Studies on the mechanisms by which pentagalloylglucose inhibits tumor cell growth and hop bitter acids induce apoptosis

• Main Authors: Wei-Jen Chen, 陳威仁
• Other Authors: 林仁混
• Format: Others
• Language: en_US
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/11072784340338715035

博士 === 國立臺灣大學 === 生物化學暨分子生物學研究所 === 92 === Pentagalloylglucose (5GG), which contains five gallate moieties linked with a glucose core by ester bond, is natural hydrolysable tannin and widely exists in many medical plants. Previous studies indicated that 5GG is a potent and specific inhibitor of NADPH dehydrogenase or xanthine oxidase. Our previous work showed that 5GG can induce human leukemic cell HL-60 undergoing apoptosis through mitochondria- and caspase-3-mediated pathway and block cell cycle progression at G1 phase in human breast caner cell line MCF-7 through inhibition of kinase activities of cyclin-dependent kinases (CDK) 4 and 2, and induction of cyclin-dependent kinase inhibitors (CKIs) p27Kip and p21Cip; yet little is known of the mechanism(s) by which 5GG induces these effects. Herein, we indicated that 5GG could potently and selectively inhibit the activities of purified 20S and 26S proteasome in vitro, 26S proteasome in Jurkat T cell lysates and downregulate the chymotrypsin-like activity of 26S proteasome in intact Jurkat T cells. The turnover of CKIs p27Kip and p21Cip necessary for cell cycle progression mediated by proteasome degradation was disrupted by treatment of human Jurkat T cells with 5GG as shown by cycloheximide treatment and in vivo pulse-chase labeling experiments, and this effect was in tight connection with suppressing proliferation of Jurkat T cells at G1 phase. Additionally, inhibition of proteasome by 5GG as well as by well-established proteasome inhibitor, MG132, resulted in accumulation of ubiquitin-tagged proteins in Jurkat T cells. Furthermore, addition of 5GG in Jurkat T cells also enhanced the stability of proteasome substrate, Bax, and raised the consequent cytochrome c release and apoptosis. Taken together, our results provide a new insight into the molecular mechanism underlying mitogenic cell cycle control down-regulated by 5GG through proteasome-mediated pathway in human leukemic cells and imply that 5GG appears to be a proteasome inhibitor. Hop bitter acids, purified from hops (Humulus lupulus L.), mainly comprise ?acids, β-acids, and their oxidation products that contribute the special aroma of the beer beverage. Our study showed that hop bitter acids had a strong growth inhibitory effect against human leukemia HL-60 cells, with an estimated IC50 value of 8.67 μg/mL, but were less effective against human histolytic lymphoma U937 cells. DNA fragmentation assay and flow cytometric analysis demonstrated that hop bitter acids induces apoptosis in HL-60 cells. The mechanisms by which hop bitter acids triggers HL-60 apoptosis might involve dissipation of mitochondrial membrane potential, cytochrome c release, and subsequent induction of pro-caspase-9 and -3 processing and activation, which resulted in the cleavage of PARP and DFF-45. Hop bitter acids also affected the expression of Bcl-2, Bcl-XL, and Bax. In comparison with the protein level of Bcl-2, the protein level of Bcl-XL was obviously decreased after hop bitter acids treatment, whereas the protein level of Bax was dramatically increased, suggesting that the control of Bcl-2 family proteins by hop bitter acids might involve in the disruption of mitochondrial integrity. Moreover, our results displayed that hop bitter acids promoted the expression of Fas and FasL before the processing and activation of pro-caspase-8 and cleavage of Bid, indicating the participation of a Fas-mediated pathway in hop bitter acids-induced apoptosis. Taken all together, our findings suggest that a certain cross-talk might exist between receptor- and mitochondria-mediated death signalings after commitment to cell death induced by hop bitter acids. The induction of apoptosis by hop bitter acids may provide a significant mechanism for their chemopreventive action.