Rheum undulatum 에서 추출한 rhapontigenin의 항산화 활성을 통한 세포손상에 대한 보호 작용

Abstract

Rheum undulatum 에서 추출한 항산화 활성 물질인 rhapontigenin 과 rhaponticin 에 대해 연구를 진행하였다. Rhapontigenin은 세포 내 ROS 소거활성, DPPH 라디칼 소거활성 및 과산화수소 소거활성을 나타내었으며 rhapontigenin은 rhaponticin보다 더 높은 라디칼 소거활성을 나타내었다. Thiobarbituric acid reaction method 와 comet assay를 통하여 rhapontigenin은 세포손상의 주요 타깃으로 되는 과산화수소에 의한 세포막의 지질과산화 및 DNA 손상으로부터 세포를 보호함을 관찰하였다. Rhapontigenin는 과산화수소에 노출된 Chinese hamster lung fibroblast (V79-4) cells의 apoptosis를 저해함을 MTT assay, Hoechst 33342 staining assay 및 flow cytometry analysis 로 측정하였다. Rhapontigenin는 serum starvation에 의해 유도되는 세포손상을 저해하였으며 catalase의 활성을 증가시켰고 상응한 protein의 expression을 증가시켰다. 또한 western blot analysis로부터 rhapontigenin은 extracellular signal regulated kinase (ERK)의 phosphorylation을 증가시킴을 알 수 있었으며 EMSA의 결과로부터 rhapontigenin은 activator protein 1 (AP-1)- a redox sensitive transcription factor 의 활성을 저해하였음을 관찰하였다. 이러한 결과들로부터 rhapontigenin은 세포의 항산화활성을 증가시키고 signal pathways를 조절함으로서 V79-4 세포의 산화적 손상을 저해함을 알 수 있었다.

The antioxidant properties of rhapontigenin and rhaponticin isolated from Rheum undulatum were investigated. Rhapontigenin was found to scavenge intracellular reactive oxygen species (ROS), the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals, and hydrogen peroxide (H₂O₂) by 2',7'-dichlorofluorescin dictate (DCF-DA) method, DPPH radical scavenging activity assay, and H₂O₂ scavenging activity assay respectively. The radical scavenging effect of rhapontigenin was more effective than rhaponticin. Rhapontigenin protected against H₂O₂ induced membrane lipid peroxidation and cellular DNA damage, the main targets of oxidative stress-induced cellular damage, which were detected by thiobarbituric acid reaction (TBAR) method and comet assay. The radical scavenging activity of rhapontigenin protected Chinese hamster lung fibroblast (V79-4) cells exposed to H₂O₂ by inhibiting apoptosis in MTT assay, Hoechst 33342 staining assay, and flow cytometry analysis. Rhapontigenin inhibited cell damage induced by serum starvation and also increased the activity of catalase and its protein expression. Further, western blot analysis of cell lysates indicated that rhapontigenin increased phosphorylation of extracellular signal regulated kinase (ERK) and the result of electrophoretic mobility shift assay (EMSA) showed that rhapontigenin inhibited the activity of activator protein 1 (AP-1), a redox sensitive transcription factor. In summary, these results suggest that rhapontigenin protects V79-4 cells against oxidative damage by enhancing the cellular antioxidant activity and modulating cellular signal pathways.