Arecoline-induced growth arrest and p21WAF1 expression are dependent on p53 in rat hepatocytes.
Betel-quid use is associated with the risk of liver cirrhosis and hepatocellular carcinoma and arecoline, the major alkaloid of betel-quid, is hepatotoxic in mice. Therefore, we studied the cytotoxic and genotoxic effects of arecoline in normal rat hepatocytes (Clone-9 cells). Arecoline dose-dependently (0.1-1mM) decreased cell cycle-dependent proliferation while inducing DNA damage at 24h. Moreover, arecoline (1mM)-induced apoptosis and necrosis at 24h. Arecoline dose-dependently (0.1-0.5mM) increased transforming growth factor-beta (TGF-beta) mRNA, gene transcription and bioactivity and neutralizing TGF-beta antibody attenuated arecoline (0.5mM)-inhibited cell proliferation at 24h. Arecoline (0.5mM) also increased p21(WAF1) protein expression and p21(WAF1) gene transcription. Moreover, arecoline (0.5mM) time-dependently (8-24h) increased p53 serine 15 phosphorylation. Pifithrin-alpha (p53 inhibitor) and the loss of the two p53-binding elements in the p21(WAF1) gene promoter attenuated arecoline-induced p21(WAF1) gene transcription at 24h. Pifithrin-alpha also attenuated arecoline (0.5mM)-inhibited cell proliferation at 24h. We concluded that arecoline induces cytotoxicity, DNA damage, G(0)/G(1) cell cycle arrest, TGF-beta1, p21(WAF1) and activates p53 in Clone-9 cells. Moreover, arecoline-induced p21(WAF1) is dependent on p53 while arecoline-inhibited growth is dependent on both TGF-beta and p53.
Chou
Wen-Wen
WW
Graduate Institute of Medicine, Faculty of Medicine, Kaohsiung Medical University, Taiwan.
Guh
Jinn-Yuh
JY
Tsai
Jung-Fa
JF
Hwang
Chi-Ching
CC
Chen
Hung-Chun
HC
Huang
Jau-Shyang
JS
Yang
Yu-Lin
YL
Hung
Wen-Chun
WC
Chuang
Lea-Yea
LY
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001114:Areca / Q000737:chemistry / Q000633:toxicity; D001115:Arecoline / Q000633:toxicity*; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000096:biosynthesis / Q000235:genetics; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D009153:Mutagens / Q000633:toxicity*; D051381:Rats; D013997:Time Factors; D014158:Transcription, Genetic; D016212:Transforming Growth Factor beta / Q000096:biosynthesis / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
C493958:Cdkn1a protein, rat; D050759:Cyclin-Dependent Kinase Inhibitor p21; D009153:Mutagens; D016212:Transforming Growth Factor beta; D016159:Tumor Suppressor Protein p53; D001115:Arecoline