Aristolochic acid suppresses DNA repair and triggers oxidative DNA damage in human kidney proximal tubular cells.
Aristolochic acid (AA), derived from plants of the Aristolochia genus, has been proven to be associated with aristolochic acid nephropathy (AAN) and urothelial cancer in AAN patients. In this study, we used toxicogenomic analysis to clarify the molecular mechanism of AA-induced cytotoxicity in normal human kidney proximal tubular (HK-2) cells, the target cells of AA. AA induced cytotoxic effects in a dose-dependent (10, 30, 90 microM for 24 h) and time-dependent manner (30 microM for 1, 3, 6, 12 and 24 h). The cells from those experiments were then used for microarray experiments in triplicate. Among the differentially expressed genes analyzed by Limma and Ingenuity Pathway Analysis software, we found that genes in DNA repair processes were the most significantly regulated by all AA treatments. Furthermore, response to DNA damage stimulus, apoptosis, and regulation of cell cycle, were also significantly regulated by AA treatment. Among the differentially expressed genes found in the dose-response and time-course studies that were involved in these biological processes, two up-regulated (GADD45B, NAIP), and six down-regulated genes (TP53, PARP1, OGG1, ERCC1, ERCC2, and MGMT) were con-firmed by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). AA exposure also caused a down-regulation of the gene expression of anti-oxidant enzymes, such as superoxide dismutase, glutathione reductase, and glutathione peroxidase. Moreover, AA treatment led to increased frequency of DNA strand breaks, 8-hydroxydeoxyguanosine-positive nuclei, and micronuclei in a dose-dependent manner in HK-2 cells, possibly as a result of the inhibition of DNA repair. These data suggest that oxidative stress plays a role in the cytotoxicity of AA. In addition, our results provide insight into the involvement of down-regulation of DNA repair gene expression as a possible mechanism for AA-induced genotoxicity.
Chen
Ya-Yin
YY
School of Chinese Medicine, China Medical University, Taichung, Taiwan.
Chung
Jing-Gung
JG
Wu
Hsiu-Ching
HC
Bau
Da-Tian
DT
Wu
Kuen-Yuh
KY
Kao
Shung-Te
ST
Hsiang
Chien-Yun
CY
Ho
Tin-Yun
TY
Chiang
Su-Yin
SY
D034341:Aristolochic Acids / Q000494:pharmacology*; D002273:Carcinogens / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004249:DNA Damage / Q000187:drug effects*; D004260:DNA Repair / Q000187:drug effects* / Q000235:genetics; D015536:Down-Regulation / Q000187:drug effects; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000187:drug effects* / Q000378:metabolism; D009152:Mutagenicity Tests; D020411:Oligonucleotide Array Sequence Analysis; D018384:Oxidative Stress / Q000187:drug effects* / Q000235:genetics
D034341:Aristolochic Acids; D002273:Carcinogens; C000228:aristolochic acid I