Cucurbitacin B Exerts Antiaging Effects in Yeast by Regulating Autophagy and Oxidative Stress

Abstract

The budding yeast Saccharomyces cerevisiae has been used as a model organism for the basic mechanism of aging, which provides useful assay systems for measuring both replicative and chronological lifespans. In the course of our screening program for substances that extend replicative lifespan, cucurbitacin B (CuB) was found as a hit compound from a compound library, which contains cerebrosides, phenols, sesquiterpenoid, triterpenoids, and sterols isolated from natural products by our research group. Importantly, it prolonged not only the replicative lifespan but also the chronological lifespan in yeast. CuB increased ATG32 gene expression, suggesting that CuB induces autophagy. Indeed, the GFP signal generated from the cleavage of GFP-Atg8, which is a signature of autophagy, was increased upon CuB treatment. On the other hand, CuB failed to increase the chronological lifespans when either ATG2 or ATG32, essential autophagy genes, was deleted, indicating that the lifespan extension by CuB depends on autophagy induction. Furthermore, CuB significantly increased superoxide dismutase (Sod) activity and the survival rate of yeast under oxidative stress, while it decreased the amount of reactive oxygen species (ROS) and malondialdehyde (MDA) production, indicating that CuB has activity to antagonize oxidative stress. Additionally, CuB did not affect replicative lifespans of sod1, sod2, uth1, and skn7 mutants with the K6001 background, indicating that aging-related genes including SOD1, SOD2, UTH1, and SKN7 participate in the antiaging effect of CuB. These results suggest that CuB exerts antiaging activity by regulating autophagy, ROS, antioxidative ability, and aging-related genes. Finally, we discuss the possible intracellular targets of CuB based on the phenotypic comparison between the CuB and global gene deletion databases.

Amarogentin from Gentiana rigescens Franch Exhibits Antiaging and Neuroprotective Effects through Antioxidative Stress

“…The liquid culture medium contained yeast extract, peptone, and D-glucose (YPD, 1% yeast extract, 2% peptone and 2% glucose) or galactose (YPG, 1% yeast extract, 2% peptone, 3% galactose) media. Replicative lifespan assays were performed as described in our previous study [ 20 ]. In brief, the K6001 yeast strain was inoculated in 5 ml of galactose medium and incubated in a shaking incubator at 180 rpm for 24 h at 28°C.…”

“…Natural products are important sources of drugs to reduce or prevent oxidative stresses in humans. In our previous studies, 19 molecules were isolated from natural products which were able to extend the replicative lifespan of yeast via antioxidative stress [20][21][22][23][24][25]. Therefore, searching for effective molecules with antioxidative stress potential should be considered an important strategy to treat aging and neurodegenerative disorders.…”

“…A unique characteristic of K6001 is that only mother cells can produce daughter cells in a glucose medium but not in a galactose medium [26]. In our previous study, K6001, which is a yeast mutant strain, was used as a bioassay system to evaluate the antiaging activity of natural products [20][21][22][23][24][25]. Furthermore, the PC12 cell line, which was derived from rat pheochromocytoma cells, is one of the useful models for studying the nervous system at the cellular level [27].…”

“…Replicative lifespan assays were performed as described in our previous study [20]. In brief, the K6001 yeast strain was inoculated in 5 ml of galactose medium and incubated in a shaking incubator at 180 rpm for 24 h at 28°C.…”

“…To confirm whether these obtained antiaging compounds have an anti-AD effect, PC12 cells were used to evaluate neuritogenic activity of them. Cucurbitacin B which is a triterpenoid and isolated from Pedicellus melo, like amarogentin, not only exerted antiaging effects on yeasts but also induced neurogenesis in PC12 cells and improved the memory of APP/PS1 transgenic mice [20,44]. Amarogentin, which has a structure that is completely different to that of CuB, exhibits significant antiaging activity and NGF-mimic activity and enhances the NGF activity.…”