A novel in vitro pancreatic carcinogenesis model |
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Authors: | Kang Hyo Jin Hong Young Bin Kim Hee Jeong Yi Yong Weon Nath Raghu G Chang Young Soo Cho Ho-Chan Bae Insoo |
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Institution: | a Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, 3970 Reservoir Road, NW, Washington, DC 20057-1469, USA b Department of Radiation Medicine, Lombardi Comprehensive Cancer Center, Georgetown University, 3970 Reservoir Road, NW, Washington, DC 20057-1469, USA c WCU (World Class University) Research Center of Nanobiomedical Science, Dankook University, San 29, Anseo-Dong, Cheonan 330-714, Republic of Korea d Department of Internal Medicine, Dongsan Medical Center, Jungri-Dong, Seo-Gu, Deagu, Republic of Korea |
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Abstract: | Environmental factors (e.g., BaP) have been pointed out as one of the etiologies of pancreatic cancer. However, very limited experimental assays are available to identify pancreatic specific environmental mutagens or susceptibility genes. In this study, we have developed a simple in vitro cell culture model system that can be used to study the molecular and biochemical aspects of carcinogenesis in a near-normal immortalized pancreatic ductal epithelial cell lines. In order to demonstrate that xenobiotic stress response is intact in these cells, we employed standard molecular biology techniques. For examples, luciferase reporter and/or real-time quantitative PCR assays were used to determine stress-induced CYP1A1 and CYP1B1 gene expression. Western blotting and immunocytochemistry assays were used to demonstrate that TCDD or BaP could activate AhR signaling. For exploring the carcinogenesis mechanism, we incubated cells with 3H]BaP and determined BaP-DNA binding activity by measuring its radioactivity. BaP-DNA adduct formation was further confirmed by 32P]-postlabeling assay. Finally, we demonstrated the effects of endogenous AhR or BRCA1 in BaP-DNA adduct accumulation in our cell system. As results, no apparent BaP-DNA adduct accumulation by 32P]-postlabeling assay was found in either control-siRNA or AhR-siRNA pretreated cells. On the other hand, a significant increase of BaP-DNA adduct accumulation was found in BRCA1 knockdown cells. In conclusion, we suggest that this in vitro model may provide the feasibility for future studies on the molecular basis of pancreatic ductal cell carcinogenesis caused by dietary mutagens. |
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Keywords: | CYP1A1 cytochrome P450 family 1 subfamily A polypeptide 1 CYP1B1 cytochrome P450 family 1 subfamily B polypeptide 1 AhR arylhydrocarbon receptor TCDD 2 3 7 8-tetrachlorodibenzodioxin BaP benzo(a)pyrene AhRR arylhydrocarbon receptor repressor BRCA1 breast cancer susceptibility gene 1 |
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