Natural Potent NAAA Inhibitor Atractylodin Counteracts LPS-Induced Microglial Activation.
Natural Potent NAAA Inhibitor Atractylodin Counteracts LPS-Induced Microglial Activation.
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天然强效 NAAA 抑制剂苍术碱可抵消 LPS 诱导的小胶质细胞激活。
DOI：
10.3389/fphar.2020.577319
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发表时间：
2020
影响因子：
5.6
通讯作者：
Qiu Y
中科院分区：
文献类型：
--
作者：
Yang L;Ji C;Li Y;Hu F;Zhang F;Zhang H;Li L;Ren J;Wang Z;Qiu Y
N-酰基乙醇胺水解酸性酰胺酶（NAAA）是一种溶酶体酶，可抑制棕榈酰乙醇胺（PEA）的降解，PEA是一种内源性脂质，通过激活PPARα诱导镇痛、抗炎和抗多发性硬化。迄今为止，只有少数有效的NAAA抑制剂被报道，这主要是由于NAAA的限制性底物结合位点。在这里，我们建立了一个高通量的荧光为基础的分析NAAA抑制剂筛选。从天然产物的小文库中发现了几种新的NAAA抑制剂。其中之一是白藜芦醇苷，一种来自蓝茅根的聚乙烯炔化合物，其显著抑制NAAA活性，IC 50为2.81 µM。动力学分析和透析实验表明，Escheridin通过与酶的可逆反应参与竞争性抑制。对接分析表明，Escheridin占据催化腔的NAAA，其中Escheridin呋喃头基团具有疏水相关的相互作用与骨干的Trp 181和Leu 152残基的人NAAA。进一步的研究表明，在BV-2小胶质细胞中，Escheridin显著增加PEA和OEA水平，并剂量依赖性地抑制LPS诱导的硝酸盐、TNF-α、IL-1β和IL-6促炎细胞因子释放。我们的研究结果表明，白藜芦醇苷提高细胞PEA水平，并通过抑制NAAA活性来抑制小胶质细胞活化，这反过来又有助于NAAA功能研究。
N-acylethanolamine-hydrolyzing acid amidase (NAAA) is a lysosomal enzyme that inhibits the degradation of palmitoylethanolamide (PEA), an endogenous lipid that induces analgesic, anti-inflammation, and anti-multiple sclerosis through PPARα activation. Only a few potent NAAA inhibitors have been reported to date, which is mainly due to the restricted substrate-binding site of NAAA. Here, we established a high-throughput fluorescence-based assay for NAAA inhibitor screening. Several new classes of NAAA inhibitors were discovered from a small library of natural products. One of these is atractylodin, a polyethylene alkyne compound from the root of Atractylodes lancea (Thunb) DC., which significantly inhibits NAAA activity and has an IC50 of 2.81 µM. Kinetic analyses and dialysis assays suggested that atractylodin engages in competitive inhibition via reversible reaction to the enzyme. Docking assays revealed that atractylodin occupies the catalytic cavity of NAAA, where the atractylodin furan head group has a hydrophobic-related interaction with the backbone of the Trp181 and Leu152 residues of human NAAA. Further investigation indicated that atractylodin significantly increases PEA and OEA levels and dose-dependently inhibits LPS-induced nitrate, TNF-α, IL-1β, and IL-6 pro-inflammatory cytokine release in BV-2 microglia. Our results show that atractylodin elevates cellular PEA levels and inhibits microglial activation by inhibiting NAAA activity, which in turn could contribute to NAAA functional research.