Aristolochic acid I-induced DNA damage and cell cycle arrest in renal tubular epithelial cells in vitro.
DNA damage is a critical event preceding cellular apoptosis or necrosis. This study was carried out to investigate the effect of aristolochic acid I (AAI) on DNA damage and cell cycle in porcine proximal tubular epithelial cell lines (LLC-PK1 cells). LLC-PK1 cells were stimulated with AAI at the concentrations of 80, 320, and 1,280 ng/ml for 24 h. DNA damage was examined by comet assay and the cell cycle was assayed by flow cytometry (FCM), cellular apoptosis and lysis were examined simultaneously. Cellular nuclear changes were observed by electron microscopy and the expression of wild-type p53 protein and mRNA were measured by FCM and RT-PCR. We found that AAI-induced DNA damage prior to apoptosis and lysis in LLC-PK1 cells in a dose-dependent manner (P<0.01). The percentage of cells in the G2/M phase that were treated with AAI (320 and 1,280 ng/ml) for 24 h increased significantly (P<0.01). Electron micrographs showed the nuclear abnormalities in AAI-treated cells. The expression of p53 protein and mRNA did not change in the AAI-treated cells. AAI may cause DNA damage and cell cycle arrest in LLC-PK1 cells through a wild-type p53-independent pathway, prior to apoptosis or necrosis. This study on the molecular mechanism of AAI-induced toxicity may explain why tubular epithelial cells present limited proliferation and regeneration abilities in the clinical presentation of AAI-associated nephrotoxicity.
Li
Ying
Y
Research Institute of Nephrology, Jinling Hospital, Nanjing University School of Medicine, 210002 Nanjing, China.
Liu
Zhihong
Z
Guo
Xiaohua
X
Shu
Jian
J
Chen
Zhaohong
Z
Li
Leishi
L
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D034341:Aristolochic Acids / Q000633:toxicity*; D002273:Carcinogens / Q000633:toxicity*; D002453:Cell Cycle / Q000187:drug effects*; D002467:Cell Nucleus / Q000187:drug effects / Q000648:ultrastructure; D020552:Comet Assay; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D018374:LLC-PK1 Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D009336:Necrosis; D012333:RNA, Messenger / Q000378:metabolism; D013552:Swine; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D034341:Aristolochic Acids; D002273:Carcinogens; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C000228:aristolochic acid I