title: Agarol, an ergosterol derivative from Agaricus blazei, induces caspase-independent apoptosis in human cancer cells.
abstract: Agaricus blazei (A. blazei) is a mushroom with many biological effects and active ingredients. We purified a tumoricidal substance from A. blazei, an ergosterol derivative, and named it 'Agarol'. Cytotoxic effects of Agarol were determined by the MTT assay using A549, MKN45, HSC-3, and HSC-4 human carcinoma cell lines treated with Agarol. Apoptosis was detected by flow cytometry analysis. Reactive oxygen species (ROS) levels and mitochondria membrane potential (∆ψm) were also determined by flow cytometry. Western blot analysis was used to quantify the expression of apoptosis-related proteins. Agarol predominantly induced apoptosis in two p53-wild cell lines (A549 and MKN45) compared to the other p53-mutant cell lines (HSC-3 and HSC-4). Further mechanistic studies revealed that induction of apoptosis is associated with increased generation of ROS, reduced ∆ψm, release of apoptosis-inducing factor (AIF) from the mitochondria to the cytosol, upregulation of Bax, and downregulation of Bcl-2. Caspase-3 activities did not increase, and z-VAD-fmk, a caspase inhibitor, did not inhibit the Agarol-induced apoptosis. These findings indicate that Agarol induces caspase-independent apoptosis in human carcinoma cells through a mitochondrial pathway. The in vivo anticancer activity of Agarol was confirmed in a xenograft murine model. This study suggests a molecular mechanism by which Agarol induces apoptosis in human carcinoma cells and indicates the potential use of Agarol as an anticancer agent.
authors.affiliation: Mushroom Research Laboratory, Hokuto Corporation, Nagano 381-0008, Japan.
authors.forename: Takamitsu
authors.initials: T
authors.lastname: Shimizu
authors.affiliation: Mushroom Research Laboratory, Hokuto Corporation, Nagano 381-0008, Japan.
authors.forename: Junya
authors.initials: J
authors.lastname: Kawai
authors.affiliation: Mushroom Research Laboratory, Hokuto Corporation, Nagano 381-0008, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Ouchi
authors.affiliation: Laboratory of Natural Product Chemistry, Tohoku University Graduate School of Pharmaceutical Sciences, Sendai 980-8578, Japan.
authors.forename: Haruhisa
authors.initials: H
authors.lastname: Kikuchi
authors.affiliation: Laboratory of Natural Product Chemistry, Tohoku University Graduate School of Pharmaceutical Sciences, Sendai 980-8578, Japan.
authors.forename: Yoshiteru
authors.initials: Y
authors.lastname: Osima
authors.affiliation: Department of Microbiology and Immunology, Tohoku University Graduate School of Dentistry, Sendai 980-8575, Japan.
authors.forename: Rikiishi
authors.initials: R
authors.lastname: Hidemi
mesh_terms: D000364:Agaricus / Q000737:chemistry*; D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000494:pharmacology; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004875:Ergosterol / Q000008:administration & dosage* / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects*; D051379:Mice; D009369:Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D017382:Reactive Oxygen Species; D020169:Caspases; D004875:Ergosterol