Clinical and Experimental Reproductive Medicine

The Korean Society for Reproductive Medicine (대한생식의학회)
권호 표지

Effects of Reactive Oxygen Species on Sperm Function, Lipid Peroxidation and DNA Fragmentation in Bovine Spermatozoa

소 정자에 있어서 활성산소계가 정자 기능과 지방산화 및 DNA 절편화에 미치는 영향

  • Ryu, Buom-Yong (Department of Animal Science & Technology Chung-Ang University) ;
  • Chung, Yung-Chai (Department of Animal Science & Technology Chung-Ang University) ;
  • Kim, Chang-Keun (Department of Animal Science & Technology Chung-Ang University) ;
  • Shin, Hyun-A (Department of Animal Science & Technology Chung-Ang University) ;
  • Han, Jung-Ho (Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University) ;
  • Pang, Myung-Geol (Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University) ;
  • Oh, Sun-Kyung (Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University) ;
  • Kim, Seok-Hyun (Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University) ;
  • Moon, Shin-Yong (Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University)
  • 류범용 (중앙대학교 동물자원과학과) ;
  • 정영채 (중앙대학교 동물자원과학과) ;
  • 김창근 (중앙대학교 동물자원과학과) ;
  • 신현아 (중앙대학교 동물자원과학과) ;
  • 한정호 (서울대학교 의학연구원 인구의학연구소) ;
  • 방명걸 (서울대학교 의학연구원 인구의학연구소) ;
  • 오선경 (서울대학교 의학연구원 인구의학연구소) ;
  • 김석현 (서울대학교 의학연구원 인구의학연구소) ;
  • 문신용 (서울대학교 의학연구원 인구의학연구소)
  • Published : 2002.06.30
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Abstract

Objective : To evaluate the effects of the reactive oxygen species (ROS) generated with a xanthine (X) and xanthine oxidase (XO) system on sperm function, the change of sperm characteristics, lipid peroxidation, and DNA fragmentation in bovine spermatozoa. Materials and Methods: ROS were produced using a combination of 1000 uM X and 50 mU/ml XO. The ROS scavengers: superoxide dismu tase (SOD) (200 U/ml) and catalase (500 U/ml) were also tested. Spermatozoa were incubated for 2 hours in BWW medium with a combination of X-XO supplemented with or without ROS scavengers at $37^{circ}C$ under 5% $CO_2$ incubator. Sperm movement characteristics by CASA (computer-aided sperm analysis), HOST (hypoosmotic swelling test), Caionophore induced acrosome reaction, malondialdehyde formation for the analysis of lipid peroxidation, the percentage of DNA fragmentation using the method of TdT-mediated nick end labelling (TUNEL) by flow cytometry were determined after 2 hours incubation. Results: The action of ROS on bovine spermatozoa resulted in a decreased in capacity for sperm motility, Ca-ionophore induced acrosome reaction and membrane integrity, an increased in malondialdehyde formation and the percentage of sperm with DNA fragmentation. In the effects of antioxidant, catalase completely alleviated the toxic effects induced by the ROS in terms of sperm function and characteristics, however SOD exhibited no capacity to reduce the toxic effects. Conclusion: The ROS can induce significant damages to sperm functions and characteristics. The useful ROS scavengers can minimized the defects of sperm function and various damages of spermatozoa.

Keywords

References

  1. Jones R, Mann T, Sherins RJ. Peroxidative breakdown of phospholipids in human spermatozoa: spermicidal effects of fatty acid peroxides and protective action of seminal plasma. Fertil Steril 1979; 31: 531-7 https://doi.org/10.1016/S0015-0282(16)43999-3
  2. Bennet PJ, Moatti JP, Mansat A, Ribbes H, Cayrac JC, Pontonnier F, et al. Evidence for the activation of phospholipase during acrosome reaction of human sperm elicited by calcium ionophore A23187. Biochim Biophys Acta 1987;919:255-65 https://doi.org/10.1016/0005-2760(87)90265-7
  3. Goldman R, Ferber E, Zort U. Reactive oxygen species are involved in the activation of cellular phospholipase A2. FEBS Lett 1992; 309: 190-2 https://doi.org/10.1016/0014-5793(92)81092-Z
  4. De Lamirande E, Gagnon C. Impact of reactive oxygen species on spermatozoa; a balancing act between beneficial and detrimental effects. Hum Reprod 1995; 12 Suppl 1: 15-21
  5. Aitken RJ, Clarkson JS, Hargreave TB, Irvine DS, Wu FC. Analysis of the relationship between defective sperm function and the generation of reactive oxygen species in cases of oligozoospermia. J Androl 1989; 10:214-20 https://doi.org/10.1002/j.1939-4640.1989.tb00091.x
  6. De Lamirande E, Gagnon C. Reactive oxygen species and human spermatozoa I. Effects on the motility of intact spermatozoa and on sperm axoneme; and II. Depletion of adenosine triphosphate plays an important role in the inhibition of sperm motility. J Androl 1992; 13:368-86
  7. Comporti M. Three models of free radical induced cell injury. Chem Biol Interactions 1989; 72: 1-56 https://doi.org/10.1016/0009-2797(89)90016-1
  8. Hinshaw DB, Sklar LA, Schraufstatter IU, Hyslop PA, Rossi MW, Spragg RG, et al. Cytoskeletal and morphologic impact of cellular oxidant injury. Am J Pathol 1986; 123: 454-64
  9. Aitken RJ, Buckingham DW, Harkiss D. Use of a xanthine oxidase free radical generating system to investigate the cytotoxic effects of reactive oxygen species on human spermatozoa. J Reprod Fetil 1993; 97:441-50 https://doi.org/10.1530/jrf.0.0970441
  10. Biggers JS, Whitten WK, Whittingham DG. The culture of mouse embryos in vitro. In: Daniel JC, editor. Method in mammalian embryology. San Francisco: Freeman; 1971. p.86-116
  11. Mortimer ST, Mortimer D. Kinematics of human spermatozoa incubated under capacitating conditions. J Androl 1990; 11: 195-203
  12. Jeyendran RS, Van der Yen HR, Perez-Pelaez M, Crabo BG, Zaneveld LJ. Development of an assay to assess the functional integrity of the human sperm membrane and its relationship to other semen characteristics. J Reprod Fertil 1984; 70: 219-28 https://doi.org/10.1530/jrf.0.0700219
  13. Cummins JM, Pember SM, Jequier AM, Yovich JL. A test of the human sperm acrosome reaction following ionophore challenge. J Androl 1991; 12:98-103
  14. Aitken RJ, Harkiss D, Buckingham DW. Analysis of lipid peroxidation mechanism in human spermatozoa. Mol Reprod Dev 1993; 35: 302-15 https://doi.org/10.1002/mrd.1080350313
  15. Slater TF, Sawyer BC. The stimulatory effects of carbon tetrachloride and other halogenoalkanes on peroxidative in rat liver fractions in vitro. Biochem J 1971; 123: 805-14 https://doi.org/10.1042/bj1230805
  16. Aitken RJ, Buckingham DW, Katrine MW. Reactive oxygen species and human spermatozoa: Analysis of the cellular mechanism involved in luminol- and lucigenin-dependent chemiluminescence. J Cell Physiol 1992; 151: 466-77 https://doi.org/10.1002/jcp.1041510305
  17. Alvarez JG, Touchstone JC, Blasco L, Storey BT. Spontaneous lipid peroxidation and production of hydrogen peroxide in human spermatozoa: superoxide dismutase as major enzyme protectant against oxygen toxicity. J Androl 1987; 8: 338-48 https://doi.org/10.1002/j.1939-4640.1987.tb00973.x
  18. Aitken RJ, Irvine DS, WU FC. Prospective analysis of sperm-oocyte fusion and reactive oxygen species generation as criteria for the diagnosis of infertility. A J Ob & Gyn 1991; 164: 542-51 https://doi.org/10.1016/S0002-9378(11)80017-7
  19. Iwasaki A, Gagnon e. Formation of reactive oxygen species in spermatozoa of infertility patients. Fertil Steril 1992; 57: 2409-16
  20. Alvarez JG, Storey BT. Evidence for increased peroxidative damage and loss of super oxide dis-mutase activity as a model of sublethal cryodamage to human sperm during cryopreservation. J Androl 1992; 13: 232-41
  21. hyashiki T, Tsuka T, Mohri T. Increase of the molecular rigidity of the protein conformation in the intestinal brush-border membranes by lipid peroxidation. Biochim Biophys Acta 1988; 939: 383-92 https://doi.org/10.1016/0005-2736(88)90084-3
  22. Block ER. Hydrogen peroxide alters the physical state and function of the plasma membrane of pulmonary artery endothelial cells. J Cell Physiol 1991; 20: 362-9
  23. Twigg J, Fulton N, Gomez E, Irvine DS, Aitken RJ. Analysis of the impact of intracellular reactive oxygen species generation on the structural and functional integrity of human spermatozoa: lipid peroxidation, DNA fragmentation and effectiveness of antioxidants. Hum Reprod 1998; 13: 1429-36 https://doi.org/10.1093/humrep/13.6.1429
  24. Terada LS, Leff JA, Guidot OM, Willingham IR, Repine JE. Inactivation of xanthine oxidase by hydrogen peroxide involves sites-directed hydroxyl radical formation. Free Radicals in Biol Med 1991; 10: 61-8 https://doi.org/10.1016/0891-5849(91)90022-U
  25. Griveau JF, Dumont E, Renard P, Callegari JP, Le Lannou D. Reactive oxygen species, lipid peroxidation and enzymatic defence systems in human spermatozoa. J Reprod Fertil 1995; 103: 17-26 https://doi.org/10.1530/jrf.0.1030017
  26. Blondin P, Coenen K, Sirard MA. The impact of reactive oxygen species on bovine sperm fertilizing ability and oocyte maturation. J Androl 1997; 18: 454-60
  27. Lopes S, Jurisicova A, Sun JG, Casper RF. Reactive oxygen species: potential cause for DNA fragmentation in human spermatozoa. Hum Reprod 1998; 13: 896-900 https://doi.org/10.1093/humrep/13.4.896