Abstract
Sperm DNA fragmentation (SDF) refers to breaks or damage in the DNA strand(s) in sperm cells, which can lead to an increased risk of male infertility. The extent of DNA fragmentation can vary widely, from mild to severe, and can be caused by a number of factors, including oxidative stress (OS), environmental toxins, and genetic abnormalities. SDF testing can help identify potential fertility issues even in men who have normal semen parameters or unexplained infertility. Moreover, SDF testing can also help predict the success of certain fertility treatments. For example, men with high levels of DNA fragmentation may be less likely to have success with in vitro fertilization (IVF) but may benefit from intracytoplasmic sperm injection (ICSI). SDF analysis is a relatively new field of study, and there are several different methods used to measure sperm DNA fragmentation. Each test has its advantages and limitations, and the results may vary depending on the specific circumstances of the individual being tested. At a time of constantly emerging ideas, policies, and protocols for SDF testing in male infertility, it is important to have an updated understanding of the contemporary SDF tests to ensure that research, diagnoses, and treatment are based on the latest concepts in this field. The present chapter describes updated concepts on SDF and the significance of SDF testing to better identify the underlying mechanisms of male infertility, develop tailored male infertility treatment, predict treatment success, and reduce the risk of unsuccessful pregnancy and ART outcomes.
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References
Abdelbaki SA, Sabry JH, Al-Adl AM, Sabry HH (2017) The impact of coexisting sperm DNA fragmentation and seminal oxidative stress on the outcome of varicocelectomy in infertile patients: a prospective controlled study. Arab J Urol 15(2):131–139
Agarwal A, Varghese AC, Sharma RK (2009) Markers of oxidative stress and sperm chromatin integrity. In: Park-Sarge OK, Curry TE (eds) Molecular endocrinology: methods and protocols. Humana Press, Totowa, NJ, pp 377–402
Agarwal A, Sharma R, Ahmad G (2017) Sperm chromatin assessment textbook of assisted reproductive techniques, 5th edn. CRC Press, Boca Raton, FL, pp 65–87
Agarwal A, Cho CL, Majzoub A, Esteves SC (2017a) Frontiers in clinical andrology. Transl Androl Urol 6(Suppl 4):S343–S345
Agarwal A, Cho CL, Majzoub A, Esteves SC (2017b) The Society for Translational Medicine: clinical practice guidelines for sperm DNA fragmentation testing in male infertility. Transl Androl Urol 6:S720–S733
Agarwal A, Panner Selvam MK, Baskaran S, Cho CL (2019) Sperm DNA damage and its impact on male reproductive health: a critical review for clinicians, reproductive professionals and researchers. Expert Rev Mol Diagn 19(6):443–457
Baker H, Liu D (1996) Assessment of nuclear maturity. CRC Press, London
Balasuriya A, Speyer B, Serhal P, Doshi A, Harper JC (2011) Sperm chromatin dispersion test in the assessment of DNA fragmentation and aneuploidy in human spermatozoa. Reprod Biomed Online 22(5):428–436
Benchaib M, Braun V, Lornage J, Hadj S, Salle B, Lejeune H, Guérin JF (2003) Sperm DNA fragmentation decreases the pregnancy rate in an assisted reproductive technique. Hum Reprod 18(5):1023–1028
Bibi R, Jahan S, Razak S, Hammadeh ME, Almajwal A, Amor H (2022) Protamines and DNA integrity as a biomarkers of sperm quality and assisted conception outcome. Andrologia 54(6):e14418
Bungum M, Humaidan P, Axmon A, Spano M, Bungum L, Erenpreiss J et al (2007) Sperm DNA integrity assessment in prediction of assisted reproduction technology outcome. Hum Reprod 22(1):174–179
Bungum M, Spano M, Humaidan P, Eleuteri P, Rescia M, Giwercman A (2008) Sperm chromatin structure assay parameters measured after density gradient centrifugation are not predictive for the outcome of ART. Hum Reprod 23(1):4–10
Caliskan Z, Kucukgergin C, Aktan G, Kadioglu A, Ozdemirler G (2022) Evaluation of sperm DNA fragmentation in male infertility. Andrologia 54:e14587
Carrell DT, Liu L, Peterson C, Jones K, Hatasaka H, Erickson L et al (2003) Sperm DNA fragmentation is increased in couples with unexplained recurrent pregnancy loss. Arch Androl 49(1):49–55
Collins JA, Barnhart KT, Schlegel PN (2008) Do sperm DNA integrity tests predict pregnancy with in vitro fertilization? Fertil Steril 89(4):823–831
Cui ZL, Zheng DZ, Liu YH, Chen LY, Lin DH, Feng-Hua L (2015) Diagnostic accuracies of the TUNEL, SCD, and comet based sperm DNA fragmentation assays for male infertility: a meta-analysis study. Clin Lab 61(5–6):525–535
De Iuliis GN, Thomson LK, Mitchell LA, Finnie JM, Koppers AJ, Hedges A et al (2009) DNA damage in human spermatozoa is highly correlated with the efficiency of chromatin remodeling and the formation of 8-Hydroxy-2′-Deoxyguanosine, a marker of oxidative Stress1. Biol Reprod 81(3):517–524
Dutta S, Henkel R, Sengupta P, Agarwal A (2020) Physiological role of ROS in sperm function male infertility. Springer, Berlin, pp 337–345
Duty SM, Singh NP, Ryan L, Chen Z, Lewis C, Huang T et al (2002) Andrology. Reliability of the comet assay in cryopreserved human. Hum Reprod 17(5):1274–1280
Enciso M, Sarasa J, Agarwal A, Fernández JL, Gosálvez J (2009) A two-tailed comet assay for assessing DNA damage in spermatozoa. Reprod Biomed Online 18(5):609–616
Erenpreisa J, Erenpreiss J, Freivalds T, Slaidina M, Krampe R, Butikova J et al (2003) Toluidine blue test for sperm DNA integrity and elaboration of image cytometry algorithm. Cytomet Pt A 52A(1):19–27
Erenpreiss J, Jepson K, Giwercman A, Tsarev I, Erenpreisa J, Spano M (2004) Toluidine blue cytometry test for sperm DNA conformation: comparison with the flow cytometric sperm chromatin structure and TUNEL assays. Hum Reprod 19(10):2277–2282
Esterhuizen A, Franken D, Lourens J, Prinsloo E, Van Rooyen L (2000) Sperm chromatin packaging as an indicator of in-vitro fertilization rates. Hum Reprod 15(3):657–661
Esteves SC, Agarwal A, Cho CL, Majzoub A (2017a) A strengths-weaknesses-opportunities-threats (SWOT) analysis on the clinical utility of sperm DNA fragmentation testing in specific male infertility scenarios. Transl Androl Urol 6:S734–S760
Esteves SC, Agarwal A, Majzoub A (2017b) The complex nature of the sperm DNA damage process. Transl Androl Urol 6:S557–S559
Esteves SC, Zini A, Coward RM, Evenson DP, Gosálvez J, Lewis SE et al (2021) Sperm DNA fragmentation testing: summary evidence and clinical practice recommendations. Andrologia 53(2):e13874
Esteves SC, López-Fernández C, MartÃnez MG, Silva EA, Gosálvez J (2022) Reliability of the sperm chromatin dispersion assay to evaluate sperm deoxyribonucleic acid damage in men with infertility. Fertil Steril 117(1):64–73
Evenson DP (2022) Sperm chromatin structure assay (SCSA®) for fertility assessment. Curr Prot 2(8):e508
Evenson DP, Wixon R (2006) Clinical aspects of sperm DNA fragmentation detection and male infertility. Theriogenology 65(5):979–991
Evenson DP, Jost LK, Marshall D, Zinaman MJ, Clegg E, Purvis K et al (1999) Utility of the sperm chromatin structure assay as a diagnostic and prognostic tool in the human fertility clinic. Hum Reprod 14(4):1039–1049
Fernández JL, Muriel L, Rivero MT, Goyanes V, Vazquez R, Alvarez JG (2003) The sperm chromatin dispersion test: a simple method for the determination of sperm DNA fragmentation. J Androl 24(1):59–66
Fernández JL, Cajigal D, López-Fernández C, Gosálvez J (2011) Assessing sperm DNA fragmentation with the sperm chromatin dispersion test. In: Didenko VV (ed) DNA damage detection in situ, ex vivo, and in vivo: methods and protocols. Humana Press, Totowa, NJ, pp 291–301
Gorczyca W, Gong J, Darzynkiewicz Z (1993) Detection of DNA strand breaks in individual apoptotic cells by the in situ terminal deoxynucleotidyl transferase and nick translation assays. Cancer Res 53(8):1945–1951
Gupta S, Sharma R, Agarwal A (2017) Inter-and intra-laboratory standardization of TUNEL assay for assessment of sperm DNA fragmentation. Curr Protoc Toxicol 74(1):16.11.11–16.11.22
Hammadeh M, Zeginiadov T, Rosenbaum P, Georg T, Schmidt W, Strehler E (2001) Predictive value of sperm chromatin condensation (aniline blue staining) in the assessment of male fertility. Arch Androl 46(2):99–104
Hellman B, Vaghef H, Boström B (1995) The concepts of tail moment and tail inertia in the single cell gel electrophoresis assay. Mutat Res 336(2):123–131
Henkel RR, Franken DR (2011) Sperm DNA fragmentation: origin and impact on human reproduction. J Reprod Stem Cell Biotechnol 2(2):88–108
Henkel R, Hoogendijk CF, Bouic PJD, Kruger TF (2010) TUNEL assay and SCSA determine different aspects of sperm DNA damage. Andrologia 42(5):305–313
Hoshi K, Katayose H, Yanagida K, Kimura Y, Sato A (1996) The relationship between acridine orange fluorescence of sperm nuclei and the fertilizing ability of human sperm. Fertil Steril 66(4):634–639
Hosseinifar H, Yazdanikhah S, Modarresi T, Totonchi M, Sadighi Gilani M, Sabbaghian M (2015) Correlation between sperm DNA fragmentation index and CMA 3 positive spermatozoa in globozoospermic patients. Andrology 3(3):526–531
Irez T, Durmus T (2022) Sperm chromatin condensation level: depending on fixatives. Andrology 11:262
Ishikawa T, Fujioka H, Ishimura T, Takenaka A, Fujisawa M (2007) Increased testicular 8-hydroxy-2′-deoxyguanosine in patients with varicocele. BJU Int 100(4):863–866
Jarow J, Sigman M, Kolettis P (2011) The optimal evaluation of the infertile male: best practice statement reviewed and validity confirmed 2011. In: Paper presented at the American Urological Association
Kazerooni T, Asadi N, Jadid L, Kazerooni M, Ghanadi A, Ghaffarpasand F et al (2009) Evaluation of sperm’s chromatin quality with acridine orange test, chromomycin A3 and aniline blue staining in couples with unexplained recurrent abortion. J Assist Reprod Genet 26(11):591–596
Kodama H, Yamaguchi R, Fukuda J, Kasai H, Tanaka T (1997) Increased oxidative deoxyribonucleic acid damage in the spermatozoa of infertile male patients. Fertil Steril 68(3):519–524
Leuchtenberger C, Schrader F, Weir DR, Gentile DP (1953) The deoxyribosenucleic acid (DNA) content in spermatozoa of fertile and infertile human males. Chromosoma 6(1):61–78
Lin MH, Kuo-Kuang Lee R, Li SH, Lu CH, Sun FJ, Hwu YM (2008) Sperm chromatin structure assay parameters are not related to fertilization rates, embryo quality, and pregnancy rates in in vitro fertilization and intracytoplasmic sperm injection, but might be related to spontaneous abortion rates. Fertil Steril 90(2):352–359
Liu K, Mao X, Pan F, Chen Y, An R (2023) Correlation analysis of sperm DNA fragmentation index with semen parameters and the effect of sperm DFI on outcomes of ART. Sci Rep 13(1):2717
Lopes S, Jurisicova A, Sun JG, Casper RF (1998) Reactive oxygen species: potential cause for DNA fragmentation in human spermatozoa. Hum Reprod 13(4):896–900
Manicardi GC, Bianchi P, Pantano S, Azzoni P, Bizzaro D, Bianchi U et al (1995) Presence of endogenous nicks in DNA of ejaculated human spermatozoa and its relationship to chromomycin A3 accessibility. Biol Reprod 52(4):864–867
Meseguer M, Santiso R, Garrido N, Gil-Salom M, Remohà J, Fernandez JL (2009) Sperm DNA fragmentation levels in testicular sperm samples from azoospermic males as assessed by the sperm chromatin dispersion (SCD) test. Fertil Steril 92(5):1638–1645
Mitchell LA, De Iuliis GN, Aitken RJ (2011) The TUNEL assay consistently underestimates DNA damage in human spermatozoa and is influenced by DNA compaction and cell vitality: development of an improved methodology. Int J Androl 34(1):2–13
Mohammed EM, Mosad E, Zahran AM et al (2015) Acridine orange and flow cytometry: which is better to measure the effect of varicocele on sperm DNA integrity? Adv Urol 2015:814150
Muratori M, Tamburrino L, Tocci V, Costantino A, Marchiani S, Giachini C et al (2010) Small variations in crucial steps of TUNEL assay coupled to flow cytometry greatly affect measures of sperm DNA fragmentation. J Androl 31(4):336–345
Muriel L, Meseguer M, Fernández JL, Alvarez J, Remohà J, Pellicer A, Garrido N (2006) Value of the sperm chromatin dispersion test in predicting pregnancy outcome in intrauterine insemination: a blind prospective study. Hum Reprod 21(3):738–744
Patel AS, Leong JY, Ramasamy R (2018) Prediction of male infertility by the World Health Organization laboratory manual for assessment of semen analysis: a systematic review. Arab J Urol 16(1):96–102
Sakkas D, Alvarez JG (2010) Sperm DNA fragmentation: mechanisms of origin, impact on reproductive outcome, and analysis. Fertil Steril 93(4):1027–1036
Sakkas D, Urner F, Bizzaro D, Manicardi G, Bianchi P, Shoukir Y et al (1998) Sperm nuclear DNA damage and altered chromatin structure: effect on fertilization and embryo development. Hum Reprod 13(suppl 4):11–19
Selvam MKP, Sengupta P, Agarwal A (2020) Sperm DNA fragmentation and male infertility genetics of male infertility. Springer, Berlin, pp 155–172
Serrano Berenguer L, Hincapié Monsalve S, Lara Cerrillo S, Rosado Iglesias C, Vegas Lozano E, Reverter Comes F et al (2022) P-101 machine and deep learning models to classify comet assay tests for sperm DNA fragmentation evaluation. Hum Reprod 37(Supplement_1):deac107.097
Shamsi MB, Imam SN, Dada R (2011) Sperm DNA integrity assays: diagnostic and prognostic challenges and implications in management of infertility. J Assist Reprod Genet 28(11):1073–1085
Sharma RK, Sabanegh E, Mahfouz R, Gupta S, Thiyagarajan A, Agarwal A (2010) TUNEL as a test for sperm DNA damage in the evaluation of male infertility. Urology 76(6):1380–1386
Sharma R, Ahmad G, Esteves SC, Agarwal A (2016) Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay using bench top flow cytometer for evaluation of sperm DNA fragmentation in fertility laboratories: protocol, reference values, and quality control. J Assist Reprod Genet 33(2):291–300
Sharma R, Gupta S, Henkel R, Agarwal A (2019) Critical evaluation of two models of flow cytometers for the assessment of sperm DNA fragmentation: an appeal for performance verification. Asian J Androl 21(5):438
Sharma R, Iovine C, Agarwal A, Henkel R (2021) TUNEL assay—standardized method for testing sperm DNA fragmentation. Andrologia 53(2):e13738
Shen HM, Ong CN (2000) Detection of oxidative DNA damage in human sperm and its association with sperm function and male infertility. Free Radic Biol Med 28(4):529–536
Simon L, Lutton D, McManus J, Lewis SEM (2011) Sperm DNA damage measured by the alkaline comet assay as an independent predictor of male infertility and in vitro fertilization success. Fertil Steril 95(2):652–657
Simon L, Zini A, Dyachenko A, Ciampi A, Carrell D (2017) A systematic review and meta-analysis to determine the effect of sperm DNA damage on in vitro fertilization and intracytoplasmic sperm injection outcome. Asian J Androl 19(1):80–90
Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175(1):184–191
Spanò M, Kolstad AH, Larsen SB, Cordelli E, Leter G, Giwereman A et al (1998) The applicability of the flow cytometric sperm chromatin structure assay in epidemiological studies. Hum Reprod 13(9):2495–2505
Sun JG, Jurisicova A, Casper RF (1997) Detection of deoxyribonucleic acid fragmentation in human sperm: correlation with fertilization in vitro. Biol Reprod 56(3):602–607
Tsarev I, Bungum M, Giwercman A, Erenpreisa J, Ebessen T, Ernst E et al (2009) Evaluation of male fertility potential by toluidine blue test for sperm chromatin structure assessment. Hum Reprod 24(7):1569–1574
Vandekerckhove FWRC, De Croo I, Gerris J, Vanden Abbeel E, De Sutter P (2016) Sperm chromatin dispersion test before sperm preparation is predictive of clinical pregnancy in cases of unexplained infertility treated with intrauterine insemination and induction with clomiphene citrate. Front Med 3:63
Zini A, Fischer MA, Sharir S, Shayegan B, Phang D, Jarvi K (2002) Prevalence of abnormal sperm DNA denaturation in fertile and infertile men. Urology 60(6):1069–1072
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Sengupta, P., Dutta, S., Samrot, A.V. (2023). Sperm DNA Fragmentation Testing in Infertility. In: Singh, R. (eds) Genetic Testing in Reproductive Medicine. Springer, Singapore. https://doi.org/10.1007/978-981-99-7028-5_4
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DOI: https://doi.org/10.1007/978-981-99-7028-5_4
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