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Aristolochic acid I-induced DNA damage and cell cycle arrest in renal tubular epithelial cells in vitro

  • Genotoxicity and Carcinogenicity
  • Published:
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Abstract

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.

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References

  • Arlt VM, Wiessler M, Schmeiser HH (2000) Using polymerase arrest to detect DNA binding specificity of aristolochic acid in the mouse H-ras gene. Carcinogenesis 21:235–242

    Article  PubMed  CAS  Google Scholar 

  • Arlt VM, Schmeiser HH, Pfeifer GP (2001) Sequence-specific detection of aristolochic acid-DNA adducts in the human p53 gene by terminal transferase-dependent PCR. Carcinogenesis 22:133–140

    Article  PubMed  CAS  Google Scholar 

  • Arlt VM, Stiborová M, Schmeiser HH (2002) Aristolochic acid as a probable cancer hazard in herbal remedies: a review. Mutagenesis 17:265–277

    Article  PubMed  CAS  Google Scholar 

  • Bache M, Dunst J, Wurl P, Frode D, Meye A, Schmidt H, Rath FW, Taubert H (1999) G2/M checkpoint is p53-dependent and independent after irradiation in five human sarcoma cell lines. Anticancer Res 19:1827–1832

    PubMed  CAS  Google Scholar 

  • Balachandran P, Wei F, Lin RC, Khan IA, Pasco DS (2005) Structure activity relationships of aristolochic acid analogues: toxicity in cultured renal epithelial cells. Kidney Int 67:1797–1805

    Article  PubMed  CAS  Google Scholar 

  • But PP, Ma SC (1999) Chinese-herb nephropathy. Lancet 354:1731–1732

    Article  PubMed  CAS  Google Scholar 

  • Chang CH, Wang YM, Yang AH, Chiang SS (2001) Rapidly progressive interstitial renal fibrosis associated with Chinese herbal medications. Am J Nephrol 21:441–448

    Article  PubMed  CAS  Google Scholar 

  • Clifford B, Beljin M, Stark GR, Taylor WR (2003) G2 arrest in response to topoisomerase II inhibitors: the role of p53. Cancer Res 63:4074–4081

    PubMed  CAS  Google Scholar 

  • Cosyns JP (2002) When is “aristolochic acid nephropathy” more accurate than “Chinese herbs nephropathy”? Kidney Int 61:1178

    Article  PubMed  Google Scholar 

  • Cosyns JP (2003) Aristolochic acid and ‘Chinese herbs nephropathy’: a review of the evidence to date. Drug Saf 26:33–48

    Article  PubMed  CAS  Google Scholar 

  • Cosyns JP, Jadoul M, Squifflet JP, Wese FX, van Ypersele de Strihou C (1999) Urothelial lesions in Chinese-herb nephropathy. Am J Kidney Dis 33:1011–1017

    Article  PubMed  CAS  Google Scholar 

  • De Broe ME (1999) On a nephrotoxic and carcinogenic slimming regimen. Am J Kidney Dis 33:1171–1173

    Article  PubMed  Google Scholar 

  • Depierreux M, Van Damme B, Vanden Houte K, Vanherweghem JL (1994) Pathologic aspects of a newly described nephropathy related to the prolonged use of Chinese herbs. Am J Kidney Dis 24:172–180

    PubMed  CAS  Google Scholar 

  • Gao R, Zheng F, Liu Y, Zheng D, Li X, Bo Y, Liu Y (2000) Aristolochic acid I-induced apoptosis in LLC-PK1 cells and amelioration of the apoptotic damage by calcium antagonist. Chin Med J (Engl) 113:418–424

    CAS  Google Scholar 

  • Guh JY, Yang ML, Yang YL, Chang CC, Chuang LY (1996) Captopril reverses high-glucose-induced growth effects on LLC-PK1 cells partly by decreasing transforming growth factor-beta receptor protein expressions. J Am Soc Nephrol 7:1207–1215

    PubMed  CAS  Google Scholar 

  • Hattori H, Kuroda M, Ishida T, Shinmura K, Nagai S, Mukai K, Imakiire A (2004) Human DNA damage checkpoints and their relevance to soft tissure sarcoma. Pathol Int 54:26–31

    Article  PubMed  CAS  Google Scholar 

  • Jagus R, Joshi B, Barber GN (1999) PRK, apoptosis and cancer. Int J Biochem Cell Biol 31:123–138

    Article  PubMed  CAS  Google Scholar 

  • Kabanda A, Jadoul M, Lauwerys R, Bernard A, van Ypersele de Strihou C (1995) Low molecular weight proteinuria in Chinese herbs nephropathy. Kidney Int 48:1571–1576

    Article  PubMed  CAS  Google Scholar 

  • Kaufmann WK, Paules RS (1996) DNA damage and cell cycle checkpoints. FASEB J 10:238–247

    PubMed  CAS  Google Scholar 

  • Lally C, Healy E, Ryan MP (1999) Cyclosporine A-induced cell cycle arrest and cell death in renal epithelial cells. Kidney Int 56:1254–1257

    Article  PubMed  CAS  Google Scholar 

  • Lebeau C, Arlt VM, Schmeiser HH, Boom A, Verroust PJ, Devuyst O, Beauwens R (2001) Aristolochic acid impedes endocytosis and induces DNA adducts in proximal tubule cells. Kidney Int 60:1332–1342

    Article  PubMed  CAS  Google Scholar 

  • Li X, Yang L, Yu Y (2001) An analysis of the clinical and pathological characteristics of mu-tong (a Chinese herb) induced tubulointerstitial nephropathy. Zhonghua Nei Ke Za Zhi 40:681–687

    PubMed  CAS  Google Scholar 

  • Lord GM, Tagore R, Cook T, Gower P, Pusey CD (1999) Nephropathy caused by Chinese herbs in the UK. Lancet 354:481–482

    Article  PubMed  CAS  Google Scholar 

  • McCauley J, Farkus Z, Prasad SJ, Plummer HA, Murray SA (1991) Cyclosporine and FK506 induced inhibition of renal epithelial cell proliferation. Transplant Proc 23:2829–2830

    PubMed  CAS  Google Scholar 

  • Minemoto Y, Uchida S, Ohtsubo M, Shimura M, Sasagawa T, Hirata M, Nakagama H, Ishizaka Y, Yamashita K (2003) Loss of p53 induces M-phase retardation following G2 DNA damage checkpoint abrogation. Arch Biochem Biophys 412:13–19

    Article  PubMed  CAS  Google Scholar 

  • Nortier JL, Deschodt-Lanckman MM, Simon S, Thielemans NO, de Prez EG, Depierreux MF, Tielemans CL, Richard C, Lauwerys RR, Bernard AM, Vanherweghem JL (1997) Proximal tubular injury in Chinese herbs nephropathy: monitoring by neutral endopeptidase enzymuria. Kidney Int 51:288–293

    Article  PubMed  CAS  Google Scholar 

  • Nortier JL, Martinez MC, Schmeiser HH, Arlt VM, Bieler CA, Petein M, Depierreux MF, De Pauw L, Abramowicz D, Vereerstraeten P, Vanherweghem JL (2000) Urothelial carcinoma associated with the use of a Chinese herb (Aristolochia fangchi). N Engl J Med 342:1686–1692

    Article  PubMed  CAS  Google Scholar 

  • Pippin JW, Durvasula R, Petermann A, Hiromura K, Couser WG, Shankland SJ (2003) DNA damage is a novel response to sublytic complement C5b-9-induced injury in podocytes. J Clin Invest 111:877–885

    Article  PubMed  CAS  Google Scholar 

  • Saldeen J, Tillmar L, Karlsson E, Welsh N (2003) Nicotinamide- and caspase-mediated inhibition of poly(ADP-ribose) polymerase are associated with p53-independent cell cycle (G2) arrest and apoptosis. Mol Cell Biochem 243:113–122

    Article  PubMed  CAS  Google Scholar 

  • Sancar A, Lindsey-Boltz LA, Unsal-Kacmaz K, Linn S (2004) Molecular mechanisms of mammalian DNA repair and the DNA damage checkpoints. Annu Rev Biochem 73:39–85

    Article  PubMed  CAS  Google Scholar 

  • Schaneberg BT, Khan IA (2004) Analysis of products suspected of containing Aristolochia or Asarum species. J Ethnopharmacol 94:245–249

    Article  PubMed  CAS  Google Scholar 

  • Schmeiser HH, Bieler CA, Wiessler M, van Ypersele de Strihou C, Cosyns JP (1996) Detection of DNA adducts formed by aristolochic acid in renal tissue from patients with Chinese herbs nephropathy. Cancer Res 56:2025–2028

    PubMed  CAS  Google Scholar 

  • Schmeiser HH, Frei E, Wiessler M, Stiborova M (1997) Comparison of DNA adduct formation by aristolochic acids in various in vitro activation systems by 32P-post-labelling: evidence for reductive activation by peroxidases. Carcinogenesis 18:1055–1062

    Article  PubMed  CAS  Google Scholar 

  • Stark GR, Taylor WR ( 2004) Analyzing the G2/M checkpoint. Methods Mol Biol 280:51–82

    PubMed  CAS  Google Scholar 

  • Su Z, Xu S, Zheng F, Li Y (2002) Aristolochic acid induced transdifferentiation and apoptosis in human tubular epithelial cells in vitro. Zhong hua Yu Fang Yi Xue Za Zhi 36:301–304

    CAS  Google Scholar 

  • Tanaka A, Nishida R, Yoshida T, Koshikawa M, Goto M, Kuwahara T (2001) Outbreak of Chinese herb nephropathy in Japan: are there any differences from Belgium? Intern Med 40:296–300

    Article  PubMed  CAS  Google Scholar 

  • Taylor WR, Stark GR (2001) Regulation of the G2/M transition by p53. Oncogene 20:1803–1815

    Article  PubMed  CAS  Google Scholar 

  • Tice RR, Agurell E, Anderson D, Burlinson B, Hartmann A, Kobayashi H, Miyamae Y, Rojas E, Ryu JC, Sasaki YF (2000) Single cell gel/comet assay: guidelines for in vitro and in vivo genetic toxicology testing. Environ Mol Mutagen 35:206–221

    Article  PubMed  CAS  Google Scholar 

  • Van Engeland M, Ramaekers FC, Schutte B, Reutelingsperger CP (1996) A novel assay to measure loss of plasma membrane asymmetry during apoptosis of adherent cells in culture. Cytometry 24:131–139

    Article  PubMed  Google Scholar 

  • Vincent F, Deplanque G, Ceraline J, Duclos B, Bergerat JP (1999) p53-Independent regulation of cyclin B1 in normal human fibroblasts during UV-induced G2-arrest. Biol Cell 91:665–674

    Article  PubMed  CAS  Google Scholar 

  • Zheng F, Wen X, Li X, Gao R, Zhang X (2000) Synergistic effect of monocyte chemotactic protein-1 and aristolochic acid I on transdifferentiation of human tubular epithelial cells in vitro. Zhonghua Nei Ke Za Zhi 39:831–834

    PubMed  CAS  Google Scholar 

Download references

Acknowledgement

We are grateful to Biomed Proofreading for their language revision.

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Author notes

  1. Ying Li

    Present address: Division of Nephrology, The Second Xiangya Hospital, Central South University, 410011, Changsha, China

Authors and Affiliations

  1. Research Institute of Nephrology, Jinling Hospital, Nanjing University School of Medicine, 210002, Nanjing, China

    Ying Li, Zhihong Liu, Xiaohua Guo, Jian Shu, Zhaohong Chen & Leishi Li

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  1. Ying Li
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  2. Zhihong Liu
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Correspondence to Zhihong Liu.

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Li, Y., Liu, Z., Guo, X. et al. Aristolochic acid I-induced DNA damage and cell cycle arrest in renal tubular epithelial cells in vitro. Arch Toxicol 80, 524–532 (2006). https://doi.org/10.1007/s00204-006-0090-4

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  • DOI: https://doi.org/10.1007/s00204-006-0090-4

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