3-Oxoolean-12-en-27-oic Acid Isolated from Aceriphyllum rossii Induces Caspase-8-Dependent Apoptosis in Human Promyelocytic Leukemia HL-60 Cells
College of Pharmacy, Kyung-Hee University, Seoul, Republic of Korea.Biological & Pharmaceutical Bulletin (Impact Factor: 1.83). 02/2009; 32(1):91-8.
In the present study, we investigated the effects of 3-oxoolean-12-en-27-oic acid (3-OA) isolated from the underground parts of Aceriphyllum rossii (Saxifragaceae) on the viability and apoptosis of HL-60 human promyelocytic leukemia cells, and the mechanisms underlying its action. 3-OA-treated HL-60 cells and HeLa human cervix adenocarcinoma cells displayed several apoptotic features, such as, DNA fragmentation, DNA laddering by agarose gel electrophoresis, and hypodiploid DNA contents by flow cytometry, and 3-OA also caused the activations of caspase-8, -9 and -3. Pretreatment with z-VAD-fmk (a broad-caspase inhibitor) almost completely suppressed 3-OA-induced DNA ladder formation and hypodiploid DNA contents, thereby implicating the caspase cascade in the apoptotic process. In addition, z-IETD-fmk (a caspase-8 inhibitor) and z-DEVD-fmk (a caspase-3 inhibitor) also completely neutralized the apoptotic effect of 3-OA in HL-60 cells. Furthermore, 3-OA increased Fas-related protein contents and the mRNA expressions of Fas ligand (FasL), Fas, and Fas-associated death domain (FADD). Preincubation with anti-Fas or anti-FasL blocking antibodies completely prevented 3-OA-induced apoptosis. Taken together, these results suggest that 3-oxoolean-12-en-27-oic acid induces apoptosis by activating caspase-8 via FasL-stimulated death receptor signaling.
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ABSTRACT: In the present study, we investigated the effect of 3alpha,23-isopropylidenedioxyolean-12-en-27-oic acid (IPA), an active compound isolated from Aceriphyllum rossii, on the apoptotic activity and the molecular mechanism of the action in human cervical cancer HeLa cells. Treatment with IPA significantly increased externalization of phosphatidylserine residues and apoptotic DNA fragmentation as shown by Annexin V staining and 4',6-diamidino-2-phenylindole-dihydrochloride (DAPI) staining, respectively. In addition, IPA induced the activations of caspase-8, -9, -3, and cleavage of poly(ADP ribose) polymerase (PARP-1) in HeLa cells. Pretreatment with a specific caspase-8, -9, or -3 inhibitor neutralized the pro-apoptotic activity of IPA in HeLa cells. Furthermore, IPA was found to induce the loss of mitochondrial membrane potential, the release of cytochrome c to the cytosol, and the increased ratio of mitochondrial Bax/Bcl-2. Moreover, we demonstrated that IPA triggered endoplasmic reticulum (ER) stress, as shown by changes in cytosol-calcium level, activation of mu-calpain and caspase-12, and up-regulation of glucose-regulated protein 78 (GRP78) and growth arrest DNA damage-inducible gene 153 (GADD153). IPA-induced apoptosis was substantially reduced in the presence of an intracellular calcium chelator BAPTA/AM. Taken together, these results suggest that both mitochondrial dysfunction and ER stress contribute to IPA-induced apoptosis of human cervical cancer HeLa cells.
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ABSTRACT: 20(S)-ginsenoside Rg3 [20(S)-Rg3)], one of the main constituents isolated from Panax ginseng, has been shown to have an anti-cancer effect and to induce apoptosis by interfering with several signaling pathways. However, the molecular mechanisms of AMP-activated protein kinase (AMPK) associated with apoptosis in HT-29 colon cancer cells remain unclear. In the present study, we investigated whether 20(S)-Rg3 exerts an anti-proliferative effect and induces apoptosis by modulating the AMPK signaling pathway in HT-29 cells. 20(S)-Rg3-treated cells displayed several apoptotic features, including DNA fragmentation, proteolytic cleavage of poly (ADP-ribose) polymerase (PARP) and morphological changes. 20(S)-Rg3 down-regulated the expression of anti-apoptotic protein B-cell CLL/lymphoma 2 (Bcl2), up-regulated the expression of pro-apoptotic protein of p53 and Bcl-2-associated X protein (Bax), and caused the release of mitochondrial cytochrome c, PARP, caspase-9 and caspase-3. However, 20(S)-Rg3-induced apoptosis was completely abolished in the presence of compound C (AMPK inhibitor) or small interfering RNA for AMPK (siAMPK). In addition, STO-609 (CaMKKβ inhibitor) attenuated 20(S)-Rg3-induced AMPK activation and apoptosis. These results suggest that 20(S)-Rg3-induced apoptosis in HT-29 cells is mediated via the AMPK signaling pathway, and that 20(S)-Rg3 is capable of inducing apoptosis in colon cancer.
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