Usage of polyvinylalcohol and polyvinylpyrrolidone for preparation of thawed ejaculated boar sperm for fertilization by ICSI

O. Ju. Lyzohub

This email address is being protected from spambots. You need JavaScript enabled to view it.

Institute of Animal Breeding and Genetics named after M. V. Zubets NAAS,
1 Pogrebnyaka str, Chubynske village, Boryspil district, Kyiv Region, 08321, Ukraine

The aim of the study was to explore the effect of PVP (polyvinylpyrrolidone) and PVA (polyvinyl alcohol) media on deconserved ejaculated boar sperm and their preparation for artificial insemination to optimize biotechnological approaches. The studies used ejaculated cryopreserved sperm of a boar of the Myrhorod breed Dnipro 641. Genetic material was stored in the Bank of Genetic Resources of Animals IABG named after M. V. Zubets NAAS for eight years. The sperm suspension was thawed in a water bath at +37°C for 5 min until completely thawed. Separation of sperm from cryopreservative agent and diluent was performed using the swim up method in Sp-TALP medium. After the presence of sperm in the 10.0% solution of PVP for 10 min, motility decreased by 68.2% (P<0.05) and amounted to 3.4%, and after the next 10 min of incubation decreased to 1.4% (P<0.01), which is 10 times lower than the initial mobility. In 10.0% of PVA mobility after 10 min of incubation decreased by 37.4% (P<0.05) and amounted to 6.7%, and after 10 min decreased to 5.7% (P<0.01), which is 1.8 times lower than the initial mobility. It was found that in the case of 10.0% of PVP solution ejaculated deconserved boar sperm lose motility by 86.9% (P<0.01) from the initial motility, which makes it impossible to select a suitable sperm for fertilization by ICSI (Intra Cytoplasmic Sperm Injection). It is shown that 10.0% PVA solution can be used for immobilization of boar sperm, as it reduces motility by 46.7% (P<0.01) of the initial sperm motility. It is proved that the mobility in the case of incubation of deconserved ejaculated boar sperm in 5.0% PVA solution decreases only by 28.0% (P<0.05) from the initial, which is optimal when using cryopreserved boar sperm, material which are limited and convenient for the operator and safe for oocytes.

Key words: ICSI, sperm motility, oocyte, polyvinylpyrrolidone, polyvinylalcohol, pig embryos

1. Association of Clinical Embryologists. Guidelines on good practice in clinical embryology laboratories. Human Fertil. 2012; 15 (4): 174–189. DOI: 10.3109/14647273.2012.747891.
2. Avalos-Durán G, Cañedo-Del Ángel AME, Rivero-Murillo BJ, Zambrano-Guerrero JE, Carballo-Mondragón E, Checa-Vizcaíno MÁ. Physiological ICSI (PICSI) vs. Conventional ICSI in couples with male factor: a systematic review. JBRA Assist. Reprod. 2018; 22 (2): 139–147. DOI: 10.5935/1518-0557.20180027.
3. Casillas F, Betancourt M, Cuello C, Ducolomb Y, López A, Juárez-Rojas L, Retana-Márquez An efficiency comparison of different in vitro fertilization methods: IVF, ICSI, and PICSI for embryo development to the blastocyst stage from vitrified porcine immature oocytes. Porc. Health Manag. 2018; 4: 16. DOI: 10.1186/s40813-018-0093-6.
4. Ding D, Wang Q, Li X, Chen B, Zou W, Ji D, Hao Y, Xue R, Zou H, Wei Z, Zhou P, Cao Y, Zhang Z. Effects of different polyvinylpyrrolidone concentrations on intracytoplasmic sperm injection. Zygote. 2020; 28 (2): 148–153. DOI: 10.1017/S0967199419000820.
5. Kato Y, Nagao Y. Effect of polyvinylpyrrolidone on sperm function and early embryonic development following intracytoplasmic sperm injection in human assisted reproduction. Med. Biol. 2012; 11 (4): 165–176. DOI: 10.1007/s12522-012-0126-9.
6. Kouba AJ, Abeydeera LR, Alvarez IM, Day BN, Buhi WC. Effects of the porcine oviduct-specific glycoprotein on fertilization, polyspermy, and embryonic development in vitro. Reprod. 2000; 63 (1): 242–250. DOI: 10.1095/biolreprod63.1.242.
7. Li XX, Lee DS, Kim KJ, Lee JH, Kim EY, Park JY, Kim MK. Leptin and nonessential amino acids enhance porcine preimplantation embryo development in vitro by intracytoplasmic sperm injection. Theriogenol. 2013; 79 (2): 291–298. DOI: 10.1016/j.theriogenology.08.019.
8. Macedo MP, Glanzner WG, Rissi VR, Gutierrez K, Currin L, Baldassarre H, Bordignon V. A fast and reliable protocol for activation of porcine oocytes. 2018; 123: 22–29. DOI: 10.1016/j.theriogenology.2018.09.021.
9. Mandryk-Grad I, Kosenyuk J, Gajda B. Developmental competence and quality of pig embryos obtained from standart in vitro fertilization or intracytoplasmic sperm injection. Fertil. Dev. 2012; 25 (1): 260–261. DOI: 10.1071/RDv25n1Ab225.
10. Nabi A, Entezari F, Miresmaeili SM, Vahidi S, Lorian K, Anbari F, Motamedzadeh L. Evaluation of sperm parameters and DNA integrity following different incubation times in PVP medium. J. 2022; 19 (3): 232–237. DOI: 10.22037/uj.v18i.6936.
11. Palermo GD, Nagy ZP (eds). Manual of intracytoplasmic sperm injection in human assisted reproduction with other advanced micromanipulation techniques to edit the genetic and cytoplasmic content of the oocyte. Cambridge, Cambridge university press, 2021: 171 p. DOI: 10.1017/9781108887595.
12. Parmegiani L, Cognigni GE, Ciampaglia W, Pocognoli P, Marchi F, Filicori M. Efficiency of hyaluronic acid (HA) sperm selection. Assist. Reprod. Genet. 2010; 27: 6–13. DOI: 10.1007/s10815-009-9380-0.
13. Petelak A, Krylov A. Surface sperm cell ubiquitination directly impaired blastocyst formation rate after intracytoplasmic sperm injection in pig. Theriogenol. 2019; 135: 115–120. DOI: 10.1016/theriogenology.2019.06.014.
14. Pinto S, Carrageta DF, Alves MG, Rocha A, Agarwal A, Barros A, Oliveira PF. Sperm selection strategies and their impact on assisted reproductive technology outcomes. 2020; 53 (2): e13725. DOI: 10.1111/and.13725.
15. Rajska I. Intra-cytoplasmic sperm injection (ICSI) as an alternative to standard in vitro fertilization in pigs. Ann. Polish Soc. Anim. Prod. 2015; 11 (3): 55–66. Available at: http://rn.ptz.icm.edu.pl/wp-content/uploads/2016/12/Rajska-ang..pdf
16. Ribas-Maynou J, Yeste M, Salas-Huetos A. The relationship between sperm oxidative stress alterations and IVF/ICSI outcomes: a systematic review from nonhuman mammals. 2020; 9 (7): 178. DOI: 10.3390/biology9070178.
17. Roychoudhury S, Maldonado-Rosas I, Agarwal A, Esteves S, Sharma R, Gupta S, Assidi M. Parthenogenetic activation and developmental potential of mouse oocytes after intracytoplasmic injection (ICSI) of PVP (polyvinylpyrrolidone) and HA (hyaluronic acid). Steril. 2016; 106 (3): E311. DOI: 10.1016/j.fertnstert.2016.07.883.
18. Wu J, Carrell DT, Wilcox AL. Development of in vitro-matured oocytes from porcine preantral follicles following intracytoplasmic sperm injection. Reprod. 2001; 65 (5): 1579–1585. DOI: 10.1095/biolreprod65.5.1579.
19. Yamochi T, Hashimoto S, Morimoto Y. Mural granulosa cells support to maintain the viability of growing porcine oocytes and its developmental competence after insemination. Assist. Reprod. Genet. 2021; 38: 2591–2599. DOI: 10.1007/s10815-021-02212-2.
20. Yong HY, Pyo BS, Hong JY, Kang KS, Lee BC, Lee ES, Hwang WS. A modifed method for ICSI in the pig: injection of head membrane-damaged sperm using a 3–4 mm diameter injection pipette. Human Reprod. 2003; 18 (11): 2390–2396. DOI: 10.1093/humrep/deg442.