是治疗AD的潜在治疗剂。
