Mechanism of patulin-induced apoptosis in human leukemia cells (HL-60).
Patulin (PAT) is a fungal secondary metabolite that exhibits potential cellular and animal toxicities. In this study, human promyelocytic leukemia (HL-60) cells were used to elucidate the mechanism and death mode associated with PAT. Morphological evidence of apoptosis, including membrane blebbing, nuclei fragmentation and DNA laddering formation was clearly observed 6h after exposure to PAT. The results of Western blotting indicated that PAT activated various processed caspases, and cleaved DFF45 and poly (ADP-ribose) polymerase (PARP) in a dose-dependent manner; it also induced a time-dependent increase in caspase 3 and 9 catalytic activities. The apoptosis mediated by PAT in HL-60 was accompanied with cytochrome c release from mitochondria and Bcl-2 expression decrease. The presence of thiol-containing compounds with PAT dramatically reduced the caspase 3 activity that was triggered by PAT; the addition of antioxidants, including mannitol and Tiron, had a similar effect. However, the suppression of p53 protein expression by RNA interference (RNAi) in human embryonic kidney (HEK293) cells did not significantly modify PAT-elicited caspase 3 activity. These findings suggest that PAT-induced apoptosis is mediated through the mitochondrial pathway without the involvement of p53; the interaction with sulfhydryl groups of macromolecules by PAT and the subsequent generation of reactive oxygen species (ROS) plays a primary role in the apoptotic process.
Wu
Ting-Shuan
TS
Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan, ROC.
Liao
Yi-Chun
YC
Yu
Feng-Yih
FY
Chang
Chia-Hao
CH
Liu
Biing-Hui
BH
D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt / Q000494:pharmacology; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D061945:Caspase Inhibitors; D002470:Cell Survival / Q000187:drug effects; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D016166:Free Radical Scavengers / Q000494:pharmacology; D018922:HL-60 Cells; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism / Q000473:pathology; D008353:Mannitol / Q000494:pharmacology; D008858:Microscopy, Phase-Contrast; D015394:Molecular Structure; D009183:Mycotoxins / Q000737:chemistry / Q000494:pharmacology; D010365:Patulin / Q000737:chemistry / Q000494:pharmacology*; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D016166:Free Radical Scavengers; D009183:Mycotoxins; D016159:Tumor Suppressor Protein p53; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D008353:Mannitol; D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; D045304:Cytochromes c; D010365:Patulin; D053148:Caspase 3; D053453:Caspase 9