The effects of protamine deficiency on ultrastructure of human sperm nucleus

Author

Anatomical Sciences Department, Isfahan University of Medical Sciences, Isfahan, Iran

Abstract

Background: Chromomycin A3 (CMA3) staining is one of the staining methods for detecting protamine deficiency in sperm nucleus. CMA3 is a fluorochrome that competes with protamines for binding to DNA double helix. It has been shown in our previous studies that percentage of CMA3 positive spermatozoa in semen has a close significant relationship with the fertilization rate in in vitro fertilization (IVF). The aim of this study was to examine the ultrastructural differences between sperms in patients who had high fluorescent percentages of yellow or red in CMA3 staining (protamine deficient) with patients with low fluorescent percentages.
Materials and Methods: Semen samples are taken from five patients with high fluorescent percentages and five patients with low fluorescent percentages. Then the samples are passed for the different steps of preparing for electron microscopy. After the sectioning and mounting on grids, they are investigated under the transmission electron microscope.
Results: Sperms in patients with low percentages of positive spermatozoa often have a normal appearance. Sperms in high fluorescent samples frequently have unpacked chromatin. Furthermore acrosomes of these sperms are thinner or disturbed. Also sometimes there are irregularities in sperm head membrane.
Conclusion: Protamine deficiency in sperm nucleus can cause ultrastructural anomalies in sperm chromatin such as unpacking of it. It also is concomitant with acrosome and sperm membrane disturbances.

Keywords

1. Baccetti B, Bruni E, Collodel G, Gambera L, Moretti E, Marzella R, et al. 10,15 reciprocal translocation in an infertile man: Ultrastructural and fluorescence in-situ hybridization sperm study: Case report. Hum Reprod 2003;18:2302-8.  Back to cited text no. 1
[PUBMED]    
2. Collodel G, Moretti E, Capitani S, Estenoz M, Manca D, Pimboni P, et al. Ultrastructural sperm study in infertile males with microdeletions of Y chromosome. J Submicrosc Cytol Pathol 2006;38:45-50.  Back to cited text no. 2
    
3. Baccetti B, Collodel G, Marzella R, Moretti E, Pimboni P, Scapigliati G, et al. Ultrastructural studies of spermatozoa from infertile males with Robertsonian translocations and 18, X,Y aneuploidies. Hum Reprod 2005;20:2295-300.  Back to cited text no. 3
    
4. Collodel G, Cosci I, Pascarelli AN, Moretti E. Sperm ultrastructure and 18, X, Y aneuploidies in man with a 46 XY, 47 XY+18 mosaic karyotype: Case report. J Assist Reprod Genet 2007;24:373-6.  Back to cited text no. 4
[PUBMED]    
5. Moretti E, Pascarelli NA, Giannerini V, Geminiani M, Anichini C, Colledel G. 18, X, Y aneuploidies and transmission electron microscopy studies in spermatozoa from five carries of different reciprocal translocations. Asian J Androl 2009;11:325-32.  Back to cited text no. 5
    
6. Pimboni P, Bruni E, Capitani S, Gambera L, Moretti E, La Marca A, et al. Ultrastructural and DNA fragmentation analyses in swim-up selected human sperm. Arch Androl 2006;52:51-9.  Back to cited text no. 6
    
7. Mantas D, Angelopoulou R, Msaouel P, Plastria K. Evaluation of sperm chromatin quality and screening of Y chromosome microdeletions in Greek males with severe oligozoospermia. Arch Androl 2007;53:5-8.  Back to cited text no. 7
    
8. Prisant N, Escalier D, Soufir JC, Morillon M, Schoevaert D, Misrahi M, et al. Ultrastructural nuclear defects and increased chromosome aneuploidies in spermatozoa with elongated heads. Hum Reprod 2007;22:1052-9.  Back to cited text no. 8
[PUBMED]    
9. Collodel G, Giannerini V, Pascarelli AN, Federico MG, Comodo F, Moretti E. TEM and FISH studies in sperm from men of couples with recurrent pregnancy loss. Andrologia 2009;41:352-60.  Back to cited text no. 9
    
10. Collodel G, Capitani S, Pammoli A, Giannerini V, Gemiani M, Moretti E. Semen quality of male idiopathic infertile smokers and non smokers: An ultrastructural study. J Androl 2010;31:108-13.  Back to cited text no. 10
    
11. Collodel G, Capitani S, Iacoponi F, Federico MG, Passcarelli NA, Moretti E. Retrospective assessment of potential negative synergistic effects of varicocele and tobacco use on ultrastructural sperm morphology. Urology 2009;74:794-9.  Back to cited text no. 11
    
12. Balhorn R. Mammalian protamines: Structure and molecular interactions. In: Adolph KW, editor. Molecular biology of chromosome function. New York: Springer-Verlag; 1989. p. 366-95.  Back to cited text no. 12
    
13. Hecht JJ. Mammalian protamines and their expression. In: Hnilica L, Stein G, Stein J, editors. Histones and other basic nuclear proteins. Boca Raton, Florida: CRC Press; 1989.  Back to cited text no. 13
    
14. Ward WS, Coffey DS. DNA packaging and organization in mammalian spermatozoa: Comparison with somatic cells. Bio Reprod 1989;44:569-74.  Back to cited text no. 14
    
15. Zamboni L. The ultrastructural pathology of the spermatozoon as a cause of infertility: The role of electron microscopy in the evaluation of semen quality. Fertil Steril 1987;48:711-34.  Back to cited text no. 15
[PUBMED]    
16. Bustos-Obergon L, Leiva S. Chromatin packing in normal and teratozoospermic human ejaculated spermatozoa. Andrologia 1983;15:468-78.  Back to cited text no. 16
    
17. Zamboni L. Sperm ultrastructural pathology and infertility. In: Gondos B, Riddick D, editors. Pathology of infertility. New York, NY: Thieme-Stratton Inc; 1987. p. 353-84.  Back to cited text no. 17
    
18. Zamboni L. Physiology and pathophysiology of the human spermatozoon: The role of electron microscopy. J Electron Microsc Tech 1991;7:412-36.  Back to cited text no. 18
    
19. Jager S. Sperm nuclear stability and male infertility. Arch Androl 1990;25:253-9.  Back to cited text no. 19
[PUBMED]    
20. Berman E, Brown SC, James TL, Shafer RH. NMR studies of chromomycin A3 interaction with DNA. Biochemistry 1985;24:6887-93.  Back to cited text no. 20
[PUBMED]    
21. Sakkas D, Manicardi GC, Bianchi PG, Bizzaro D, Bianchi U. Relationship between the presence of endogenous nicks and sperm chromatin packaging in maturing and fertilizing mouse spermatozoa. Biol Reprod 1995;52:1149-55.  Back to cited text no. 21
    
22. Iranpour FG, Nasr-Esfahani MH, Valojerdi MR, Al-Taraihi TM. Chromomycin A3 as a useful tool for evaluation of male fertility. J Assist Reprod Genet 2000;17:60-6.  Back to cited text no. 22
[PUBMED]    
23. Ward WS, Coffey DS. DNA packaging and organization in mammalian spermatozoa: Comparison with somatic cells. Biol Reprod 1991;44:569-74.  Back to cited text no. 23
[PUBMED]    
24. Bianchi PG, Manicardi GC, Urner F, Compana A, Sakkas D. Chromatin packaging and morphology in ejaculated human spermatozoa: Evidence of hidden anomalies in normal spermatozoa. Mol Hum Reprod 1996;3:139-44.  Back to cited text no. 24
    
25. Balhorn R, Reed S, Tanphaichitr N. Aberrant protamine 1/protamine 2 ratios in sperm of infertile human males. Experientia 1988;44:53-5.  Back to cited text no. 25
    
26. de Yerba L, Ballesca JL, Vornell JA, Bassas L, Oliva R. Complete selective absence of protamine P2 in humans. J Biol Chem 1993;268:10553-7.  Back to cited text no. 26
    
27. Belkopytova IA, Kostyleva EI, Tomilin AN, Vorobev VI. Human male infertility may be due to a decrease of protamine P2 content of sperm chromatin. Mol Reprod Dev 1993;34:53-7.  Back to cited text no. 27
    
28. Bianchi PG, Manicardi GC, Bizzaro D, Bianchi U, Sakkas D. Effect of deoxyribonucleic acid protamination on fluorochrome staining and in situ nick-translation of marine and human mature spermatozoa. Biol Reprod 1993;49:1083-8.  Back to cited text no. 28
[PUBMED]    
29. Sailor 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;16:80-7.  Back to cited text no. 29
    
30. Manicardi GC, Bianchi PG, Pantano S. Under protamination and nicking of DNA in ejaculated human spermatozoa are highly related phenomena. Biol Reprod 1995;52:864-7.  Back to cited text no. 30
    
31. Gorczyca W, Traganos F, Jesionowska H, Darzynkiewics Z. Presence of strand breaks and increased sensitivity of DNA in situ to denaturation in abnormal human sperm cell: Analogy to apoptosis of somatic cells. Exp Cell Res 1993;207:202-5.  Back to cited text no. 31
    
32. Marsh SK, Bolton VN, Braude PR. The effect of morphology on the ability of human spermatozoa to penetrate zona-free hamster oocytes. Hum Reprod 1987;2:499-503.  Back to cited text no. 32
[PUBMED]    
33. 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.  Back to cited text no. 33
[PUBMED]    
34. 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.  Back to cited text no. 34
[PUBMED]    
35. Chevaillier PH, Mauro N, Feneux D, Jouannet P, David G. Anomalous protein complement of sperm nuclei in some infertile men. Lancet 1978;8562:806-7.  Back to cited text no. 35
    
36. Sakkas D, Urner F, Bianchi PG, Bizzaro D, Wagner I, Jaquenoud N, et al. Sperm chromatin anomalies can influence decondensation after intracytoplasmic sperm injection. Hum Reprod 1996;4:837-43.  Back to cited text no. 36
    
37. Balhorn R. A model for the structure of chromatin in mammalian sperm. J Cell Biol 1982;93:298-305.  Back to cited text no. 37
[PUBMED]    
38. Perreault SD. Chromatin remodeling in mammalian zygotes. Mutat Res 1992;296:43-55.  Back to cited text no. 38
[PUBMED]    
39. Berman E, Brown SC, James TL, Shafer RH. NMR studies of Chromomycin A3 interaction with DNA. Biochemistry 1985;24:6887-93.  Back to cited text no. 39
[PUBMED]    
40. Behr W, Honikel K, Hartmann G. Interaction of the RNA polymerase inhibitor chromomycin with DNA. Eur J Biochem 1969;9:82-92.  Back to cited text no. 40
[PUBMED]    
41. Nasr-Esfahani MH, Razavi S, Anjomshoa M, Rozbahani S, Moulavi F, Mardani M. Effect of sperm DNA damage and sperm protamine deficiency on fertilization and embryo development post-ICSI. Reprod Biomed Online 2005;11:198-205.  Back to cited text no. 41
[PUBMED]    
42. Nasr-esfahani MH, Razavi S, Mozdarani H, Mardani M, Azvaqi H. Relationship between protamine deficiency with fertilization rate and incidence of sperm premature chromosomal condensation post-ICSI. Andrologia 2004;36:95-100.  Back to cited text no. 42
    
43. Razavi S, Nasr-Esfahani MH, Mardani M, Mafi A, Moghadam A. Effect of human sperm chromatin anomalies on fertilization outcome post-ICSI. Andrologia 2003;35:238-43.  Back to cited text no. 43
    
44. Tavalaee M, Razavi S, Nasr-Esfahani MH. Influence of sperm chromatin anomalies on assisted reproductive technology out come. Fertil Steril 2009;91:1119-26.  Back to cited text no. 44
[PUBMED]    
45. Tavalaee M, Razavi S, Nasr-Esfahani MH. Effect of sperm acrosomal integrity and protamine deficiency on In vitro fertilization and pregnancy. Iran J Fertil Steril 2007;1:27-34  Back to cited text no. 45
    
46. Gill-Sharrma MK, Choudhuri J, Dsouza S. Sperm chromatin protamination: An endocrine perspective. Protein Pept Lett 2011;18:786-801.  Back to cited text no. 46
    
47. Aleem M, Padwal V, Choudhari J, Balasinor N, Gill-Sharma MK. Sperm protamine levels as indicator of fertilizing potential in sexually mature male. Andrologia 2008;40:29-37.  Back to cited text no. 47
[PUBMED]    
48. Nasr-Esfahani MH, Naghshizadian N, Imani H, Razavi S, Mardani M, Kazemi S, et al. Can sperm protamine deficiency induce sperm premature chromosomal condensation? Andrologia 2006;38:92-8.  Back to cited text no. 48
[PUBMED]    
49. Perdix A, Trevers A, Chelli MH, Esalier D, Do Reqo JL, Milazzo JP, et al. Assessment of acrosome and nuclear abnormalities in human spermatozoa with large vacuoles. Human Reprod 2011;26:47-58.  Back to cited text no. 49