2. Academic Validation
  3. Three Polymethoxyflavones Purified from Ougan ( Citrus reticulata Cv. Suavissima) Inhibited LPS-Induced NO Elevation in the Neuroglia BV-2 Cell Line via the JAK2/STAT3 Pathway

Three Polymethoxyflavones Purified from Ougan ( Citrus reticulata Cv. Suavissima) Inhibited LPS-Induced NO Elevation in the Neuroglia BV-2 Cell Line via the JAK2/STAT3 Pathway

  • Nutrients. 2019 Apr 5;11(4):791. doi: 10.3390/nu11040791.
Yue Wang 1,  Wenjing Zang 2,  Shiyu Ji 3,  Jinping Cao 4,  Chongde Sun 5
Affiliations

Affiliations

  • 1 Laboratory of Fruit Quality Biology/The State Agriculture Ministry Laboratory of Horticultural Plant Growth, Development and Quality Improvement, Zhejiang University, Zijingang Campus, Hangzhou 310058, China. fruit@zju.edu.cn.
  • 2 Laboratory of Fruit Quality Biology/The State Agriculture Ministry Laboratory of Horticultural Plant Growth, Development and Quality Improvement, Zhejiang University, Zijingang Campus, Hangzhou 310058, China. zangwj@zju.edu.cn.
  • 3 Laboratory of Fruit Quality Biology/The State Agriculture Ministry Laboratory of Horticultural Plant Growth, Development and Quality Improvement, Zhejiang University, Zijingang Campus, Hangzhou 310058, China. jishiyu@zju.edu.cn.
  • 4 Laboratory of Fruit Quality Biology/The State Agriculture Ministry Laboratory of Horticultural Plant Growth, Development and Quality Improvement, Zhejiang University, Zijingang Campus, Hangzhou 310058, China. caojinpingabc@126.com.
  • 5 Laboratory of Fruit Quality Biology/The State Agriculture Ministry Laboratory of Horticultural Plant Growth, Development and Quality Improvement, Zhejiang University, Zijingang Campus, Hangzhou 310058, China. adesun2006@zju.edu.cn.
PMID: 30959824 DOI: 10.3390/nu11040791
Abstract

In order to establish an efficient method for separation of polymethoxyflavones (PMFs) and explore the anti-inflammatory mechanism of PMF monomers, a citrus variety rich in PMFs, Ougan (Citrus reticulata cv. Suavissima), was selected, and three monomers, including nobiletin, tangeretin, and 5-demethylnobiletin, were purified by ultrasonic-assisted extraction, solid phase extraction, and high-speed countercurrent chromatography separation. UPLC-MS was used to identify the three monomers. UPLC determined purities of 99.87% to nobiletin, 99.76% to tangeretin, and 98.75% to 5-demethylnobiletin with the standard curve method. A lipopolysaccharide (LPS)-induced NO releasing model was performed in the mouse microglia BV-2 cell line. Results illustrated that PMF monomers inhibited the NO release and the inflammation-related cytokines, including IL-1β, IL-6, and TNFα elevation. QRT-PCR revealed that PMFs alleviated LPS-induced upregulation of iNOS, IL-6, JAK2, TNFα, IL-1β, and NF-κB and LPS-induced downregulation of IκBα, while they did not affect TLR1, TLR2, TLR4, and TLR6. STAT3 expression was repressed by tangeretin and 5-demethylnobiletin, but not by nobiletin. Western blot assay also showed a suppression of expression and phosphorylation of JAK2 by all three PMF monomers, while STAT3 phosphorylation was restrained by tangeretin and 5-demethylnobiletin. The mechanism was primarily verified by the JAK2 Inhibitor Ruxolitinib and the STAT3 Inhibitor Stattic.

Keywords

5-demethylnobiletin; Citrus reticulata cv. Suavissima; JAK2/STAT3 pathway; NO release; neuroglia; nobiletin; polymethoxyflavone; purification; tangeretin.

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