title: Regulation of apoptotic effects by erythrocarpine E, a cytotoxic limonoid from Chisocheton erythrocarpus in HSC-4 human oral cancer cells.
abstract: The aim of this study was to determine the cytotoxic and apoptotic effects of erythrocarpine E (CEB4), a limonoid extracted from Chisocheton erythrocarpus on human oral squamous cell carcinoma. Based on preliminary dimethyl-2-thiazolyl-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays, CEB4 treated HSC-4 cells demonstrated a cytotoxic effect and inhibited cell proliferation in a time and dose dependent manner with an IC(50) value of 4.0±1.9 µM within 24 h of treatment. CEB4 was also found to have minimal cytotoxic effects on the normal cell line, NHBE with cell viability levels maintained above 80% upon treatment. Annexin V-fluorescein isothiocyanate (FITC), poly-ADP ribose polymerase (PARP) cleavage and DNA fragmentation assay results showed that CEB4 induces apoptosis mediated cell death. Western blotting results demonstrated that the induction of apoptosis by CEB4 appeared to be mediated through regulation of the p53 signalling pathway as there was an increase in p53 phosphorylation levels. CEB4 was also found to up-regulate the pro-apoptotic protein, Bax, while down-regulating the anti-apoptotic protein, Bcl-2, suggesting the involvement of the intrinsic mitochondrial pathway. Reduced levels of initiator procaspase-9 and executioner caspase-3 zymogen were also observed following CEB4 exposure, hence indicating the involvement of cytochrome c mediated apoptosis. These results demonstrate the cytotoxic and apoptotic ability of erythrocarpine E, and suggest its potential development as a cancer chemopreventive agent.
authors.affiliation: Faculty of Science, Institute of Biological Science (Genetics and Molecular Biology), University Malaya, Kuala Lumpur, Malaysia. hasima@um.edu.my
authors.forename: Noor Hasima
authors.initials: NH
authors.lastname: Nagoor
authors.forename: Norliza
authors.initials: N
authors.lastname: Shah Jehan Muttiah
authors.forename: Chong Soon
authors.initials: CS
authors.lastname: Lim
authors.forename: Lionel L A
authors.initials: LL
authors.lastname: In
authors.forename: Khalit
authors.initials: K
authors.lastname: Mohamad
authors.forename: Khalit
authors.initials: K
authors.lastname: Mohammad
authors.forename: Khalijah
authors.initials: K
authors.lastname: Awang
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D053938:DNA Fragmentation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D056945:Hep G2 Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D036701:Limonins / Q000737:chemistry / Q000494:pharmacology*; D029582:Meliaceae / Q000737:chemistry*; D015394:Molecular Structure; D009062:Mouth Neoplasms / Q000378:metabolism / Q000473:pathology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D036701:Limonins; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; C569836:erythrocarpine E; D011065:Poly(ADP-ribose) Polymerases; D053148:Caspase 3