DataSheet_1_C13 Megastigmane Derivatives From Epipremnum pinnatum: β-Damascenone Inhibits the Expression of Pro-Inflammatory Cytokines and Leukocyte Adhesion Molecules as Well as NF-κB Signaling.docx

In order to identify active constituents and to gain some information regarding their mode of action, extracts from leaves of Epipremnum pinnatum were tested for their ability to inhibit inflammatory gene expression in endothelial- and monocyte-like cells (HUVECtert and THP-1, respectively). Bioactivity-guided fractionation using expression of PTGS2 (COX-2) mRNA as a readout resulted in the isolation of two C13 megastigmane glycosides, gusanlungionoside C (1) and citroside A (3), and the phenylalcohol glycoside phenylmethyl-2-O-(6-O-rhamnosyl)-ß-D-galactopyranoside (2). Further analysis identified six additional megastigmane glycosides and the aglycones β-damascenone (10), megastigmatrienone (11), 3-hydroxy-β-damascenone (12), and 3-oxo-7,8-dihydro-α-ionol (13). Pharmacological analysis demonstrated that 10 inhibits LPS-stimulated induction of mRNAs encoding for proinflammatory cytokines and leukocyte adhesion molecules, such as TNF-α, IL-1β, IL-8, COX-2, E-selectin, ICAM-1, and VCAM-1 in HUVECtert and THP-1 cells. 10 inhibited induction of inflammatory genes in HUVECtert and THP-1 cells treated with different agonists, such as TNF-α, IL-1β, and LPS. In addition to mRNA, also the upregulation of inflammatory proteins was inhibited by 10 as demonstrated by immune assays for cell surface E-selectin and secreted TNF-α. Finally, using a luciferase reporter construct, it was shown, that 10 inhibits NF-κB-dependent transcription. Therefore, we hypothesize that inhibition of NF-κB by β-damascenone (10) may represent one of the mechanisms underlying the in vitro anti-inflammatory activity of Epipremnum pinnatum extracts.

为明确麒麟叶（Epipremnum pinnatum）中的有效成分并揭示其作用机制，本研究以该植物叶片提取物为对象，检测其在内皮样细胞（HUVECtert）与单核细胞样细胞（THP-1）中抑制炎症基因表达的活性。以PTGS2（COX-2）mRNA的表达作为活性评价指标，通过生物活性导向分离法，从提取物中分离得到2种C13裂环马钱烷糖苷（C13 megastigmane glycosides）：gusanlungionoside C（1）与citroside A（3），以及1种苯乙醇苷（phenylalcohol glycoside）：苯甲基-2-O-(6-O-鼠李糖基)-β-D-吡喃半乳糖苷（2）。后续分析进一步鉴定出6种额外的裂环马钱烷糖苷，以及4种苷元：β-大马士革酮（β-damascenone，10）、巨马钱三酮（megastigmatrienone，11）、3-羟基-β-大马士革酮（3-hydroxy-β-damascenone，12）与3-氧代-7,8-二氢-α-紫罗兰醇（3-oxo-7,8-dihydro-α-ionol，13）。药理实验结果显示，化合物10可抑制脂多糖（LPS）诱导的HUVECtert与THP-1细胞中促炎细胞因子及白细胞黏附分子编码mRNA的表达，涉及的分子包括肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、白细胞介素-8（IL-8）、COX-2、E-选择素（E-selectin）、细胞间黏附分子-1（ICAM-1）与血管细胞黏附分子-1（VCAM-1）。此外，化合物10可抑制经TNF-α、IL-1β及LPS等不同激动剂处理的HUVECtert与THP-1细胞中炎症基因的诱导表达。除mRNA层面的调控外，化合物10还可抑制炎症蛋白的上调，该结论通过针对细胞表面E-选择素与分泌型TNF-α的免疫检测实验得以验证。最后，借助荧光素酶报告基因构建体（luciferase reporter construct）实验证实，化合物10能够抑制核因子κB（NF-κB）依赖的转录过程。综上，本研究推测β-大马士革酮（10）对NF-κB的抑制作用，或是麒麟叶提取物体现体外抗炎活性的潜在分子机制之一。