Calcium ionophore A23187 versus progesterone: induction of acrosome reaction in thawed and prepared cryopreserved semen samples
DOI:
https://doi.org/10.18203/2320-1770.ijrcog20254268Keywords:
Acrosome reaction, Aqueous progesterone, Calcium ionophore, Cryopreservation, Fertilization failure, Induced acrosome reactionAbstract
Background: Acrosome reaction facilitates fertilization and it can be induced in-vitro using various stimuli like progesterone, calcium ionophore A23187, prostaglandins, etc. We investigated the efficacy of calcium ionophore and progesterone in inducing acrosome reaction in prepared semen samples following cryopreservation, using fluorescein isothiocyanate- labelled Arachis hypogaea (peanut) agglutinin (FITC-PNA) as a marker.
Methods: A prospective analysis was carried out on 10 normozoospermic semen samples collected between March and May 2025 at the department of reproductive medicine and andrology, Chettinad Super Speciality Hospital, Chettinad Academy of Research and Education. The collected semen samples were cryopreserved using rapid freezing method and subsequently after thawing, sperm preparation was carried out using direct swim-up method. The obtained post-wash was then subjected to capacitation and in-vitro induction of acrosome reaction using calcium ionophore A23187 and progesterone. The spermatozoa were stained with a fluorescent dye FITC-PNA and by examining under a fluorescent microscope, acrosome reacted and acrosome intact spermatozoa were scored. An independent t-test was performed to determine the statistical significance between the two groups. The statistical significance was set at p value <0.05.
Results: Our study revealed a significant difference between calcium ionophore and progesterone in inducing acrosome reaction in frozen thawed spermatozoa. Calcium ionophore demonstrated a higher acrosome reaction rate when compared to progesterone (p value <0.001).
Conclusions: The in-vitro induced acrosome reaction rate in frozen thawed spermatozoa is significantly higher with calcium ionophore when compared to progesterone.
Metrics
References
Esteves S, Verza S, Sharma R, Gosálvez J, Agarwal A. Role and significance of sperm function in men with unexplained infertility. In: Schattman G, Esteves S, Agarwal A, eds. Unexplained Infertility. Springer, New York, NY; 2015:91-119. DOI: https://doi.org/10.1007/978-1-4939-2140-9_11
Prajapati P, Kane S, McBrinn RC, Dean MS, Martins da Silva SJ, et al. Elevated and sustained intracellular calcium signalling is necessary for efficacious induction of the human sperm acrosome reaction. Int J Mol Sci. 2022;23(19):11253. DOI: https://doi.org/10.3390/ijms231911253
Brucker C, Lipford GB. The human sperm acrosome reaction: physiology and regulatory mechanisms. An update. Hum Reprod Update. 1995;1(1):51-62. DOI: https://doi.org/10.1093/humupd/1.1.51
Esteves SC, Verza Jr S. Relationship of in vitro acrosome reaction to sperm function: an update. Open Reprod Sci J. 2011;3(1):72-84. DOI: https://doi.org/10.2174/1874255601103010072
Krausz C, Bonaccorsi L, Luconi M, Fuzzi B, Criscuoli L, Pellegrini S, et al. Intracellular calcium increase and acrosome reaction in response to progesterone in human spermatozoa are correlated with in-vitro fertilization. Hum Reprod. 1995;10(1):120-4. DOI: https://doi.org/10.1093/humrep/10.1.120
Katsuki T, Hara T, Ueda K, Tanaka J, Ohama K. Prediction of outcomes of assisted reproduction treatment using the calcium ionophore-induced acrosome reaction. Hum Reprod. 2005;20(2):469-75. DOI: https://doi.org/10.1093/humrep/deh636
Pereira RJ, Tuli RK, Wallenhorst S, Holtz W. The effect of heparin, caffeine and calcium ionophore A 23187 on in vitro induction of the acrosome reaction in frozen-thawed bovine and caprine spermatozoa. Theriogenology. 2000;54(2):185-92. DOI: https://doi.org/10.1016/S0093-691X(00)00340-X
Tanphaichitr N, Hansen C. Production of motile acrosome‐reacted mouse sperm with nanomolar concentration of calcium ionophore A23187. Mol Reprod Dev. 1994;37(3):326-34. DOI: https://doi.org/10.1002/mrd.1080370312
Foresta C, Rossato M, Di Virgilio F. Ion fluxes through the progesterone-activated channel of the sperm plasma membrane. Biochem J. 1993;294(1):279-83. DOI: https://doi.org/10.1042/bj2940279
Harper CV, Barratt CL, Publicover SJ, Kirkman-Brown JC. Kinetics of the progesterone-induced acrosome reaction and its relation to intracellular calcium responses in individual human spermatozoa. Biol Reprod. 2006;75(6):933-9. DOI: https://doi.org/10.1095/biolreprod.106.054627
Sagare‐Patil V, Galvankar M, Satiya M, Bhandari B, Gupta SK, Modi D. Differential concentration and time dependent effects of progesterone on kinase activity, hyperactivation and acrosome reaction in human spermatozoa. Int J Androl. 2012;35(5):633-44. DOI: https://doi.org/10.1111/j.1365-2605.2012.01291.x
Oren-Benaroya R, Orvieto R, Gakamsky A, Pinchasov M, Eisenbach M. The sperm chemoattractant secreted from human cumulus cells is progesterone. Hum Reprod. 2008;23(10):2339-45. DOI: https://doi.org/10.1093/humrep/den265
Jaiswal BS, Eisenbach M, Tur-Kaspa I. Detection of partial and complete acrosome reaction in human spermatozoa: which inducers and probes to use? Mol Hum Reprod. 1999;5(3):214-9. DOI: https://doi.org/10.1093/molehr/5.3.214
Santo MD, Tarozzi N, Nadalini M, Borini A. Human sperm cryopreservation: update on techniques, Effect on DNA integrity, and implications for ART. Adv Urol. 2011;2012:854837. DOI: https://doi.org/10.1155/2012/854837
Practice Committees of the American Society for Reproductive Medicine and Society for Assisted Reproductive Technology. Intracytoplasmic sperm injection (ICSI) for non-male factor infertility: a committee opinion. Fertil Steril. 2012;98(6):1395-9. DOI: https://doi.org/10.1016/j.fertnstert.2012.08.026
Anger JT, Gilbert BR, Goldstein M. Cryopreservation of sperm: indications, methods and results. J Urol. 2003;170(4 Pt 1):1079-84. DOI: https://doi.org/10.1097/01.ju.0000084820.98430.b8
Mahadevan MM, Trounson AO. Relationship of fine structure of sperm head to fertility of frozen human semen. Fertil Steril. 1984;41(2):287-93. DOI: https://doi.org/10.1016/S0015-0282(16)47606-5
Esteves SC, Sharma RK, Thomas Jr AJ, Agarwal A. Improvement in motion characteristics and acrosome status in cryopreserved human spermatozoa by swim-up processing before freezing. Hum Reprod Oxf Engl. 2000;15(10):2173-9. DOI: https://doi.org/10.1093/humrep/15.10.2173
Critser JK, Huse-Benda AR, Aaker DV, Arneson BW, Ball GD. Cryopreservation of human spermatozoa. I. Effects of holding procedure and seeding on motility, fertilizability, and acrosome reaction. Fertil Steril. 1987;47(4):656-63. DOI: https://doi.org/10.1016/S0015-0282(16)59118-3
Henkel R, Müller C, Miska W, Gips H, Schill WB. Determination of the acrosome reaction in human spermatozoa is predictive of fertilization in vitro. Hum Reprod. 1993;8(12):2128-32. DOI: https://doi.org/10.1093/oxfordjournals.humrep.a137994
Agarwal A, Mulgund A, Hamada A, Chyatte MR. A unique view on male infertility around the globe. Reprod Biol Endocrinol. 2015;13:37. DOI: https://doi.org/10.1186/s12958-015-0032-1
Calvo L, Dennison-Lagos L, Banks SM, Sherins RJ. Characterization and frequency distribution of sperm acrosome reaction among normal and infertile men. Hum Reprod. 1994;9(10):1875-9. DOI: https://doi.org/10.1093/oxfordjournals.humrep.a138351
Esteves SC, Schneider DT. Male infertility and assisted reproductive technology: lessons from the IVF. Open Reprod Sci J. 2011;3(1):138-53. DOI: https://doi.org/10.2174/1874255601103010138
Balli M, Cecchele A, Pisaturo V, Makieva S, Carullo G, Somigliana E, et al. Opportunities and limits of conventional IVF versus ICSI: it is time to come off the fence. J Clin Med. 2022;11(19):5722. DOI: https://doi.org/10.3390/jcm11195722
Tavalaee M, Kiani-Esfahani A, Nasr-Esfahani MH. Relationship between potential sperm factors involved in oocyte activation and sperm DNA fragmentation with intra-cytoplasmic sperm injection clinical outcomes. Cell J. 2017;18(4):588-96. DOI: https://doi.org/10.1111/andr.12179
Popkiss S, Horta F, Vollenhoven B, Green MP, Zander-Fox D. Calcium chloride dihydrate supplementation at ICSI improves fertilization and pregnancy rates in patients with previous low fertilization: a retrospective paired treatment cycle study. J Assist Reprod Genet. 2022;39(5):1055-64. DOI: https://doi.org/10.1007/s10815-022-02407-1
Cross NL, Hanks SE. Effects of cryopreservation on human sperm acrosomes. Hum Reprod. 1991;6(9):1279-83. DOI: https://doi.org/10.1093/oxfordjournals.humrep.a137526
Chang MC. The meaning of sperm capacitation. A historical perspective. J Androl. 1984;5(2):45-50. DOI: https://doi.org/10.1002/j.1939-4640.1984.tb00775.x
Larson JL, Miller DJ. Simple histochemical stain for acrosomes on sperm from several species. Mol Reprod Dev. 1999;52(4):445-9. DOI: https://doi.org/10.1002/(SICI)1098-2795(199904)52:4<445::AID-MRD14>3.0.CO;2-6
Zeginiadou T, Papadimas J, Mantalenakis S. Acrosome reaction: methods for detection and clinical significance. Andrologia. 2000;32(6):335-43. DOI: https://doi.org/10.1046/j.1439-0272.2000.00359.x
Talbot P, Summers RG, Hylander BL, Keough EM, Franklin LE. The role of calcium in the acrosome reaction: an analysis using ionophore A23187. J Exp Zool. 1976;198(3):383-92. DOI: https://doi.org/10.1002/jez.1401980312
Contreras HR, Llanos MN. Detection of progesterone receptors in human spermatozoa and their correlation with morphological and functional properties. Int J Androl. 2001;24(4):246-52. DOI: https://doi.org/10.1046/j.1365-2605.2001.00294.x
Mbizvo MT, Thomas S, Fulgham DL, Alexander NJ. Serum hormone levels affect sperm function. Fertil Steril. 1990;54(1):113-20. DOI: https://doi.org/10.1016/S0015-0282(16)53646-2
Meizel S, Turner KO, Nuccitelli R. Progesterone triggers a wave of increased free calcium during the human sperm acrosome reaction. Dev Biol. 1997;182(1):67-75. DOI: https://doi.org/10.1006/dbio.1997.8477
World Health Organization. WHO laboratory manual for the examination and processing of human semen. 6th edn. Geneva: World Health Organization; 2021.
Merchant R, Gandhi G, Allahbadia GN. In vitro fertilization/intracytoplasmic sperm injection for male infertility. Indian J Urol. 2011;27(1):121-32. DOI: https://doi.org/10.4103/0970-1591.78430
Downloads
Published
How to Cite
Issue
Section
