The aim of this study is to compare the regulatory abilities of citrus flavonoids on the oscillating expression of circadian genes. Seven varieties of citrus fruits and twenty-five citrus flavonoids were selected and evaluated. Per2 luciferase bioluminescence report system and serum shock were used to induce circadian gene expression in mouse microglia BV-2 cells. In vivo experiments were carried out using C57BL6/J mice to evaluate the regulation of flavonoids on the oscillatory expression of liver biorhythm genes. Lipopolysaccharide was used to interfere the gene oscillating expression. QRT-PCR was performed to detect the expression of circadian rhythm-related genes, including Clock, Bmal1, Per1, Per2, Per3, Cry1, Cry2, Rev-erbα, Rev-erbβ, Rorα, Dbp, and Npas2. The results show that the polymethoxyflavones (PMFs) exerted stronger circadian gene regulatory capability, while the flavonoids containing glycosides showed no biological activity. Also, all tested flavonoids decreased LPS-induced nitric oxide release, but only polymethoxyflavones inhibited circadian rhythm disorder. PMFs inhibited Nlrp3 inflammasome-related genes and proteins, including Nlrp3, IL-1β, ASC, and Caspase1, while other flavonoids only affected IL-1β and Caspase1 expression. This mechanism was preliminarily verified using the Nlrp3 inhibitor INF39.

中文翻译：

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柑橘中的多甲氧基黄酮通过抑制 Nlrp3 表达调节脂多糖诱导的昼夜节律基因的振荡衰减

本研究的目的是比较柑橘类黄酮对昼夜节律基因振荡表达的调节能力。选择并评价了7个柑橘类水果和25个柑橘类黄酮。Per2荧光素酶生物发光报告系统和血清休克用于诱导小鼠小胶质细胞BV-2细胞的昼夜节律基因表达。使用 C57BL6/J 小鼠进行体内实验，以评估黄酮类化合物对肝脏生物节律基因振荡表达的调节作用。脂多糖用于干扰基因振荡表达。采用 QRT-PCR 检测昼夜节律相关基因的表达，包括Clock、Bmal1、Per1、Per2、Per3、Cry1、Cry2、Rev-erbα、Rev-erbβ、Rorα、Dbp和Npas2。结果表明，多甲氧基黄酮（PMFs）具有较强的昼夜节律基因调控能力，而含糖苷的黄酮类化合物则无生物活性。此外，所有测试的类黄酮都减少了 LPS 诱导的一氧化氮释放，但只有多甲氧基黄酮抑制了昼夜节律紊乱。PMFs 抑制 Nlrp3 炎症小体相关基因和蛋白质，包括 Nlrp3、IL-1 β、ASC 和 Caspase1，而其他类黄酮仅影响 IL-1 β和 Caspase1 表达。使用Nlrp3抑制剂 INF39初步验证了该机制。