sRAGE mRNA 3'未翻译区相关microRNA的鉴定及其在阿尔茨海默病治疗中的作用研究

Receptor for advanced glycation end-products (RAGE) is known to be involved in Alzheimer's disease (AD). Ligand engagement of RAGE activates multiple signaling pathways resulting in amplification of inflammatory responses and overproduction of Aβ. In contrast, a secretory splice isoform of RAGE, termed soluble RAGE (sRAGE), has the ability to inhibit the RAGE-mediated pathological effects by functioning as a decoy receptor. Using computational miRNA target prediction algorithms, we found that miR-106b could potentially target the 3'UTRs of sRAGE. Thus, in this study, we attempted to confirm whether sRAGE was a true target of miR-106b. For this purpose, luciferase activity was measured in cells transfected with pRL-3'-UTR-sRAGE (wild type) or pRL-3'-UTR(Δ-miR-122)-sRAGE (mutant) along with miR-106b mimic or control RNA. We also explore whether single nucleotide polymorphisms (SNPs) in the miR-106b binding region could affect miRNA mediated regulation of sRAGE, which may lead to altered AD susceptibility and serum sRAGE levels. On the other hand, to investigate the functional significance of miR-106b in AD brain, we will use an antagomir to silence miR-106b and analyze whether AD pathology is alleviated. The findings of this research will not only provide potential microRNA target for development of therapeutics for AD, but may also provide genetic marker to screen those AD patients who are sensitive to miR-106b antagomir.

晚期糖基化终末产物受体（RAGE）与其配体结合可激活多条信号传导通路，导致脑内炎症反应增加及Aβ过度生成；可溶性晚期糖基化终末产物受体（sRAGE）是RAGE的选择性剪接异构体，可阻止配体与RAGE结合从而减轻RAGE的有害效应。通过生物信息学分析我们发现sRAGE转录本的3'非翻译区有miR-106b作用靶点。本课题采用荧光报告基因检测miR-106b对sRAGE的调控作用及miR-106b靶序列单核苷酸多态（SNP）对这一作用的影响。同时通过AD动物模型探讨miR-106b抑制剂能否增加sRAGE表达并减轻AD症状；另一方面，研究miR-106b靶序列多态性与AD易感性的关联及对血清sRAGE表达水平的影响。这将为开发sRAGE微小RNA抑制剂治疗AD提供理论依据；miR-106b靶序列SNP可作为一个遗传标记位点，筛选对miR-106b抑制剂敏感的AD患者，为个体化治疗提供理论指导。

晚期糖基化终末产物受体（RAGE）与其配体结合可激活多条信号传导通路，导致脑内炎症反应增加及β淀粉样肽(Aβ)过度生成；可溶性晚期糖基化终末产物受体（sRAGE）是RAGE的选择性剪接异构体，可阻止配体与RAGE结合从而减轻RAGE的有害效应。本研究通过生物信息学分析确定sRAGE转录的3’非翻译区有一个（microRNA）结合位点。以此为工作基础我们在SH-SY5Y细胞中，利用实时定量PCR和Western-Blot方法，通过使用mimics和antagomir从正负两个效应方面研究转染该microRNA对sRAGE的转录后调控作用。本研究证实在SH-SY5Y细胞中过表达该microRNA后，细胞分泌sRAGE的核酸和蛋白水平均下降；在干扰该microRNA后，细胞分泌sRAGE的核酸和蛋白水平均增加，靶向调节成立。这个结果提示sRAGE很可能是该microRNA靶基因，这将为进一步开发micro RNA antagomir作为治疗AD药物提供理论基础。