Clinical and Experimental Reproductive Medicine

The Korean Society for Reproductive Medicine (대한생식의학회)
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Efficacy of testicular sperm chromatin condensation assay using aniline blue-eosin staining in the IVF-ET cycle

  • Park, Yong-Seog (Laboratory of Reproductive Biology and Infertility, Cheil General Hospital and Women's Healthcare Center, Kwandong University College of Medicine) ;
  • Kim, Myo-Kyung (Laboratory of Reproductive Biology and Infertility, Cheil General Hospital and Women's Healthcare Center, Kwandong University College of Medicine) ;
  • Lee, Sun-Hee (Laboratory of Reproductive Biology and Infertility, Cheil General Hospital and Women's Healthcare Center, Kwandong University College of Medicine) ;
  • Cho, Jae-Won (Laboratory of Reproductive Biology and Infertility, Cheil General Hospital and Women's Healthcare Center, Kwandong University College of Medicine) ;
  • Song, In-Ok (Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Kwandong University College of Medicine) ;
  • Seo, Ju-Tae (Department of Urology, Cheil General Hospital and Women's Healthcare Center, Kwandong University College of Medicine)
  • Received : 2011.06.15
  • Accepted : 2011.08.22
  • Published : 2011.09.30
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Abstract

Objective: This study was performed to evaluate testicular sperm chromatin condensation using aniline blue-eosin (AB-E) staining and its effects on IVF-ET. Methods: Chromatin condensation was analyzed using AB-E staining in 27 cases of testicular sperm extraction. There were 19 cases of obstructive azoospermia (OA) and 8 cases of non-obstructive azoospermia (NOA) in IVF-ET. Mature sperm heads were stained red-pink whereas immature sperm heads were stained dark blue. The percentage of sperm chromatin condensation was calculated from the ratio of the number of red-pink sperm to the total number of sperm analyzed. Results: The overall percentages of chromatin condensation in OA and NOA were $31.1{\pm}11.2%$ and $26.3{\pm}14.4%$, respectively. The fertilization rate was significant higher in OA than NOA ($p$ <0.05); however, the rates of good embryos and clinical pregnancy did not show statistical differences. In OA and NOA, statistical differences were not observed in the rate of chromatin condensation, fertilization, good embryos, and clinical pregnancy between the pregnant group and non-pregnant group. Conclusion: Chromatin condensation is less stable than OA and showed a low fertilization rate in NOA. While there were no significant differences in chromatin condensation results between NOA and OA, we propose that a pattern of decreased chromatin condensation in NOA is one of the factors of low fertilization results requiring further study.

Keywords

References

  1. Hofmann N, Hilscher B. Use of aniline blue to assess chromatin condensation in morphologically normal spermatozoa in normal and infertile men. Hum Reprod 1991;6:979-82. https://doi.org/10.1093/oxfordjournals.humrep.a137472
  2. Sun JG, Jurisicova A, Casper RF. Detection of deoxyribonucleic acid fragmentation in human sperm: correlation with fertilization in vitro. Biol Reprod 1997;56:602-7. https://doi.org/10.1095/biolreprod56.3.602
  3. Spano M, Bonde JP, Hjollund HI, Kolstad HA, Cordelli E, Leter G. Sperm chromatin damage impairs human fertility. The Danish First Pregnancy Planner Study Team. Fertil Steril 2000;73:43-50. https://doi.org/10.1016/S0015-0282(99)00462-8
  4. Evenson DP, Jost LK, Marshall D, Zinaman MJ, Clegg E, Purvis K, et al. Utility of the sperm chromatin structure assay as a diagnostic and prognostic tool in the human fertility clinic. Hum Reprod 1999;14:1039-49. https://doi.org/10.1093/humrep/14.4.1039
  5. Evenson DP, Larson KL, Jost LK. Sperm chromatin structure assay: its clinical use for detecting sperm DNA fragmentation in male infertility and comparisons with other techniques. J Androl 2002;23:25-43. https://doi.org/10.1002/j.1939-4640.2002.tb02599.x
  6. Wong A, Chuan SS, Patton WC, Jacobson JD, Corselli J, Chan PJ. Addition of eosin to the aniline blue assay to enhance detection of immature sperm histones. Fertil Steril 2008;90:1999-2002. https://doi.org/10.1016/j.fertnstert.2007.09.026
  7. Park YS, Shon JH, Lee JS, Seo JT. Efficacy of the aniline blue-eosin staining method for testicular sperm chromatin condensation assay in azoospermia. Korean J Androl 2010;28:107-11.
  8. Park YS, Lee SH, Song SJ, Jun JH, Koong MK, Seo JT. Influence of motility on the outcome of in vitro fertilization/intracytoplasmic sperm injection with fresh vs. frozen testicular sperm from men with obstructive azoospermia. Fertil Steril 2003;80:526-30. https://doi.org/10.1016/S0015-0282(03)00798-2
  9. Park YS, Choi SJ, Lee SH, Park DW, Lim CK, Jun JH, et al. Comparative results of embryo development and clinical pregnancy using sperm retrieved from fresh and frozen-thawed testicular tissue from patients with obstructive and non-obstructive azoospermia. Korean J Reprod Med 2009;36:301-10.
  10. Golan R, Shochat L, Weissenberg R, Soffer Y, Marcus Z, Oschry Y, et al. Evaluation of chromatin condensation in human spermatozoa: a flow cytometric assay using acridine orange staining. Mol Hum Reprod 1997;3:47-54. https://doi.org/10.1093/molehr/3.1.47
  11. Agarwal A, Said TM. Role of sperm chromatin abnormalities and DNA damage in male infertility. Hum Reprod Update 2003;9:331-45. https://doi.org/10.1093/humupd/dmg027
  12. Auger J, Mesbah M, Huber C, Dadoune JP. Aniline blue staining as a marker of sperm chromatin defects associated with different semen characteristics discriminates between proven fertile and suspected infertile men. Int J Androl 1990;13:452-62. https://doi.org/10.1111/j.1365-2605.1990.tb01052.x
  13. Hammadeh ME, al-Hasani S, Stieber M, Rosenbaum P, Kupker D, Diedrich K, et al. The effect of chromatin condensation (aniline blue staining) and morphology (strict criteria) of human spermatozoa on fertilization, cleavage and pregnancy rates in an intracytoplasmic sperm injection programme. Hum Reprod 1996;11:2468-71. https://doi.org/10.1093/oxfordjournals.humrep.a019139
  14. Hammadeh ME, Zeginiadov T, Rosenbaum P, Georg T, Schmidt W, Strehler E. Predictive value of sperm chromatin condensation (aniline blue staining) in the assessment of male fertility. Arch Androl 2001;46:99-104. https://doi.org/10.1080/01485010117363
  15. Foresta C, Zorzi M, Rossato M, Varotto A. Sperm nuclear instability and staining with aniline blue: abnormal persistence of histones in spermatozoa in infertile men. Int J Androl 1992;15:330-7. https://doi.org/10.1111/j.1365-2605.1992.tb01132.x
  16. Laberge RM, Boissonneault G. Chromatin remodeling in spermatids: a sensitive step for the genetic integrity of the male gamete. Arch Androl 2005;51:125-33. https://doi.org/10.1080/014850190518134
  17. 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;17:990-8. https://doi.org/10.1093/humrep/17.4.990
  18. Thompson-Cree ME, McClure N, Donnelly ET, Steele KE, Lewis SE. Effects of cryopreservation on testicular sperm nuclear DNA fragmentation and its relationship with assisted conception outcome following ICSI with testicular spermatozoa. Reprod Biomed Online 2003;7:449-55. https://doi.org/10.1016/S1472-6483(10)61889-5
  19. Seli E, Gardner DK, Schoolcraft WB, Moffatt O, Sakkas D. Extent of nuclear DNA damage in ejaculated spermatozoa impacts on blastocyst development after in vitro fertilization. Fertil Steril 2004;82:378-83. https://doi.org/10.1016/j.fertnstert.2003.12.039
  20. Lopes S, Sun JG, Jurisicova A, Meriano J, Casper RF. Sperm deoxyribonucleic acid fragmentation is increased in poor-quality semen samples and correlates with failed fertilization in intracytoplasmic sperm injection. Fertil Steril 1998;69:528-32. https://doi.org/10.1016/S0015-0282(97)00536-0
  21. Jones GM, Trounson AO, Lolatgis N, Wood C. Factors affecting the success of human blastocyst development and pregnancy following in vitro fertilization and embryo transfer. Fertil Steril 1998;70:1022-9. https://doi.org/10.1016/S0015-0282(98)00342-2
  22. Gandini L, Lombardo F, Paoli D, Caruso F, Eleuteri P, Leter G, et al. Full-term pregnancies achieved with ICSI despite high levels of sperm chromatin damage. Hum Reprod 2004;19:1409-17. https://doi.org/10.1093/humrep/deh233
  23. Larson KL, DeJonge CJ, Barnes AM, Jost LK, Evenson DP. Sperm chromatin structure assay parameters as predictors of failed pregnancy following assisted reproductive techniques. Hum Reprod 2000;15:1717-22. https://doi.org/10.1093/humrep/15.8.1717
  24. 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;75:674-7. https://doi.org/10.1016/S0015-0282(00)01796-9
  25. Larson-Cook KL, Brannian JD, Hansen KA, Kasperson KM, Aamold ET, Evenson DP. Relationship between the outcomes of assisted reproductive techniques and sperm DNA fragmentation as measured by the sperm chromatin structure assay. Fertil Steril 2003;80:895-902. https://doi.org/10.1016/S0015-0282(03)01116-6
  26. Tomlinson MJ, Moffatt O, Manicardi GC, Bizzaro D, Afnan M, Sakkas D. Interrelationships between seminal parameters and sperm nuclear DNA damage before and after density gradient centrifugation: implications for assisted conception. Hum Reprod 2001;16:2160-5. https://doi.org/10.1093/humrep/16.10.2160
  27. Tesarík J, Kopecny V, Plachot M, Mandelbaum J. Activation of nucleolar and extranucleolar RNA synthesis and changes in the ribosomal content of human embryos developing in vitro. J Reprod Fertil 1986;78:463-70. https://doi.org/10.1530/jrf.0.0780463
  28. Sakkas D, Manicardi GC, Tomlinson M, Mandrioli M, Bizzaro D, Bianchi PG, et al. The use of two density gradient centrifugation techniques and the swim-up method to separate spermatozoa with chromatin and nuclear DNA anomalies. Hum Reprod 2000;15:1112-6. https://doi.org/10.1093/humrep/15.5.1112
  29. Virro MR, Larson-Cook KL, Evenson DP. Sperm chromatin structure assay (SCSA) parameters are related to fertilization, blastocyst development, and ongoing pregnancy in in vitro fertilization and intracytoplasmic sperm injection cycles. Fertil Steril 2004;81:1289-95. https://doi.org/10.1016/j.fertnstert.2003.09.063

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