INDUCTION OF MITOCHONDRIAL DNA DELETION BY IONIZING RADIATION IN HUMAN LUNG FIBROBLAST IMR-90 CELLS

  • Eom, Hyeon-Soo (Radiation Biotechnology Research Division, Korea Atomic Energy Research Institute) ;
  • Jung, U-Hee (Radiation Biotechnology Research Division, Korea Atomic Energy Research Institute) ;
  • Park, Hae-Ran (Radiation Biotechnology Research Division, Korea Atomic Energy Research Institute) ;
  • Jo, Sung-Kee (Radiation Biotechnology Research Division, Korea Atomic Energy Research Institute)
  • Published : 2009.06.30

Abstract

Mitochondrial DNA (mtDNA) deletion is a well-known marker for oxidative stress and aging and also contributes to their unfavorable effects in cultured cells and animal tissues. This study was conducted to investigate the effect of ionizing radiation (IR) on mtDNA deletion and the involvement of reactive oxygen species (ROS) in this process in human lung fibroblast (IMR-90) cells. Young IMR-90 cells at population doubling (PD) 39 were irradiated with $^{137}Cs$ $\gamma$-rays and the intracellular ROS level was determined by 2',7'-dichlorofluorescein diacetate (DCFH-DA) and mtDNA common deletion (4977bp) was detected by nested PCR. Old cells at PD 55 and $H_2O_2$-treated young cells were compared as the positive control. IR increased the intracellular ROS level and mtDNA 4977 bp deletion in IMR-90 cells dose-dependently. The increases of ROS level and mtDNA deletion were also observed in old cells and $H_2O_2$-treated young cells. To confirm the increased ROS level is essential for mtDNA deletion in irradiated cells, the effects of N-acetylcysteine (NAC) on IRinduced ROS and mtDNA deletion were examined. 5 mM NAC significantly attenuated the IR-induced ROS increase and mtDNA deletion. These results suggest that IR induces the mtDNA deletion and this process is mediated by ROS in IMR-90 cells.

References

  1. Packer L. Ultraviolet radiation (UVA, UVB) and skin antioxidants. In : Rice-Evans C. A. and Burdon R. H., eds. Free Radical Damage and Its Control, USA, Elsevier Science. 1994:239-253
  2. Vijayalaxmi, Reiter RJ, Herman TS, Meltz ML. Melatonin reduces gamma radiation-induced primary DNA damage in human blood lymphocytes. Mutat. Res. 1998;397:203- 208 https://doi.org/10.1016/S0027-5107(97)00211-X
  3. Evans JL, Goldfine ID, Maddux BA, Grodsky GM. Oxidative stress and stress- activated signaling pathways : a unifying hypothesis of type 2 diabetes. Endocr Rev. 2002;23:599- 622 https://doi.org/10.1210/er.2001-0039
  4. Turrens JF. Mitochondrial formation of reactive oxygen species. J Physiol. 2003;552:335-44 https://doi.org/10.1113/jphysiol.2003.049478
  5. Harman D. Free radical theory of aging. Triangle. 1973;12:153-158
  6. Landis GN, Tower J. Superoxide dismutase evolution and life span regulation. Ageing Dev. 2005;126:907-908 https://doi.org/10.1016/j.mad.2005.05.001
  7. Corral-debrinski M, Shoffner JM, Lott MT, Wallace DC. Association of mitochondrial DNA damage with aging and coronary atherosclerotic heart disease. Mutat. Res. 1992a;275:169-180 https://doi.org/10.1016/0921-8734(92)90021-G
  8. Corral-debrinski M, Horton T, Lott MT, Shoffner JM, Beal MF, Wallace DC. Mitochondrial DNA deletions in human brain: regional variability and increase with advanced age. Nat. Genet. 1992b;2:324-329 https://doi.org/10.1038/ng1292-324
  9. Lee CM, Chung SS, Kaczkowski JM, Weindruch R, Aiken JM. Multiple mitochondrial DNA deletions associated with age in skeletal muscle of rhesus monkeys. J. Gerontol. 1993;48:B201-B205 https://doi.org/10.1093/geronj/48.6.B201
  10. Wang Y, Michikawa Y, Mallidis C, Bai Y, Woodhouse L, Yarasheski KE, Miller CA, Askanas V, Engel WK, Bhasin S, Attardi G. Muscle-specific mutations accumulate with aging in critical human mtDNA control sites for replication. Proc. Natl. Acad. Sci. 2001;98:4022-4027 https://doi.org/10.1073/pnas.061013598
  11. Wanagat J, Cao Z, Pathare P, Aiken JM. Mitochondrial DNA deletion mutations colocalize with segmental electron transport system abnormalities, muscle fiber atrophy, fibersplitting, and oxidative damage in sarcopenia. FASEB J. 2001;15:322-332 https://doi.org/10.1096/fj.00-0320com
  12. Khaidakov M, Heflich RH, Manjanatha MG, Myers MB, Aidoo A. Accumulation of point mutations in mitochondrial DNA of aging mice. Mutat. Res. 2003;526:1-7 https://doi.org/10.1016/S0027-5107(03)00010-1
  13. Cortopassi GA, Arnheim N. Detection of a specific mitochondrial DNA deletion in tissues of older humans. Nucleic Acids Res. 1990;18:6927-6933 https://doi.org/10.1093/nar/18.23.6927
  14. Ikebe S, Tanaka M, Ohno K. Increase of deleted mitochondrial DNA in the striatum in Parkinson's disease and senescence. Biochem Biophys Res Commun. 1990;170:1044-1048 https://doi.org/10.1016/0006-291X(90)90497-B
  15. Cortopassi GA, Shibata D, Soong NW, Arnheim N. A pattern of accumulation of a somatic deletion of mitochondrial DNA in aging human tissues. Proc Natl Acad Sci. 1992;89:7370-7374 https://doi.org/10.1073/pnas.89.16.7370
  16. Wallace DC. Mitochondrial genetics: a paradigm for aging and degenerative diseases? Science. 1992;256:628-632 https://doi.org/10.1126/science.1533953
  17. Lu CY, Lee HC, Fahn HJ, Wei YH. Oxidative damage elicited by imbalance of free radical scavenging enzymes is associated with largescale mtDNA deletions in aging human skin. Mutat Res. 1999;423:11-21 https://doi.org/10.1016/S0027-5107(98)00220-6
  18. Shin MH, Rhie GE, Kim YK. ${H}_2{0}_2$ accumulation by catalase reduction changes MAP kinase signaling in aged human skin in vivo. J Invest Dermatol. 2005;125:221-229 https://doi.org/10.1111/j.0022-202X.2005.23823.x
  19. Nagley P, Wei YH. Ageing and mammalian mitochondrial genetics. Trends Genet. 1998;14:513-517 https://doi.org/10.1016/S0168-9525(98)01580-7
  20. Shoffner JM, Lott MT, Voljavec AS, Soueidan SA, Costigan DA, Wallace DC. Spontaneous Kearns -Sayre/chronic external ophthalmoplegia plus syndrome associated with a mitochondrial DNA deletion: a slip-replication model and metabolic therapy. Proc Natl Acad Sci. 1989;86:7952-7956 https://doi.org/10.1073/pnas.86.20.7952
  21. Brierley EJ, Johnson MA, Lightowlers RN, James OF, Turnbull DM. Role of mitochondrial DNA mutations in human aging: implications for the central nervous system and muscle. Ann Neurol. 1998;43:217-223 https://doi.org/10.1002/ana.410430212
  22. Sciacco M, Bonilla E, Schon EA, DiMauro S, Moraes CT. Distribution of wild-type and common deletion forms of mtDNA in normal and respiration-deficient muscle fibers from patients with mitochondrial myopathy. Hum Mol Genet. 1994;3:13-19 https://doi.org/10.1093/hmg/3.1.13
  23. D. Harman. The biologic clock: the mitochondria? J. Am. Geriatr. Soc. 1974;20:145-147 https://doi.org/10.1111/j.1532-5415.1972.tb00787.x
  24. Linnane AW, Marzuki S, Ozawa T, Tanaka M. Mitochondrial DNA mutations as an important contributor to ageing and degenerative diseases. Lancet 1. 1989;642-645 https://doi.org/10.1016/S0140-6736(89)92145-4
  25. Sener G, Jahovic N, Tosun OB, Atasoy M and Yeen BC. Melatinin ameliorates ionizing radiation-induced oxidative organ damage in rat. Life Sci. 2003;74:563-572 https://doi.org/10.1016/j.lfs.2003.05.011
  26. Dumont P, Burton M, Chen QM, Gonos ES, Frippiat C, Mazarati JB, Eliaers F, Remacle J and Toussaint O. Induction of replicative senescence biomarkers by sublethal oxidative stresses in normal human fibroblast. Free Radic. Biol. Med. 2000;28:361-373 https://doi.org/10.1016/S0891-5849(99)00249-X
  27. Berneburg M, Grether-Beck S, Kurten V, Ruzicka T, Briviba K, Sies H and Krutmann J. Singlet oxygen mediates the UVA-induced generation of the photoagingassociated mitochondrial common deletion. J. Biol. Chem. 1999;274:15345-15349 https://doi.org/10.1074/jbc.274.22.15345
  28. Lu W, Yoshikazu K, Li L, Taisuke B, Ryong-woon S, Yasuhito O, Koji O and Manabu F. Analysis of common Deletion (CD) and a novel deletion of mitochondrial DNA induced by ionizing radiation. Int. J. Radiat. Biol. 2007;83:433-442 https://doi.org/10.1080/09553000701370878