The Massive Impact of Ram’s Sperm Starvation on the Fertilization and Blastocyst Rates in Terms of Sperm Quality and Capacitation

During sperm incubation in vitro, the sperm capacitation medium must play a key role in establishing conditions that perfect the required physiological requirements for sperm metabolic activities to obtain a high in vitro fertilization rate ensured by the ideal development of the embryos. Therefore, sperm collected from the caudal epididymis of rams should acquire progressive motility and fertilization. This process occurs through physiological and biochemical changes known as capacitation, a prerequisite for fertilization. In this work, we have studied a new way to incubate sperm, applied for the first time in ram, using four different treatments in terms of energy substrates and different incubation methods. In this sperm energy restriction and recovery treatment, sperm was let starving for 50 min until it lost its capacity for motility, and then was provided with glucose and incubated for 60 min. In the other three treatments, the sperm was not exposed to starvation and was incubated by the standard methods of maturation and capacitation by evaluating different energy substrates. According to the obtained results, the sperm was hyperactive and showed a significant increase in the ability to fertilize oocytes in the treatment that involved starvation and then re-activation of the sperm by adding glucose, compared to other treatments. In conclusion, the effects of this treatment persisted until after fertilization, which led to the production of a high percentage of blastocysts.

Keywords:

Sheep, Sperm Starvation, Blastocyst IVF Capacitation, Rescue Protocol,

PDF

___

Amiridis G S & Cseh S (2012). Assisted reproductive technologies in the reproductive management of small ruminants. Animal reproduction science 130(3-4): 152-161.‏ doi.org/10.1016/j.anireprosci.2012.01.009
Anderson M J, Chapman S J, Videan E N, Evans E, Fritz J, Stoinski T S, Dixson A F & Gagneux P (2007). Functional evidence for differences in sperm competition in humans and chimpanzees. American Journal of Physical Anthropolog 134(2): 274-280. doi.org/10.1002/ajpa.20674
Binder N K, Hannan N J & Gardner D K (2015). In vitro embryo outgrowth is a bioassay of in vivo embryo implantation and development. Asian Pacific Journal of Reproduction 4(3): 240-241. doi.org/10.1016/j.apjr.2015.06.009
Bohnensack R & Halangk W (1986). Control of respiration and of motility in ejaculated bull spermatozoa. Biochimica et Biophysica Acta (BBA)-Bioenergetics 850(1): 72-79. doi.org/10.1016/0005-2728(86)90010-1
Bucci D, Rodriguez-Gil J E, Vallorani C, Spinaci M, Galeati G & Tamanini C (2011). GLUTs and mammalian sperm metabolism. Journal of Andrology 32(4): 348-355. doi.org/10.2164/jandrol.110.011197
Canovas S, Ross P J, Kelsey G & Coy P (2017). DNA methylation in embryo development: epigenetic impact of ART (assisted reproductive technologies). Bioessays 39(11): 1700106. doi.org/10.1002/bies.201700106
de Souza-Fabjan J M G, Panneau B, Duffard N, Locatelli Y, de Figueiredo J R, de Figueirêdo Freitas V J & Mermillod P (2014). In vitro production of small ruminant embryos: late improvements and further research. Theriogenology 81(9): 1149-1162. doi.org/10.1016/j.theriogenology.2014.02.001
El-Shahat K H, Taysser M I, Badr M R & Zaki K A (2016). Effect of heparin, caffeine and calcium ionophore A23187 on in vitro induction of the acrosome reaction of fresh ram spermatozoa. Asian Pacific Journal of Reproduction 5(2): 148-155. doi.org/10.1016/j.apjr.2016.01.012
Eskandari F & Momeni H R (2016). Silymarin protects plasma membrane and acrosome integrity in sperm treated with sodium arsenite. International Journal of Reproductive BioMedicine 14(1): 47. doi.org/10.29252/ijrm.14.1.47
Ferré L B, Bogliotti Y, Chitwood J L, Fresno C, Ortega H H, Kjelland M E & Ross P J (2017). Effect of spermatozoa motility hyperactivation factors and gamete coincubation duration on in vitro bovine embryo development using flow cytometrically sorted spermatozoa. Reproduction Fertility and Development 29(4): 805-814. doi.org/10.1071/rd15289
Gannon J R, Emery B R, Jenkins T G & Carrell D T (2014). The sperm epigenome: implications for the embryo. Genetic Damage in Human Spermatozoa 53-66. doi.org/10.1007/978-1-4614-7783-9_4
García-Álvarez O, Maroto-Morales A, Jiménez-Rabadán P, Ramón M, Del Olmo E, Iniesta-Cuerda M, Anel-Lopez L, Fernandez-Santos M R, Garde J J & Soler A J (2015). Effect of different media additives on capacitation of frozen–thawed ram spermatozoa as a potential replacement for estrous sheep serum. Theriogenology 84(6): 948-955. doi.org/10.1016/j.theriogenology.2015.05.032
Garrett L J, Revell S G & Leese H J (2008). Adenosine triphosphate production by bovine spermatozoa and its relationship to semen fertilizing ability. Journal of andrology 29(4): 449-458. doi.org/10.2164/jandrol.107.003533
Goodson S G, Qiu Y, Sutton K A, Xie G, Jia W & O’Brien D A (2012). Metabolic substrates exhibit differential effects on functional parameters of mouse sperm capacitation. Biology of Reproduction 87(3): 75. doi.org/10.1095/biolreprod.112.102673
Gu Y H, Li Y, Huang X F, Zheng J F, Yang J, Diao H, Yuan Y, Xu Y, Liu M, Shi HJ & Xu W P (2013). Reproductive effects of two neonicotinoid insecticides on mouse sperm function and early embryonic development in vitro. Plos one 8(7): e70112. doi.org/10.1371/journal.pone.0070112
Gross N, Strillacci M G, Peñagaricano F & Khatib H (2019). Characterization and functional roles of paternal RNAs in 2–4 cell bovine embryos. Scientific reports 9(1): 1-9. doi.org/10.1038/s41598-019-55868-3
Hajihassani A, Ahmadi E, Shirazi A & Shams-Esfandabadi N (2019). Reduced glutathione in the freezing extender improves the in vitro fertility of ram epididymal spermatozoa. Small Ruminant Research 174: 13-18. doi.org/10.1016/j.smallrumres.2019.02.016
Hidalgo D M, Romarowski A, Gervasi M G, Navarrete F, Balbach M, Salicioni A M, Lonny L R, Buck J & Visconti P E (2020). Capacitation increases glucose consumption in murine sperm. Molecular Reproduction and Development 87(10): 1037-1047. doi.org/10.1002/mrd.23421
Jin S K & Yang W X (2017). Factors and pathways involved in capacitation: how are they regulated. Oncotarget 8(2): 3600-3627. doi.org/10.18632/oncotarget.12274
Kashir J, Deguchi R, Jones C, Coward K & Stricker S A (2013). Comparative biology of sperm factors and fertilization-induced calcium signals across the animal kingdom. Molecular reproduction and development 80(10): 787-815. doi.org/10.1002/mrd.22222
Küçük N, Lopes JS, Soriano-Úbeda C, Hidalgo CO, Romar R & Gadea J (2020). Effect of oviductal fluid on bull sperm functionality and fertility under non-capacitating and capacitating incubation conditions. Theriogenology 158: 406-415. doi.org/10.1016/j.theriogenology.2020.09.035
Leahy T, Rickard JP, Aitken RJ & de Graaf SP (2016). Penicillamine prevents ram sperm agglutination in media that support capacitation. Reproduction 151(2): 167-177. doi.org/10.1530/rep-15-0413
Ledda S & Gonzalez-Bulnes A (2018). ET-technologies in small ruminants. In Animal Biotechnology 1. Springer. pp. 135-166. doi.org/10.1007/978-3-319-92327-7_6
Ledda S, Kelly J M, Nieddu S, Bebbere D, Ariu F, Bogliolo L & Arav A (2019). High in vitro survival rate of sheep in vitro produced blastocysts vitrified with a new method and device. Journal of animal science and biotechnology 10(1): 1-10. doi.org/10.1186/s40104-019-0390-1
Li F, Pi W H, Zhu H Z, Zhang S S, Liu S R & Xue J L (2006). The effect of estrous ewe serum and heparin on in vitro fertilization and subsequent embryonic development in sheep. Small Ruminant Research 63(3): 226-232. doi.org/10.1016/j.smallrumres.2005.02.019
Lone FA, Islam R, Khan MZ & Sofi KA (2011). Effect of transportation temperature on the quality of caudaepididymal spermatozoa of ram. Anim Reprod Sci 123(1-2):54-59. doi.org/10.1016/j.anireprosci.2010.10.012
Merati Z & Farshad A (2020). Ginger and echinacea extracts improve the quality and fertility potential of frozen-thawed ram epididymal spermatozoa. Cryobiology 92: 138-145. doi.org/10.1016/j.cryobiol.2019.12.003
Navarrete FA, Aguila L, Martin-Hidalgo D, Tourzani DA, Luque GM, Ardestani G, Garcia-Vazquez FA, Levin LR, Buck J, Darszon A, Buffone MG, Mager J, Fissore RA, Salicioni AM, Gervasi MG & Visconti PE (2019). Transient Sperm Starvation Improves the Outcome of Assisted Reproductive Technologies. Front Cell Dev Biol 7: 262. doi.org/10.3389/fcell.2019.00262
Odet F, Gabel S, London RE, Goldberg E & Eddy EM (2013). Glycolysis and mitochondrial respiration in mouse LDHC-null sperm. Biol Reprod 88(4): 95. doi.org/10.1095/biolreprod.113.108530
Otasevic V, Stancic A, Korac A, Jankovic A & Korac B (2020). Reactive oxygen, nitrogen, and sulfur species in human male fertility. A crossroad of cellular signaling and pathology. Biofactors 46(2): 206-219 doi.org/10.1002/biof.1535
Qiu J H, Li Y W, Xie H L, Li Q, Dong H B, Sun M J, Wei-Qiang Gao & Tan J H (2016). Effects of glucose metabolism pathways on sperm motility and oxidative status during long-term liquid storage of goat semen. Theriogenology 86(3): 839-849 doi.org/10.1016/j.theriogenology.2016.03.005
Roldan E R S (2019). Sperm competition and the evolution of sperm form and function in mammals. Reprod Domest Anim 54 Suppl 4: 14-21 doi.org/10.1111/rda.13552
Sajeevadathan M (2018). ROLE OF NA/K-ATPase IN BULL SPERM CAPACITATION (Doctoral dissertation, University of Saskatchewan). https://harvest.usask.ca/handle/10388/8639
Saleh A, Kashir J, Thanassoulas A, Safieh-Garabedian B, Lai F A & Nomikos M (2020). Essential role of sperm-specific PLC-zeta in egg activation and male factor infertility: an update. Front Cell Dev Biol 8: 28. doi.org/10.3389/fcell.2020.00028
Sánchez-Cárdenas C, Romarowski A, Orta G, De la Vega-Beltrán JL, Martín-Hidalgo D, Hernández-Cruz A, Visconti PE & Darszon A (2021). Starvation induces an increase in intracellular calcium and potentiates the progesterone-induced mouse sperm acrosome reaction. FASEB J 35(4): e21528. doi.org/10.1096/fj.202100122r
Sansegundo E, Tourmente M & Roldan E R S (2022). Energy Metabolism and Hyperactivation of Spermatozoa from Three Mouse Species under Capacitating Conditions. Cells 11(2): 220. doi.org/10.3390/cells11020220
Souza-Fabjan J M, Batista R I, Correia L F, Paramio M T, Fonseca J F, Freitas V J & Mermillod P (2021). In vitro production of small ruminant embryos: Latest improvements and further research. Reproduction, Fertility and Development 33(2): 31-54. doi.org/10.1071/rd20206
Sutradhar B C, Park J, Hong G, Choi S H & Kim G (2010). Effects of trypsinization on viability of equine chondrocytes in cell culture. Pak Vet J 30(4): 232-238.
Terrell K A, Wildt D E, Anthony N M, Bavister B D, Leibo S P, Penfold L M, Marker L L & Crosier A E (2011). Oxidative phosphorylation is essential for felid sperm function, but is substantially lower in cheetah (Acinonyx jubatus) compared to domestic cat (Felis catus) ejaculate. Biol Reprod 85(3): 473-481. doi.org/10.1095/biolreprod.111.092106
Tourmente M, Varea-Sánchez M, Roldan E R S (2019). Faster and more efficient swimming: energy consumption of murine spermatozoa under sperm competition. Biol Reprod 100(2): 420-428. doi.org/10.1093/biolre/ioy197
Tourmente M, Villar-Moya P, Varea-Sánchez M, Luque-Larena J J, Rial E, Roldan E R (2015). Performance of Rodent Spermatozoa Over Time Is Enhanced by Increased ATP Concentrations: The Role of Sperm Competition. Biol Reprod 93(3): 64. doi.org/10.1095/biolreprod.114.127621
Toyoda Y & Yokoyama M (2016). The early history of the TYH medium for in vitro fertilization of mouse ova. Journal of Mammalian Ova Research 33(1): 3-10. doi.org/10.1274/032.033.0103
Umehara T, Kawai T, Goto M, Richards J S & Shimada M (2018). Creatine enhances the duration of sperm capacitation: a novel factor for improving in vitro fertilization with small numbers of sperm. Human Reproduction 33(6): 1117-1129. doi.org/10.1093/humrep/dey081
Vazquez-Levin M H & Verón G L (2020). Myo-inositol in health and disease: its impact on semen parameters and male fertility. Andrology 8(2): 277-298. doi.org/10.1111/andr.12718
Visconti P E, Krapf D, de la Vega-Beltran J L, Acevedo J J & Darszon A (2011). Ion channels, phosphorylation and mammalian sperm capacitation. Asian J Androl 13: 395-405. doi.org/10.1038/aja.2010.69
Wani N A, Wani G M, Khan M Z & Salahudin S (2000). Effect of oocyte harvesting technique on in vitro maturation and in vitro fertilization in sheep. Small Rum Res 36(1): 63-67. doi.org/10.1016/s0921-4488(99)00097-8
Zeng Y & Chen T (2019). DNA methylation reprogramming during mammalian development. Genes 10(4): 257. doi.org/10.3390/genes10040257
Zhao L L, Ru Y F, Liu M, Tang J N, Zheng J F, Wu B, Yi-hua Gu & Shi H J (2017). Reproductive effects of cadmium on sperm function and early embryonic development in vitro. PloS One 12(11): e0186727. doi.org/10.1371/journal.pone.0186727
Zheng W W, Song G, Wang Q L, Liu S W, Zhu X L, Deng S M, Zhong A T, Yu-Mei T & Ying T (2018). Sperm DNA damage has a negative effect on early embryonic development following in vitro fertilization. Asian journal of andrology 20(1): 75. doi.org/10.4103/aja.aja_19_17
Zhu J, Moawad A R, Wang C Y, Li H F, Ren J Y & Dai Y F (2018). Advances in in vitro production of sheep embryos. International Journal of Veterinary Science and Medicine 6: S15-S26. doi.org/10.1016/j.ijvsm.2018.02.003
Zhu Z, Kawai T, Umehara T, Hoque S M, Zeng W & Shimada M (2019). Negative effects of ROS generated during linear sperm motility on gene expression and ATP generation in boar sperm mitochondria. Free Radical Biology and Medicine 141: 159-171. doi.org/10.1016/j.freeradbiomed.2019.06.018