Screening of cytotoxic or cytostatic flavonoids with quantitative FUCCI-based cell cycle assay

The Fluorescence-Ubiquitin Cell Cycle Indicator (FUCCI) system can be used not only to study gene expression at a specific cell cycle stage, but also to monitor cell cycle transitions in real time. In this study, we used a single clone of FUCCI-expressing HeLa cells (FUCCI-HeLa cells) and monitored the cell cycle in individual live cells over time by determining the ratios between red fluorescence (RF) of RFP-Cdt1 and green fluorescence (GF) of GFP-Geminin. Cytotoxic and cytostatic compounds, the latter of which induced G2 or mitotic arrest, were identified based on periodic cycling of the RF/GF and GF/RF ratios in FUCCI-HeLa cells treated with anticancer drugs. With this cell cycle monitoring system, ten flavonoids were screened. Of these, apigenin and luteolin, which have a flavone backbone, were cytotoxic, whereas kaempferol, which has a flavonol backbone, was cytostatic and induced G2 arrest. In summary, we developed a system to quantitatively monitor the cell cycle in real time. Th...荧光泛素化细胞周期指示剂（Fluorescence-Ubiquitin Cell Cycle Indicator, FUCCI）系统不仅可用于研究特定细胞周期阶段的基因表达，还能实时监测细胞周期转变过程。本研究采用单克隆的表达FUCCI的海拉细胞（FUCCI-HeLa细胞），通过定量检测RFP-Cdt1的红色荧光（red fluorescence, RF）与GFP-Geminin的绿色荧光（green fluorescence, GF）的比值，对单个活细胞的细胞周期进行长时程监测。针对经抗癌药物处理的FUCCI-HeLa细胞，可通过其RF/GF与GF/RF比值的周期性波动，筛选出细胞毒性化合物与细胞生长抑制化合物，其中后者可诱导G2期或有丝分裂期阻滞。利用该细胞周期监测系统，本研究共筛选了10种黄酮类化合物：其中具有黄酮母核的芹菜素与木犀草素表现出细胞毒性；而具有黄酮醇母核的山奈酚则具有细胞生长抑制活性，可诱导G2期阻滞。综上，本研究开发了一套可实时定量监测细胞周期的实验系统。Th...