Oil A induces apoptosis of pancreatic cancer cells via caspase activation, redistribution of cell cycle and GADD expression.
OBJECTIVE
To explore the mechanisms of effects of oil A on apoptosis of human pancreatic cancer cells.


METHODS
Cellular DNA content was analyzed by flow cytometry. Western blotting was used for caspase-3 and PARP, caspase-7, caspase-9, cytochrome c, Bcl-2, Bax, Mcl-1, cyclinA, cyclin B1, cyclin D1, cyclin E, CDK2, CDK4, CDK6, P21, P27, GADD45, GADD153.


RESULTS
The caspase-3, caspase-7, and caspase-9 activities were significantly increased as well as the cleavage of caspase-3, downstream substrate poly-ADP ribose polymerase (PARP) was induced. The amount of cytochrome c in the cytosolic fraction was increased, while the amount of cytochrome c in the mitochondrial fraction was decreased after oil A treatment. The anti-apoptosis proteins Bcl-2 and Mcl-1 were decreased in parallel and Bax increased, indicating that Bcl-2 family proteins-mitochondria-caspase cascade was responsible for oil-induced apoptosis. The proportion of cells in the G0/G1 decreased in MiaPaCa-2 and AsPC-1 cells after the treatment of oil A for 24 hours. The number of cells in S phase was increased in two cancer cell lines at 24 hours. Therefore, cells were significantly accumulated in G2/M phase. The cells with a sub-G0/G1 DNA content, a hallmark of apoptosis, were seen at 24 hours both in MiaPaCa-2 and AsPC-1 cells following exposure to oil A. The expression of cyclin A and cyclin B1 was slightly decreased and cyclin D1 levels were markedly lowered in MiaPaCa-2 cells. The expression of cyclin A and cyclin B1 was markedly decreased and cyclin D1 levels were slightly lowered in AsPC-1 cells, while cyclin E was not affected and the levels of CDK2, CDK4, and CDK6 were unchanged in MiaPaCa-2 and AsPC-1 cells. In response to oil A, P21 expression was increased, but P27 expression was not affected. The expression of both GADD45 and GADD153 was increased in two cell lines following oil A treatment.


CONCLUSIONS
Oil A induces apoptosis of pancreatic cancer cells via activating caspase cascade, modifying cell cycle progress and changing cell cycle-regulating proteins and GADD expression.
Dong
Mi-Lian
ML
Affiliated Taizhou Hospital, Wenzhou Medical College, Linhai 317000, Zhejiang, Provice China. mdong2@hotmail.com
Zhu
Yue-Chun
YC
Hopkins
John V
JV
D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D022762:CCAAT-Enhancer-Binding Proteins / Q000187:drug effects / Q000378:metabolism; D020169:Caspases / Q000187:drug effects / Q000378:metabolism*; D002453:Cell Cycle / Q000187:drug effects*; D016213:Cyclins / Q000187:drug effects; D004249:DNA Damage / Q000187:drug effects; D004273:DNA, Neoplasm / Q000187:drug effects; D005434:Flow Cytometry; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008055:Lipids / Q000494:pharmacology*; D008928:Mitochondria / Q000187:drug effects / Q000473:pathology / Q000502:physiology; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000473:pathology*; D011506:Proteins / Q000187:drug effects / Q000378:metabolism; D051742:Transcription Factor CHOP; D014157:Transcription Factors / Q000187:drug effects / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000097642:GADD45 Proteins
D016588:Anticarcinogenic Agents; D022762:CCAAT-Enhancer-Binding Proteins; D020169:Caspases; D016213:Cyclins; D004273:DNA, Neoplasm; D047908:Intracellular Signaling Peptides and Proteins; D008055:Lipids; D011506:Proteins; D051742:Transcription Factor CHOP; D014157:Transcription Factors; D000097642:GADD45 Proteins; C497249:DDIT3 protein, human; C482522:Oil A