Advertisement

Cell Biology and Toxicology

, Volume 31, Issue 2, pp 121–130 | Cite as

Phlda3, a urine-detectable protein, causes p53 accumulation in renal tubular cells injured by cisplatin

  • Chan Gyu Lee
  • Yoon Jong Kang
  • Hyung Sik Kim
  • Aree Moon
  • Sang Geon Kim
Article

Abstract

Measurable indicators of renal injury are required for the assessment of kidney function after toxicant challenge. In our previous study, pleckstrin homology-like domain, family A, member 3 (Phlda3) was a most greatly up-regulated molecule downstream from p53, culminating with kidney tubular injury. This study investigated the positive feedforward effect of Phlda3 on p53 in an effort to explain the largest increase of Phlda3 in injured tubules and the potential of its urine excretion. qRT-PCR assays confirmed a rapid and substantial increase in Phlda3 messenger RNA (mRNA) in the kidney cortex of mice treated with a single dose of cisplatin. Cisplatin overexpression of Phlda3 was verified by gene set analyses of three different microarray databases. In the immunohistochemistry, Phlda3 staining intensities were augmented in the tubules as kidney injury worsened. Moreover, the urinary content of Phlda3 was increased after cisplatin treatment, as were those of other kidney injury markers (Kim-1 and Timp-1). By contrast, cisplatin failed to increase Phlda3 mRNA in the liver despite hepatocyte necrosis and ensuing increases in serum transaminase activities. In NRK52E tubular cells, siRNA knockdown of Phlda3 enhanced the ability of cisplatin to increase p-Mdm2 presumably via Akt, enforcing the interaction between Mdm2 and p53. Consistently, a deficiency in Phlda3 abrogated p53 increase by cisplatin, indicating that Phlda3 promotes p53 accumulation. Phlda3 overexpression had the opposite effect. In addition, treatment with cyclosporine A or CdCl2, other nephrotoxicants, increased Phlda3 mRNA and protein levels in NRK52E cells, as did cisplatin treatment. Overall, Phlda3 may cause p53 accumulation through a feedforward pathway, facilitating tubular injury and its urine excretion.

Keywords

Biomarker Cisplatin p53 Phlda3 Renal tubular injury 

Abbreviations

ALT

Alanine aminotransferase

AST

Aspartate aminotransferase

BUN

Blood urea nitrogen

Kim-1

Kidney injury molecule 1

NAG

N-acetyl-beta-d-glucosaminidase

Phlda3

Pleckstrin homology-like domain

family A

Member 3

qRT-PCR

Quantitative real-time polymerase chain reaction

SCr

Serum creatinine

Timp-1

TIMP metallopeptidase inhibitor 1

Notes

Acknowledgments

This work was supported by a grant [10182KFDA992] from Korea Food & Drug Administration in 2010–2012. The authors wish to thank Dr. Se Jin Hwang in the College of Medicine, Hanyang University, for histological evaluation.

Conflict of interest

The authors declare that they have no conflict of interest.

Author contributions

The overall study was conceived and designed by CGL and SGK; CGL analyzed data; AM and HSK contributed reagents or analysis tools; CGL and SGK wrote the paper.

Supplementary material

10565_2015_9299_Fig5_ESM.jpg (55 kb)
Supplementary Figure 1

Immunoprecipitation and immunoblotting analysis for Mdm2 and p53. NRK52E cells were treated with 30 μM cisplatin for the indicated times. (JPEG 55 kb)

10565_2015_9299_MOESM1_ESM.tif (153 kb)
High Resolution Image (TIFF 152 kb)

References

  1. Amin RP, Vickers AE, Sistare F, Thompson KL, Roman RJ, Lawton M, et al. Identification of putative gene based markers of renal toxicity. Environ Health Perspect. 2004;112:465–79.CrossRefPubMedCentralPubMedGoogle Scholar
  2. Brenner BM, Falchuk KH, Keimowitz RI, Berliner RW. The relationship between peritubular capillary protein concentration and fluid reabsorption by the renal proximal tubule. J Clin Invest. 1969;48:1519–31.CrossRefPubMedCentralPubMedGoogle Scholar
  3. Cavalli F, Tschopp L, Sonntag RW, Zimmermann A. A case of liver toxicity following cis-dichlorodiammineplatinum(II) treatment. Cancer Treat Rep. 1978;62:2125–6.PubMedGoogle Scholar
  4. Cersosimo RJ. Hepatotoxicity associated with cisplatin chemotherapy. Ann Pharmacother. 1993;27:438–41.PubMedGoogle Scholar
  5. Ciarimboli G, Ludwig T, Lang D, Pavenstädt H, Koepsell H, Piechota HJ, et al. Cisplatin nephrotoxicity is critically mediated via the human organic cation transporter 2. Am J Pathol. 2005;167:1477–84.CrossRefPubMedCentralPubMedGoogle Scholar
  6. Datta K, Franke TF, Chan TO, Makris A, Yang SI, Kaplan DR, et al. AH/PH domain-mediated interaction between Akt molecules and its potential role in Akt regulation. Mol Cell Biol. 1995;15:2304–10.PubMedCentralPubMedGoogle Scholar
  7. Devarajan P. Emerging urinary biomarkers in the diagnosis of acute kidney injury. Expert Opin Med Diagn. 2008;2:387–98.CrossRefPubMedCentralPubMedGoogle Scholar
  8. Dieterle F, Sistare F, Goodsaid F, Papaluca M, Ozer JS, Webb CP, et al. Renal biomarker qualification submission: a dialog between the FDA-EMEA and predictive safety testing consortium. Nat Biotechnol. 2010;28:455–62.CrossRefPubMedGoogle Scholar
  9. Fielden MR, Eynon BP, Natsoulis G, Jarnagin K, Banas D, Kolaja KL. A gene expression signature that predicts the future onset of drug-induced renal tubular toxicity. Toxicol Pathol. 2005;33:675–83.CrossRefPubMedGoogle Scholar
  10. Fox BC, Devonshire AS, Schutte ME, Foy CA, Minguez J, Przyborski S, et al. Validation of reference gene stability for APAP hepatotoxicity studies in different in vitro systems and identification of novel potential toxicity biomarkers. Toxicol In Vitro. 2010;24:1962–70.CrossRefPubMedGoogle Scholar
  11. Jiang M, Yi X, Hsu S, Wang CY, Dong Z. Role of p53 in cisplatin-induced tubular cell apoptosis: dependence on p53 transcriptional activity. Am J Physiol Ren Physiol. 2004;287:F1140–7.CrossRefGoogle Scholar
  12. Kawase T, Ohki R, Shibata T, Tsutsumi S, Kamimura N, Inazawa J, et al. PH domain-only protein PHLDA3 is a p53-regulated repressor of Akt. Cell. 2009;136:535–50.CrossRefPubMedGoogle Scholar
  13. Kim SG. Kidney: Toxicological assessment. 1st ed. London: CRC Press; 2013.Google Scholar
  14. Lee CG, Kim JG, Kim HJ, Kwon HK, Cho IJ, Choi DW, et al. Discovery of an integrative network of microRNAs and transcriptomics changes for acute kidney injury. Kidney Int. 2014a;86:943–53.CrossRefPubMedGoogle Scholar
  15. Lee CG, Kim YW, Kim EH, Meng Z, Huang W, Hwang SJ, et al. Farnesoid X receptor protects hepatocytes from injury by repressing miR-199a-3p, which increases levels of LKB1. Gastroenterology. 2012;142:1206–17.CrossRefPubMedCentralPubMedGoogle Scholar
  16. Lee YK, Park EY, Kim S, Son JY, Kim TH, Kang WG, et al. Evaluation of cadmium-induced nephrotoxicity using urinary metabolomic profiles in Sprague–Dawley male rats. J Toxicol Environ Health A. 2014b;77:1384–98.CrossRefPubMedGoogle Scholar
  17. Mayo LD, The DDB, PTEN. Mdm2, p53 tumor suppressor-oncoprotein network. Trends Biochem Sci. 2002;27:462–7.CrossRefPubMedGoogle Scholar
  18. Molitoris BA, Dagher PC, Sandoval RM, Campos SB, Ashush H, Fridman E, et al. siRNA targeted to p53 attenuates ischemic and cisplatin-induced acute kidney injury. J Am Soc Nephrol. 2009;20:1754–64.CrossRefPubMedCentralPubMedGoogle Scholar
  19. Ogawara Y, Kishishita S, Obata T, Isazawa Y, Suzuki T, et al. Akt enhances Mdm2-mediated ubiquitination and degradation of p53. J Biol Chem. 2002;277:21843–50.CrossRefPubMedGoogle Scholar
  20. O’Keefe K, Li H, Zhang Y. Nucleocytoplasmic shuttling of p53 is essential for MDM2-mediated cytoplasmic degradation but not ubiquitination. Mol Cell Biol. 2003;23:6396–405.CrossRefPubMedCentralPubMedGoogle Scholar
  21. Pollera CF, Ameglio F, Nardi M, Vitelli G, Marolla P. Cisplatin-induced hepatic toxicity. J Clin Oncol. 1987;5:318–9.PubMedGoogle Scholar
  22. Saxena A, Morozov P, Frank D, Musalo R, Lemmon MA, et al. Phosphoinositide binding by the pleckstrin homology domains of Ipl and Tih1. J Biol Chem. 2002;277:49935–4944.CrossRefPubMedGoogle Scholar
  23. Supavekin S, Zhang W, Kucherlapati R, Kaskel FJ, Moore LC, Devarajan P. Differential gene expression following early renal ischemia/reperfusion. Kidney Int. 2003;63:1714–24.CrossRefPubMedGoogle Scholar
  24. Togashi Y, Sakaguchi Y, Miyamoto M, Miyamoto Y. Urinary cystatin C as a biomarker for acute kidney injury and its immunohistochemical localization in kidney in the CDDP-treated rats. Exp Toxicol Pathol. 2012;64:797–805.CrossRefPubMedGoogle Scholar
  25. Tokumoto M, Fujiwara Y, Shimada A, Hasegawa T, Seko Y, Nagase H, et al. Cadmium toxicity is caused by accumulation of p53 through the down-regulation of Ube2d family genes in vitro and in vivo. J Toxicol Sci. 2011;36:191–200.CrossRefPubMedGoogle Scholar
  26. Wei Q, Dong G, Yang T, Megyesi J, Price PM, Dong Z. Activation and involvement of p53 in cisplatin-induced nephrotoxicity. Am J Physiol Ren Physiol. 2007;293:F1282–91.CrossRefGoogle Scholar
  27. Vaidya VS, Ferguson MA, Bonventre JV. Biomarkers of acute kidney injury. Annu Rev Pharmacol Toxicol. 2008;48:463–93.CrossRefPubMedCentralPubMedGoogle Scholar
  28. Vaidya VS, Ramirez V, Ichimura T, Bobadilla NA, Bonventre JV. Urinary kidney injury molecule-1: a sensitive quantitative biomarker for early detection of kidney tubular injury. Am J Physiol Ren Physiol. 2005;290:F517–29.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Chan Gyu Lee
    • 1
  • Yoon Jong Kang
    • 2
  • Hyung Sik Kim
    • 2
  • Aree Moon
    • 3
  • Sang Geon Kim
    • 1
  1. 1.College of Pharmacy and Research Institute of Pharmaceutical SciencesSeoul National UniversityGwanak-guSouth Korea
  2. 2.School of PharmacySungkyunkwan UniversitySuwonSouth Korea
  3. 3.College of PharmacyDuksung Women’s UniversitySeoulSouth Korea

Personalised recommendations