Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
권호 표지
DOI QR코드

DOI QR Code

NADPH oxidase inhibitor diphenyleneiodonium induces p53 expression and cell cycle arrest in several cancer cell lines

NADPH oxidase 저해제인 diphenyleneiodonium의 p53 발현 및 암세포의 성장억제에 대한 연구

  • Jo, Hong-Jae (Department of General Surgery, Pusan National University School of Medicine) ;
  • Kim, Kang-Mi (Department of Microbiology & Immunology and Medical Research Institute, Pusan National University School of Medicine) ;
  • Song, Ju-Dong (Department of Microbiology & Immunology and Medical Research Institute, Pusan National University School of Medicine) ;
  • Park, Young-Chul (Department of Microbiology & Immunology and Medical Research Institute, Pusan National University School of Medicine)
  • 조홍재 (부산대학교 의학전문대학원 외과학교실) ;
  • 김강미 (미생물학 및 면역학교실) ;
  • 송주동 (미생물학 및 면역학교실) ;
  • 박영철 (미생물학 및 면역학교실)
  • Published : 2007.06.25
Download PDF
⟨ Previous Next ⟩

Abstract

The Diphenyleneiodonium (DPI) is widely used as an inhibitor of flavoenzymes, particularly NADPH oxidase. In this study, we investigated the effect of DPI on the cell growth progression of human colon cancer cells HCT-116 (wild-type p53), HT-29 (p53 mutant) and human breast cancer cells MCF-7 (wild-type p53). DPI treatment in cancer cells evoked a dose- and time-dependent growth inhibition, and also induced the cell cycle arrest in C2/M phase. The peak of cell population arrested in C2/M phase was observed at12 hr after treatment of DPI. In addition, DPI significantly induced the expression of p53, which induces proapoptotic genes in response to DNA damage or irreparable cell cycle arrest, at 6 hr in DPI-stimulated cells. However, a catechol apocynin, which inhibits the assembly of NADPH oxidase, did not induce p53 expression. This suggest that p53 expression induced by DPI is not associated with the inhibition of NADPH oxidase. In conclusion, we suggest that DPI induces the expression of wild-type p53 by ROS-in-dependent mechanism in several cancer cells, and upregulated p53 may be involved in regulatory mechanisms for growth inhibition and cell cycle arrest at C2/M phase in DPI-stimulated cells.

Diphenyleneiodonium (DPI)는 NADPH oxidase 같은 flavoenzymes의 저해제로써 널리 사용되고 있다. 본 연구에서는 인간 대장암 세포주 HCT-116 (wild-type p53)와 HT-29 (p53 mutant) 및 인간 유방암 세포주인 MCF-7(wild-type p53)의 세포성장 과정에서의 DPI의 효과를 살펴보았다. DPI는 농도 및 시간 의존적으로 암세포주의성장을 막았으며 G2/M phase에서 cell cycle arrest를 일으켰다. Cell cycle arrest의 가장 높은 값은 DPI 처리후 12 시간에서 관찰할 수 있었다. 한편 DPI는 아폽토시스 그리고 cell cycle arres 에 관여하는 유전자 발현에 관여하는 p53의 표현을 크게 증가시켰으며, 이는 DPI처리 후 6시간 후 부터 관찰할 수 있었다. 그러나 NADPH oxidase의 조합을 억제하는 catechol 계인 apocynin은 p53의 발현을 유도하지 못하였다. 이것은 DPI에 의해 유도되는 p53의 발현증가는 NADPH oxidase활성의 저해와 관련되어 있지 않다는 것을 의미한다. 결론적으로 DPI는 HCT-116, HCT-15 및 MCF-7 암세포주에서 ROS에 비 의존적으로 wild-type p53 발현의 증가를 유도하며, 이 증가된 p53은 DPI에 의해 유도되는 성장 억제 및 C2/M phase에서의 cell cycle arrset과정의 조절기전에 관여한다는 것을 시사한다.

Keywords

References

  1. Chen, Y., Z. H. Miao, W. M. Zhao and J. Ding. 2005. The p53 pathway is synergized by p38 MAPK signaling to mediate 11,11'-dideoxyverticillin-induced $G_2/M $arrest. FEBS Lett. 579, 3683-3690 https://doi.org/10.1016/j.febslet.2005.05.053
  2. Cross, A. R. and O. T. Jones. 1986. The effect of the inhibitor diphenylene iodonium on the superoxide-generating sys tem of neutrophils. Specific labelling of a component polypeptide of the oxidase. Biochem. J. 237, 111-116 https://doi.org/10.1042/bj2370111
  3. Doussiere, J., J. Gaillard and P. V. Vignais. 1999. The heme component of the neutrophil NADPH oxidase complex is a target for aryliodonium compounds. Biochemistry 38, 3694-3703 https://doi.org/10.1021/bi9823481
  4. Eastman, A. 2004. Cell cycle checkpoints and their impact on anticancer therapeutic strategies. J. Cell. Biochem. 91, 223-231 https://doi.org/10.1002/jcb.10699
  5. Finkel, T. 2000. Redox-dependent signal transduction. FEBS Lett. 476, 52-54 https://doi.org/10.1016/S0014-5793(00)01819-6
  6. Flatt, P. M., L. J. Tang, C. D. Scatena, S. T. Szak and J. A. Pietenpol. 2000. p53 regulation of $G_2$ checkpoint is retinoblastoma protein dependent. Mol. Cell. Biol. 20, 4210- 4223 https://doi.org/10.1128/MCB.20.12.4210-4223.2000
  7. Ianzini, F., A. Bertoldo, E. A. Kosmacek, S. L. Phillips and M. A. Mackey. 2006. Lack of p53 function promotes radiation- induced mitotic catastrophe in mouse embryonic fibroblast cells. Cancer Cell Int. 6, 11 (p1-8) https://doi.org/10.1186/1475-2867-6-1
  8. Levine, A. J. 1997. p53, the cellular gatekeeper for growth and division. Cell 88, 323-331
  9. Li, N., K. Ragheb, G. Lawler, J. Sturgis, B. Rajwa, J. A. Melendez and J. P. Robinson. 2003. DPI induces mitochondrial superoxide-mediated apoptosis. Free Radic. Biol. Med. 34, 465-477 https://doi.org/10.1016/S0891-5849(02)01325-4
  10. Li, Y. and M. A. Trush. 1998. Diphenyleneiodonium, an NAD(P)H oxidase inhibitor, also potently inhibits mitochondrial reactive oxygen species production. Biochem. Biophys. Res. Commun. 253, 295-299 https://doi.org/10.1006/bbrc.1998.9729
  11. Maya, R., M. Balass, S. T. Kim, D. Shkedy, J. F. Leal, O. Shifman, M. Moas, T. Buschmann, Z. Ronai, Y. Shiloh, M. B. Kastan, E. Katzir and M. Oren. 2001. ATM-dependent phosphorylation of Mdm2 on serine 395: role in p53 activation by DNA damage. Genes Dev. 15, 1067-1077. https://doi.org/10.1101/gad.886901
  12. Meng, L. H., H. Zhang, L. Hayward, H. Takemura, R. G. Shao and Y. Pommier. 2004. Tetrandrine induces early $G_1$ arrest in human colon carcinoma cells by down-regulating the activity and inducing the degradation of $G_1$-S-specific cyclin-dependent kinases and by inducing p53 and $p21^{Cip1}$. Cancer Res. 64, 9086-9092 https://doi.org/10.1158/0008-5472.CAN-04-0313
  13. Nakamura, Y., K. Tsuji, M. Shuto, K. Ogita, Y. Yoneda, K. Shimamoto, T. Shibata and K. Kataoka. 1997. Protection by diphenyliodonium against glutamate neurotoxicity due to blocking of N-methyl-D-aspartate receptors. Neuroscience 76, 459-466 https://doi.org/10.1016/S0306-4522(96)00375-2
  14. O'Donnell, V. B., D. G. Tew, O. T. Jones and P. J. England. 1993. Studies on the inhibitory mechanism of iodonium compounds with special reference to neutrophil NADPH oxidase. Biochem. J. 290, 41-49 https://doi.org/10.1042/bj2900041
  15. Ormerod, M. G. 1990. pp. 69-81, Flow cytometry: A practical approach, Oxford University Press, New York
  16. Pandian, R. P., V. K. Kutala, A. Liaugminas, N. L. Parinandi and P. Kuppusamy. 2005. Lipopolysaccharideinduced alterations in oxygen consumption and radical generation in endothelial cells. Mol. Cell. Biochem. 278, 119-127-172
  17. Park, B. S., Y. S. Song, S. B. Yee, B. G. Lee, S. Y. Seo, Y. C. Park, J. M. Kim, H. M. Kim and Y. H. Yoo. 2005. Phospho-ser 15-p53 translocates into mitochondria and interacts with Bcl-2 and Bcl-xL in eugenol-induced apoptosis. Apoptosis 10, 193-200 https://doi.org/10.1007/s10495-005-6074-7
  18. Riganti, C., E. Gazzano, M. Polimeni, C. Costamagna, A. Bosia and D. Ghigo. 2004. Diphenyleneiodonium inhibits the cell redox metabolism and induces oxidative stress. J. Biol. Chem. 279, 47726-47731 https://doi.org/10.1074/jbc.M406314200
  19. Sanders, S. A., R. Eisenthal and R. Harrison. 1997. NADH oxidase activity of human xanthine oxidoreductase--generation of superoxide anion. Eur. J. Biochem. 245, 541-548 https://doi.org/10.1111/j.1432-1033.1997.00541.x
  20. Scaife, R.M. (2005) Selective and irreversible cell cycle inhibition by diphenyleneiodonium. Mol. Cancer Ther. 4, 876-884 https://doi.org/10.1158/1535-7163.MCT-05-0009
  21. Shuto, M., K. Ogita and Y. Yoneda. 1997. Protection by diphenyliodonium against glutamate neurotoxicity due to blocking of N-methyl-D-aspartate receptors. Neurochem. Int. 31, 73-82 https://doi.org/10.1016/S0197-0186(96)00140-4
  22. Stuehr, D. J., O. A. Fasehun, N. S. Kwon, S. S. Gross, J. A. Gonzalez, R. Levi and C. F. Nathan. 1991. Inhibition of macrophage and endothelial cell nitric oxide synthase by diphenyleneiodonium and its analogs. FASEB J. 5, 98-103 https://doi.org/10.1096/fasebj.5.1.1703974
  23. Tew, D. G. 1993. Inhibition of cytochrome P450 reductase by the diphenyliodonium cation. Kinetic analysis and covalent modifications. Biochemistry 32, 10209-10215 https://doi.org/10.1021/bi00089a042
  24. Wang, X. W., Q. Zhan, J. D. Coursen, M. A. Khan, H. U. Kontny, L. Yu, M. C. Hollander, P. M. O'Connor, A. J. Fornace and C. C. Harris. 1999. GADD45 induction of a G2/M cell cycle checkpoint. Proc. Natl. Acad. Sci. USA 96, 3706-3711 https://doi.org/10.1073/pnas.96.7.3706
  25. Weir, E. K., C. N. Wyatt, H. L. Reeve, J. Huang, S. L. Archer and C. Peers. 1994. Diphenyleneiodonium inhibits both potassium and calcium currents in isolated pulmonary artery smooth muscle cells. J. Appl. Physiol. 76, 2611-2615 https://doi.org/10.1152/jappl.1994.76.6.2611