从日本玉露 Ophiopogon japonicus(莲草)中提取的甾体类皂苷酮鲨目根（Ruscogenin，rus）具有缓解脑缺血再灌注损伤（cerebral ischemia-reperfusion injury，IRI）和急性肺损伤的效果。目前，rus的时间药效学作用仍未知。本研究探讨了不同时点 rus 给药对小鼠脑IRI的缓解效果及机制。不同组别的动物在不同时段（23:00-01:00、05:00-07:00、11:00-13:00、17:00-19:00）分别经口饲服rus（10mg/kg），连续3天。第4天，在5:00-7:00进行大脑中动脉阻断（middle cerebral artery occlusion，MCAO）手术。进行行为测试，并收集脑组织进行梗死体积、qPCR和免疫印迹检测。通过qPCR检测肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）、白细胞介素6（interleukin 6，IL-6）、白细胞介素1β（interleukin-1beta，IL-1β）和诱导型一氧化氮合酶（inducible nitric oxide synthase，iNOS）的水平。检测血清和大脑皮层中的谷胱甘肽（glutathione，GSH）、超氧化物歧化酶（superoxide dismutase，SOD）活力和丙二醛（malondialdehyde，MDA）含量。通过免疫印迹检测时钟基因。根据这些结果，选择17:00-19:00时段继续研究rus的机制，并将Nrf2阻断剂ATRA（all-trans-retinoic acid）连续3天于14:00给予处理。rus的给药降低了脑梗死体积，改善了行为评分，并上调了Per1、Bmal1、Clock、Rev-erbα、转录因子核因子红系2相关因子2（transcription factor nuclear factor erythroid 2-related factor 2，Nrf2）、血红素加氧酶1（heme oxygenase-1，HO-1）和醌氧还原酶1（quinone oxidoreductase 1，NQO1）的mRNA和蛋白表达。17:00-19:00时段给予rus的预防效果优于其他三个时间点。ATRA的联合给药削弱了rus的预防效果。给药时间影响rus的预防效果，由Nrf2信号通路调节。综上所述，我们提供了坚实的证据，表明不同给药时间点影响rus在缓解IRI中的有效性。版权归2023年Elsevier GmbH所有。保留所有权利。

Ruscogenin (Rus), a steroidal sapogenin extracted from Ophiopogon japonicus (L. f.) Ker-Gawl., has the effect of alleviating cerebral ischemia-reperfusion injury (IRI), acute lung injury. At present, the chronopharmacological effects of Rus are still unknown.This study explored the alleviating effect and mechanism of Rus timing administration on mice cerebral IRI.The animals in different groups were administrated Rus (10 mg/kg) by gavage at four time points (23:00-01:00, 05:00-07:00, 11:00-13:00, 17:00-19:00) respectively for 3 days. On the 4th day, middle cerebral artery occlusion (MCAO) surgery was operated during 5:00-7:00. Behavioral tests were executed and the brain was collected for infarct volume, qPCR and immunoblot detection. The levels of tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), interleukin-1beta (IL-1β) and inducible nitric oxide synthase (iNOS) were detected by qPCR. Glutathione (GSH), superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in serum and cerebral cortex were detected. The clock genes were tested by western blot. Based on these results, 17:00-19:00 was selected to administrate Rus for further mechanism study and Nrf2 blocker group was administrated all-trans-retinoic acid (ATRA) at 14:00 for 3 days.Administration of Rus reduced cerebral infarcted volume, ameliorated the behavior score and upregulated the mRNA and protein expression of Per1, Bmal1, Clock, Rev-erbα, transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), quinone oxidoreductase 1 (NQO1). Administration of Rus during 17:00-19:00 had better preventive effect than other three time points. Combined administration of ATRA blunted the preventive effect of Rus.The preventive effect of Rus is affected by the time of administration, which was regulated by Nrf2 pathway. Taken together, we provide solid evidence to suggest that different administration time point affect the effectiveness of Rus in alleviating IRI.Copyright © 2023 Elsevier GmbH. All rights reserved.