Abstract
BackgroundAlthough it has been reported to contain high polyphenols, the pharmacological studies of the calyx of Diospyros kaki Thunb (DKC) have not been elucidated in detail. In this study, we elucidated anti-cancer activity and potential molecular mechanism of DKC against human colorectal cancer cells.MethodsAnti-cell proliferative effect of 70% ethanol extracts from the calyx of Diospyros kaki (DKC-E70) was evaluated by MTT assay. The effect of DKC-E70 on the expression of cyclin D1 in the protein and mRNA level was evaluated by Western blot and RT-PCR, respectively.ResultsDKC-E70 suppressed the proliferation of human colorectal cancer cell lines such as HCT116, SW480, LoVo and HT-29. Although DKC-E70 decreased cyclin D1 expression in protein and mRNA level, decreased level of cyclin D1 protein by DKC-E70 occurred at the earlier time than that of cyclin D1 mRNA, which indicates that DKC-E70-mediated downregulation of cyclin D1 protein may be a consequence of the induction of degradation and transcriptional inhibition of cyclin D1. In cyclin D1 degradation, we found that cyclin D1 downregulation by DKC-E70 was attenuated in presence of MG132. In addition, DKC-E70 phosphorylated threonine-286 (T286) of cyclin D1 and T286A abolished cyclin D1 downregulation by DKC-E70. We also observed that DKC-E70-mediated T286 phosphorylation and subsequent cyclin D1 degradation was blocked in presence of the inhibitors of ERK1/2, p38 or GSK3β. In cyclin D1 transcriptional inhibition, DKC-E70 inhibited the expression of β-catenin and TCF4, and β–catenin/TCF-dependent luciferase activity.ConclusionsOur results suggest that DKC-E70 may downregulate cyclin D1 as one of the potential anti-cancer targets through cyclin D1 degradation by T286 phosphorylation dependent on ERK1/2, p38 or GSK3β, and cyclin D1 transcriptional inhibition through Wnt signaling. From these findings, DKC-E70 has potential to be a candidate for the development of chemoprevention or therapeutic agents for human colorectal cancer.
Highlights
It has been reported to contain high polyphenols, the pharmacological studies of the calyx of Diospyros kaki Thunb (DKC) have not been elucidated in detail
Calyx of Diospyros kaki Thunberg (DKC)-E70 suppresses the proliferation of human colorectal cancer cells To assess whether DKC-E70 possesses anti-cancer property, we evaluated anti-proliferative activity in human colorectal cancer cell lines such as HCT116, SW480, LoVo and HT-29 cells using MTT assay
Because we observed that DKC-E70 inhibited the expression of cyclin D1 mRNA, we investigated the effect of DKC-E70 on Wnt signaling using Western blot against β-catenin and T-cell factor-4 (TCF4), and β –catenin/TCF-dependent luciferase activity
Summary
It has been reported to contain high polyphenols, the pharmacological studies of the calyx of Diospyros kaki Thunb (DKC) have not been elucidated in detail. We elucidated anti-cancer activity and potential molecular mechanism of DKC against human colorectal cancer cells. The detection approaches has been advanced, the incidence of human colorectal cancer with high morbidity and mortality rate remains high [1]. The annual incidence of human colorectal cancer is estimated to be ~1 million, with ~500,000 mortalities [2]. Many plants have been reported to exert anti-cancer activity [5,6,7,8,9]
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