Fertility and Sterility
Volume 80, Issue 6 , Pages 1420-1430 , December 2003

Effects of age on DNA double-strand breaks and apoptosis in human sperm

  • Narendra P Singh, M.B.B.S., M.S.

      Affiliations

    • Corresponding Author InformationReprint requests: Narendra P. Singh, M.B.B.S., M.S., Department of Bioengineering, Box 357962, University of Washington, Seattle, Washington 98195-7962, USA (FAX: 206-685-2060).
    • Bioengineering, University of Washington, Seattle, Washington, USA
  • ,
  • Charles H Muller, Ph.D.

      Affiliations

    • Department of Urology, University of Washington, Seattle, Washington, USA
  • ,
  • Richard E Berger, M.D.

      Affiliations

    • Department of Urology, University of Washington, Seattle, Washington, USA

Received 6 February 2003 ,Revised 30 April 2003 ,Accepted 30 April 2003.

References 

  1. Evans HJ. Mutation and mutagenesis in inherited and acquired human disease. Mutat Res. 1996;351:89–103
  2. Aitken RJ, Gordon E, Harkiss D, Twigg JP, Milne P, Jennings Z, et al.  Relative impact of oxidative stress on the functional competence and genomic integrity of human spermatozoa. Biol Reprod. 1998;59:1037–1046
  3. Wyrobek AJ, Meistrich ML, Furer R, Bruce WR. Physical characteristics of mouse sperm nuclei. Biophys J. 1976;16:811–825
  4. Ward WS, Coffey DS. DNA packaging and organization in mammalian spermatozoa (comparison with somatic cells). Biol Reprod. 1991;44:569–574
  5. Ward WS. The structure of the sleeping genome (implication of sperm DNA organization for somatic cells). J Cell Biochem. 1994;55:77–82
  6. Ward WS, Zalensky AO. The unique complex organization of the transcriptionally silent sperm chromatin. Crit Rev Eukaryot Gene Expr. 1996;6:139–147
  7. Tanphaichitr N, Sobborn P, Taluppeth N, Chalermisarachai P. Basic nuclear proteins in testicular cells and ejaculated spermatozoa in man. Exp Cell Res. 1978;117:247–256
  8. Balhorn R. A model for the structure of chromatin in mammalian sperm. J Cell Biol. 1982;93:298–305
  9. Shen HM, Chia SE, Ong CN. Evaluation of oxidative DNA damage in human sperm and its association with male infertility. J Androl. 1999;20:718–723
  10. Sun JG, Jurisicova A, Casper RF. Detection of deoxyribonucleic acid fragmentation in human sperm (correlation with fertilization in vitro). Biol Reprod. 1997;56:602–607
  11. Morris ID, Ilott S, Dixon L, Brison DR. The spectrum of DNA damage in human sperm assessed by single cell gel electrophoresis (Comet assay) and its relationship to fertilization and embryo development. Hum Reprod. 2002;17:990–998
  12. Gray LE, Ostby J, Linder R, Goldman J, Rehnberg G, Cooper R. Carbendazim-induced alterations of reproductive development and function in the rat and hamster. Fundam Appl Toxicol. 1990;15:281–297
  13. Bonduelle M, Liebaers I, Deketelaere V, Derde MP, Carnus M, Devroey P, et al.  Neonatal data on a cohort of 2889 infants born after ICSI (1991–1999) and of 2995 infants born after IVF (1983–1999). Hum Reprod. 2002;17:671–694
  14. Maher ER, Brueton LA, Bowdin SC, Luharia A, Cooper W, Cole TR, et al.  Beckwith-Wiedemann syndrome and assisted reproduction technology (ART). J Med Genet. 2003;40:62–64
  15. De Jonge C. The clinical value of sperm nuclear DNA assessment. Hum Fertil (Camb). 2002;5:51–53
  16. Ford WCL, North K, Taylor H, Farrow A, Hull MGR, Golding J, et al.  Increasing paternal age is associated with delayed conception in a large population of fertile couples (evidence for declining fecundity in older men). Hum Reprod. 2000;15:1703–1708
  17. de La Rochebrochard E, Thonneau P. Paternal age and maternal age are risk factors for miscarriage; results of a multicentre European study. Hum Reprod. 2002;17:1649–1656
  18. Rosenbusch B, Strehler E, Sterzik K. Cytogenetics of human spermatozoa (correlations with sperm morphology and age of fertile men). Fertil Steril. 1992;58:1071–1072
  19. Joshi DS, Yick J, Murray D, Meistrich ML. Stage-dependent variation in the radiosensitivity of DNA in developing male germ cells. Radiat Res. 1990;121:274–281
  20. Sega GA, Generoso EE. Measurement of DNA breakage in specific germ-cell stages of male mice exposed to acrylamide, using an alkaline-elution procedure. Mutat Res. 1990;242:79–87
  21. Qiu J, Hales BF, Robaire B. Damage to rat spermatozoal DNA after chronic cyclophosphamide exposure. Biol Reprod. 1995;53:1465–1473
  22. Sakkas D, Moffatt O, Manicardi GC, Mariethoz E, Tarozzi N, Bizzaro D. Nature of DNA damage in ejaculated human spermatozoa and the possible involvement of apoptosis. Biol Reprod. 2002;66:1061–1067
  23. Singh NP, Danner DB, Tice RR, McCoy MT, Collins GD, Schneider EL. Abundant alkali-sensitive sites in DNA of human and mouse sperm. Exp Cell Res. 1989;184:461–470
  24. Singh NP, Stephens RE. X-ray induced DNA double-strand breaks in human sperm. Mutagenesis. 1998;13:75–79
  25. Donnelly ET, McClure N, Lewis SE. The effect of ascorbate and alpha-tocopherol supplementation in vitro on DNA integrity and hydrogen peroxide-induced DNA damage in human spermatozoa. Mutagenesis. 1999;14:505–512
  26. Hughes CM, McKelvey-Martin VJ, Lewis SE. Human sperm DNA integrity assessed by the comet and ELISA assays. Mutagenesis. 1999;14:71–75
  27. Irvine DS, Twigg JP, Gordon EL, Fulton N, Milne PA, Aitken RJ. DNA integrity in human spermatozoa (relationships with semen quality). J Androl. 2000;21:33–44
  28. Donnelly ET, McClure N, Lewis SE. Cryopreservation of human semen and prepared sperm (effects on motility parameters and DNA integrity). Fertil Steril. 2001;76:892–900
  29. Chen H, Cheung MP, Chow PH, Cheung AL, Liu W, O WS. Protection of sperm DNA against oxidative stress in vivo by accessory sex gland secretions in male hamsters. Reproduction. 2002;124:491–499
  30. Haines GA, Hendry JH, Daniel CP, Morris ID. Germ cell and dose-dependent DNA damage measured by the comet assay in murine spermatozoa after testicular X-irradiation. Biol Reprod. 2002;67:854–861
  31. O'Connell M, McClure N, Lewis SE. A comparison of mitochondrial and nuclear DNA status in testicular sperm from fertile men and those with obstructive azoospermia. Hum Reprod. 2002;17:1571–1577
  32. Sierens J, Hartley JA, Campbell MJ, Leathem AJ, Woodside JV. In vitro isoflavone supplementation reduces hydrogen peroxide-induced DNA damage in sperm. Teratog Carcinog Mutagen. 2002;22:227–234
  33. Marty MS, Singh NP, Holsapple MP, Gollapudi BB. Influence of p53 zygosity on select sperm parameters of the mouse. Mutat Res. 1999;427:39–45
  34. Larson KL, Brannian JD, Singh NP, Burbach JA, Jost LK, Hansen KP, et al.  Chromatin structure in globozoospermia (a case study). J Androl. 2001;22:424–431
  35. Duty SM, Singh NP, Ryan L, Chen Z, Lewis C, Huang T, et al.  Reliability of the comet assay in cryopreserved human sperm. Hum Reprod. 2002;17:1274–1280
  36. Roosen-Runge EC. Germinal-cell loss in normal metazoan spermatogenesis. Reprod Fertil. 1973;35:339–348
  37. Allan DJ, Harmon BV, Kerr JFR. Cell death in spermatogenesis. In:  Porter CS editors. Perspective on mammalian cell death. Oxford, United Kingdom: Oxford University Press; 1987;p. 229–258
  38. Lee J, Richburg JH, Younkin SC, Boekelheide K. The Fas system is a key regulator of germ cell apoptosis in the testis. Endocrinology. 1997;138:2081–2088
  39. Sinha Hikim AP, Swerdloff RS. Hormonal and genetic control of germ cell apoptosis in the testis. Rev Reprod. 1999;4:38–47
  40. Print CG, Loveland KL. Germ cell suicide (new insights into apoptosis during spermatogenesis). Bioessays. 2000;22:423–430
  41. Martincic DS, Klun IV, Zorn B, Vrtovec HM. Germ cell apoptosis in the human testis. Pflugers Arch. 2001;442:R159–160
  42. Anzar M, He L, Buhr MM, Kroetsch TG, Pauls KP. Sperm apoptosis in fresh and cryopreserved bull semen detected by flow cytometry and its relationship with fertility. Biol Reprod. 2002;66:354–360
  43. Baccetti B, Collodel G, Piomboni P. Apoptosis in human ejaculated sperm cells (notulae seminologicae 9). J Submicrosc Cytol Pathol. 1996;28:587–596
  44. Weil M, Jacobson MD, Raff MC. Are caspases involved in the death of cells with a transcriptionally inactive nucleus? Sperm and chicken erythrocytes. J Cell Sci. 1998;111:2707–2715
  45. Gandini L, Lombardo F, Paoli D, Caponecchia L, Familiari G, Verlengia C, et al.  Study of apoptotic DNA fragmentation in human spermatozoa. Hum Reprod. 2000;15:830–839
  46. Muratori M, Piomboni P, Baldi E, Filimberti E, Pecchioli P, Moretti E, et al.  Functional and ultrastructural features of DNA-fragmented human sperm. J Androl. 2000;21:903–912
  47. Oosterhuis GJ, Mulder AB, Kalsbeek-Batenburg E, Lambalk CB, Schoemaker J, Vermes I. Measuring apoptosis in human spermatozoa (a biological assay for semen quality?). Fertil Steril. 2000;74:245–250
  48. Koji T. Male germ cell death in mouse testes (possible involvement of Fas and Fas ligand). Med Electron Microsc. 2001;34:213–222
  49. Heyer BS, MacAuley A, Behrendtsen O, Werb Z. Hypersensitivity to DNA damage leads to increased apoptosis during early mouse development. Genes Dev. 2000;14:2072–2084
  50. Hoyes KP, Lord BI, McCann C, Hendry JH, Morris ID. Transgenerational effects of preconception paternal contamination with (55)Fe. Radiat Res. 2001;156:488–494
  51. Ricci G, Perticarari S, Fragonas E, Giolo E, Canova S, Pozzobon C, et al.  Apoptosis in human sperm (its correlation with semen quality and the presence of leukocytes). Hum Reprod. 2002;17:2665–2672
  52. Weng SL, Taylor SL, Morshedi M, Schuffner A, Duran EH, Beebe S, et al.  Caspase activity and apoptotic markers in ejaculated human sperm. Mol Hum Reprod. 2002;8:984–991
  53. Shen HM, Dai J, Chia SE, Lim A, Ong CN. Detection of apoptotic alterations in sperm in subfertile patients and their correlations with sperm quality. Hum Reprod. 2002;17:1266–1273
  54. Singh NP. A simple method for accurate estimation of apoptotic cells. Exp Cell Res. 2000;256:328–337
  55. Bacso Z, Eliason JF. Measurement of DNA damage associated with apoptosis by laser scanning cytometry. Cytometry. 2001;45:180–186
  56. Nigro M, Frenzilli G, Scarcelli V, Gorbi S, Regoli F. Induction of DNA strand breakage and apoptosis in the eel Anguilla anguilla. Mar Environ Res 2002;54:517–20
  57. World Health Organization . WHO laboratory manual for the examination of human semen and semen-cervical mucus interactions, 4th ed. Cambridge, United Kingdom: Cambridge University Press; 1999;
  58. Coetzee K, Kruger TF, Lombard CJ, Shaughnessy D, Oehninger S, Özgür K, et al.  Assessment of interlaboratory and intralaboratory sperm morphology readings with the use of a Hamilton Thorne Research integrated visual optical system semen analyzer. Fertil Steril. 1999;71:80–84
  59. Singh NP. Microgels for estimation of DNA strand breaks, DNA protein crosslinks, and apoptosis. Mutat Res. 2000;455:119–135
  60. Singh NP. The DNA diffusion assay for apoptosis assessment. In: Rosalyn Blumenthal, ed. Methods in molecular medicine. Totowa, NJ: Humana Press: In press
  61. Evenson DP, Jost LK. Sperm chromatin structure assay (DNA denaturability). In:  Darzynkiewicz Z,  Robinson JP,  Crissman HA editor. Methods in cell biology. Vol. 42. Flow cytometry 2E. Orlando, FL: Academic Press; 1994;p. 159–176
  62. Fernandez JL, Muriel L, Rivero MT, Goyanes V, Vazquez R, Alvares JG. The sperm chromatin dispersion test (a simple method for the determination of sperm DNA fragmentation). J Androl. 2003;24:59–66
  63. Gorczyca W, Traganos F, Jesionowska H, Darzynkiewicz Z. Presence of DNA strand breaks and increased sensitivity of DNA in situ to denaturation in abnormal human sperm cells (analogy to apoptosis of somatic cells). Exp Cell Res. 1993;207:202–205
  64. Sailer BL, Jost LK, Evenson DP. Mammalian sperm DNA susceptibility to in situ denaturation associated with the presence of DNA strand breaks as measured by the terminal deoxynucleotidyl transferase assay. J Androl. 1995;16:80–87
  65. Sega GA, Sluder AE, McCoy LS, Owens JG, Generoso EE. The use of alkaline elution procedures to measure DNA damage in spermiogenic stages of mice exposed to methyl methanesulfonate. Mutat Res. 1986;159:55–63
  66. Sega GA, Generoso EE. Measurement of DNA breakage in spermiogenic germ-cell stages of mice exposed to ethylene oxide, using an alkaline elution procedure. Mutat Res. 1988;197:93–99
  67. Singh NP, McCoy MT, Tice RR, Schneider EL. A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res. 1988;175:184–191
  68. Fraga CG, Motchnik PA, Wyrobek AJ, Rempel DM, Ames BN. Smoking and low antioxidant levels increase oxidative damage to sperm DNA. Mutat Res. 1996;351:199–203
  69. Barnes CJ, Covington BW, Cameron IL, Lee M. Effect of aging on spontaneous and induced mouse testicular germ cell apoptosis. Aging (Milano). 1998;10:497–501
  70. Barnes CJ, Covington BW, Lee M. Effect of aging and dietary restriction on rat testicular germ cell apoptosis. J Gerontol A Biol Sci Med Sci. 1999;54:B199–204
  71. Brinkworth MH. Paternal transmission of genetic damage (findings in animals and humans). Int J Androl. 2000;23:123–135
  72. Brinkworth MH, Nieschlag E. Association of cyclophosphamide-induced male-mediated, foetal abnormalities with reduced paternal germ-cell apoptosis. Mutat Res. 2000;447:149–154
  73. Yin Y, Stahl BC, DeWolf WC, Morgentaler A. p53-mediated germ cell quality control in spermatogenesis. Dev Biol. 1998;204:165–171
  74. 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;13:1429–1436
  75. Shen H, Ong C. Detection of oxidative DNA damage in human sperm and its association with sperm function and male infertility. Free Radic Biol Med. 2000;28:529–536
  76. Ramos L, Wetzels AM. Low rates of DNA fragmentation in selected motile human spermatozoa assessed by the TUNEL assay. Hum Reprod. 2001;16:1703–1707
  77. Payne CM, Bernstein C, Bernstein H. Apoptosis overview emphasizing the role of oxidative stress, DNA damage and signal-transduction pathways. Leuk Lymphoma. 1995;19:43–93
  78. Li GM. The role of mismatch repair in DNA damage-induced apoptosis. Oncol Res. 1999;11:393–400
  79. Maione B, Pittoggi C, Achene L, Lorenzini R, Spadafora C. Activation of endogenous nucleases in mature sperm cells upon interaction with exogenous DNA. DNA Cell Biol. 1997;16:1087–1097
  80. McCarthy S, Ward WS. Functional aspects of mammalian sperm chromatin. Hum Fertil (Camb). 1999;2:56–60
  81. Ricote M, Alfaro JM, Garcia-Tunon I, Arenas MI, Fraile B, Paniagua R, et al.  Control of the annual testicular cycle of the marbled-newt by p53, p21, and Rb gene products. Mol Reprod Dev. 2002;63:202–209
  82. Blanco-Rodriguez J. DNA replication and germ cell apoptosis during spermatogenesis in the cat. J Androl. 2002;23:484–490
  83. Blanco-Rodriguez J, Martinez-Garcia C. Apoptosis pattern elicited by several apoptogenic agents on the seminiferous epithelium of the adult rat testis. J Androl. 1998;19:487–497
  84. Blanco-Rodriguez J. Deoxyribonucleic acid replication and germ cell apoptosis during spermatogenesis in the rabbit. J Androl. 2002;23:182–187
  85. Brash DE. Cellular proofreading. Nat Med. 1996;2:525–526
  86. Woolveridge I, Morris ID. Apoptosis in male reproductive toxicology. In:  Roberts R editors. Apoptosis in toxicology. New York: Taylor and Francis; 2000;p. 72–87
  87. Sakkas D, Mariethoz E, St. John JC. Abnormal sperm parameters in humans are indicative of an abortive apoptotic mechanism linked to the Fas-mediated pathway. Exp Cell Res. 1999;251:350–355
  88. Kemal Duru N, Morshedi M, Oehninger S. Effects of hydrogen peroxide on DNA and plasma membrane integrity of human spermatozoa. Fertil Steril. 2000;74:1200–1207
  89. Barroso G, Morshedi M, Oehninger S. Analysis of DNA fragmentation, plasma membrane translocation of phosphatidylserine and oxidative stress in human spermatozoa. Hum Reprod. 2000;15:1338–1344
  90. Sega GA, Sotomayor RE, Owens JG. A study of unscheduled DNA synthesis induced by X-rays in the germ cells of male mice. Mutat Res. 1978;49:239–257
  91. Ono T, Okada S. Radiation-induced DNA single-strand scission and its rejoining in spermatogonia and spermatozoa of mouse. Mutat Res. 1977;43:25–36
  92. Tateno H, Iijima S, Nakanishi Y, Kamiguchi Y, Asaka A. No induction of chromosome aberrations in human spermatozoa exposed to extremely low frequency electromagnetic fields. Mutat Res. 1998;414:31–35
  93. Singh NP, Danner DB, Tice RR, Brant L, Schneider EL. DNA damage and repair with age in individual human lymphocytes. Mutat Res. 1990;237:123–130
  94. Singh NP, Danner DB, Tice RR, Pearson JD, Brant LJ, Morrell CH, et al.  Basal DNA damage in human lymphocytes with age. Mutat Res. 1991;256:1–6
  95. Pollard I. Substance abuse and parenthood (biological mechanisms—bioethical challenges). Women Health. 2000;30:1–24
  96. Shu XO, Reaman GH, Lampkin B, Sather HN, Pendergrass TW, Robison LL. Association of paternal diagnostic X-ray exposure with risk of infant leukemia. Investigators of the Childrens Cancer Group. Cancer Epidemiol Biomarkers Prev. 1994;3:645–653
  97. Ji BT, Shu XO, Linet MS, Zheng W, Wacholder S, Gao YT, et al.  Paternal cigarette smoking and the risk of childhood cancer among offspring of nonsmoking mothers. J Natl Cancer Inst. 1997;89:238–244
  98. Hu J, Many Y, Ugnat AM. Parental cigarette smoking, hard liquor consumption and the risk of childhood brain tumors—a case-control study in northeast China. Acta Oncol. 2000;39:979–984
  99. Malaspina D, Corcoran C, Fahim C, Berman A, Harkavy-Friedman J, Yale S, et al.  Paternal age and sporadic schizophrenia (evidence for de novo mutations). Am J Med Genet. 2002;114:299–303
  100. Roeleveld N, Vingerhoets E, Zielhuis GA, Galbreels F. Mental retardation associated with parental smoking and alcohol consumption before, during and after pregnancy. Prev Med. 1992;21:110–119
  101. Klonoff-Cohen H, Bleha J, Lam-Kruglick P. A prospective study of the effects of female and male caffeine consumption on the reproductive endpoints of IVF and gamete intra-Fallopian transfer. Hum Reprod. 2002;17:1746–1754
  102. Cai L, Hales BF, Robaire B. Induction of apoptosis in the germ cells of adult male rats after exposure to cyclophosphamide. Biol Reprod. 1997;56:1490–1497
  103. Takagi S, Itoh N, Kimura M, Sasao T, Tsukamoto T. Spermatogonial proliferation and apoptosis in hypospermatogenesis associated with nonobstructive azoospermia. Fertil Steril. 2001;76:901–907
  104. Zini A, Bielecki R, Phang D, Zenzes MT. Correlations between two markers of sperm DNA integrity, DNA denaturation and DNA fragmentation, in fertile and infertile men. Fertil Steril. 2001;75:674–677

 Supported by the Paul G. Allen Foundation for Medical Research.Authors Singh, Muller, and Berger contributed equally to this work.

PII: S0015-0282(03)02249-0

doi: 10.1016/j.fertnstert.2003.04.002

Fertility and Sterility
Volume 80, Issue 6 , Pages 1420-1430 , December 2003