Ultrastructural and Functional Characteristics of Human Spermatozoa After Cryopreservation by Vitrification

Authors

  • Olena Pavlovych Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv
  • Hanna Hapon Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv
  • Taisiia Yurchuk Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv
  • Mykola Repin Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv
  • Larysa Marchenko Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv
  • Tetiana Govorukha Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv
  • Maryna Petrushko Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

DOI:

https://doi.org/10.15407/cryo30.01.024

Keywords:

spermatozoa, cryopreservation, vitrification, asthenozoospermia, polyvinylpyrrolidone

Abstract

Abstract: Low temperature preservation of spermatozoa is widely applied in infertility treatment when using the assisted reproductive technology (ART). There are the standard methods for ejaculated spermatozoa cryopreservation in normozoospermia, but their application for ejaculate-derived spermatozoa, having spermatogenesis defects, is impossible. Therefore, of high priority are to design the cryopreservation methods and assess morphofunctional and ultrastructural characteristics of spermatozoa after cryopreservation. Here, we have evaluated the impact of cryopreservation by vitrification using non-penetrating cryoprotectants of polyvinylpyrrolidone (PVP) and sucrose on morphofunctional and ultrastructural state of human spermatozoa in pathospermia. In (22.3 ± 3.4),(26.8 ± 4.2), (18.6 ± 2.1)% of spermatozoa after vitrifi cation with 0.25M sucrose; 10% PVP and a mixture of 0.25 M sucrose, 10% PVP, 10% HSA, respectively, we have observed the appearance of vacuoles in nuclear chromatin of spermatozoa. For freshly isolated spermatozoa this index was (16.6 ± 1.4)%. In the studied groups, the ultrastructural characteristics of acrosome, axoneme, periaxonemal structures, and spermatozoa flagella did not significantly differ from the control samples. The cryopreservation of human spermatozoa by vitrification using sucrose and PVP solutions was shown to preserve their viability and ultrastructural integrity and to be promising in ART.

 

Probl Cryobiol Cryomed 2020; 30(1): 024–033

Author Biographies

Olena Pavlovych , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Department of Cryobiology of Reproductive System

Hanna Hapon , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Department of Cryobiology of Reproductive System

Taisiia Yurchuk , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Department of Cryobiology of Reproductive System

Mykola Repin , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Laboratory of Cryomorphology

Larysa Marchenko , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Laboratory of Cryomorphology

Tetiana Govorukha , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Laboratory of Cryomorphology

Maryna Petrushko , Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv

Department of Cryobiology of Reproductive System

References

Afzelius B. Electron microscopy of the sperm tail results obtained with a new fixative. J Bioph Biochem Cytol. 1959;5(2):269-78. CrossRef

Aizpurua J, Medrano L, Enciso M et al. New permeable cryoprotectant-free vitrification method for native human sperm. Hum Reprod. 2017;32(10):2007-15. CrossRef

Bragina EE, Sorokina TM, Arifulin EA, Kurilo LF. [Genetically determined patozoospermia.] Moscow: SOPEK-polygrapfia. 2002. 93 p. Russian.

Chemes HE, Rawe VY. The making of abnormal spermatozoa: cellular and molecular mechanisms underlying pathological spermiogenesis. Cell Tissue Res. 2010;341(3):349-57. CrossRef

Chohan KR, Griffin JT, Carrell DT. Evaluation of chromatin integrity in human sperm using acridine orange staining with different fixatives and after cryopreservation. Androl. 2004;36(5):321-6. CrossRef

Conklin B, Richter E, Kreutziger K et al. Development and evaluation of a novel decellularized vascular xenograft. Med Eng Physics. 2002;24(3):173-83. CrossRef

Isachenko V, Isachenko E, Petrunkina AM, Sánchez R. Human spermatozoa vitrified in the absence of permeable cryoprotectants: birth of two healthy babies. Reprod Fertil. 2012;24(2):323-6. CrossRef

Isachenko V, Isachenko E, Katkov I, et al. Cryoprotectant-free cryopreservation of human spermatozoa by vitrification and freezing in vapor: effect on motility, DNA integrity, and fertilization ability. Biol Reprod. 2004;71(4):1167-73. CrossRef

Isachenko V, Maettner R, Petrunkina AM, et al. Cryoprotectant free vitrification of human spermatozoa in large (to 0.5 mL) volume: novel technology. Clin Lab. 2011;57(9-10):643-50. PubMed

Isachenko EP, Mallmann G, Rahimi J, et al. Vitrification technique-new possibilities for male gamete low-temperature storage. In: Katkov II, editor. Current Frontiers in Cryobiology. InTech Open Access Books; 2012. Chapter 1; p. 41-76. [Cited 01.03.2019]. Available from: https://www.intechopen.com/books/current-frontiers-in-cryobiology/vitrification-of-fish-and-dog-spermatozoa. CrossRef

Fahy GM, Levy DI, Ali SE. Some emerging principles underlying the physical properties, biological actions, and utility of vitrification solutions. Cryobiology. 1987,24(3):196-213. CrossRef

Khayat SSh, Bragina EE, Kurilo LF. [Ultrastructural investigation of human sperm from asthenozoospermic men.] J Andr Genit Surg. 2012,13(4):54-61. Russian.

Mohamed MSA. Slow cryopreservation is not superior to vitrification in human spermatozoa; an experimental controlled study. Int J Reprod Med. 2015;13(10):633-44. PubMed

Nawroth F, Isachenko V, Dessole S et al. Vitrification of human spermatozoa without cryoprotectants. CryoLetters. 2002,23(2):93-102. PubMed

Ozkavukcu S, Erdemli E, Isik A, et al. Effects of cryopreservation on sperm parameters and ultrastructural morphology of human spermatozoa. J Assist Reprod Genet. 2008; 25(8): 403-11. CrossRef

Petrushko MP. [Сurrent state of cryopreservation of reproductive cells and embryos.] Visn Nac Akad Nauk Ukr. 2017;(7):44-52. Ukrainian. CrossRef

Petrushko MP The cytogenetic analysis of unfertilized human oocytes. Tsitol Genet. 2003;37(6): 60-5.

Petrushko МP, Pavlovich EV, Pinyaev VІ et al. Apoptosis and processes of DNA fragmentation in native and cryopreserved human sperm cells at normo- and pathospermia. Tsitol Genet. 2017, 51(4): 278-81. CrossRef

Repin NV, Marchenko LN, Govorukha TP, Mikhailova IP, Manchenko AA, Sandomirsky BP. Ultrastructure of porcine pericardium and aortic valve tissues, devitalized with cryogenic and irradiation exposures. Probl Cryobiol Cryomed. 2015,25(3):246-54. CrossRef

Virant-Klun I, Tomazevic T, Meden-Vrtovec H. Sperm single-stranded DNA, detected by acridine orange staining, reduces fertilization and quality of ICSI-derived embryos. J Assist Reprod Genet. 2002,19(7):319-28. CrossRef

WHO laboratory manual for the Examination and processing of human semen. Fifth edition. World Health Organization, Department of Reproductive Health and Research. 2010. 287p.

Zheng WW, Song G, Wang QL, et al. Sperm DNA damage has a negative effect on early embryonic development following in vitro fertilization. Asian J Androl. 2018;20(1):75-9. CrossRef

Published

2020-03-28

How to Cite

Pavlovych , O., Hapon , H., Yurchuk , T., Repin , M., Marchenko , L., Govorukha , T., & Petrushko , M. (2020). Ultrastructural and Functional Characteristics of Human Spermatozoa After Cryopreservation by Vitrification. Problems of Cryobiology and Cryomedicine, 30(1), 24-33. https://doi.org/10.15407/cryo30.01.024

Issue

Section

Theoretical and Experimental Cryobiology