The effect of cult-active medium on pregnancy outcomes after intracytoplasmic sperm injection in azoospermic men: A case-control study
Background: Failed oocyte activation following intracytoplasmic sperm injection (ICSI) as a result of calcium deficiency is a major challenge.
Objective: We compared the effect of cult-active medium (CAM) on ICSI outcomes in obstructive azoospermia cases.
Materials and Methods: The present study was conducted with 152 ICSI cases, classified into CAM and control groups. The injected oocytes in the control group were cultured in the cleavage medium, while in the artificial oocyte activation group, oocytes were chemically activated through exposure to 200 µL of CAM for 15 min. Fertilization and cleavage rates, quality of embryos, and biochemical pregnancy and live birth rates were assessed in both groups.
Results: There were significant differences between the groups in terms of fertilization and cleavage rates after using the CAM in the percutaneous epididymal sperm aspiration (PESA) subgroup (p = 0.05, p ≤ 0.001) and in the testicular sperm extraction subgroup (p = 0.02, p = 0.04), compared to their control groups. Also, the pregnancy rate was significantly higher in the PESA-CAM subgroup (p = 0.03). The PESA-CAM subgroup demonstrated a significant difference in embryo quality after ICSI (p = 0.04). Unsuccessful embryo transfer and abortion were lower in both subgroups compared to the control groups, but this difference was not significant. Surprisingly, live birth rate was higher in the PESA-CAM subgroup (p = 0.03).
Conclusion: CAM treatment could improve fertilization and cleavage rates in obstructive azoospermia participants. It had a significant effect on embryo quality, and pregnancy and live birth rates in PESA cases.
Key words: Calcium ionophore, Obstructive azoospermia, Fertilization, ICSI.
 Borges Jr E, de Almeida Ferreira Braga DP, de Sousa Bonetti TC, Iaconelli Jr A, Franco Jr JG. Artificial oocyte activation with calcium ionophore A23187 in intracytoplasmic sperm injection cycles using surgically retrieved spermatozoa. Fertil Steril 2009; 92: 131–136.
 Verheyen G, Popovic-Todorovic B, Tournaye H. Processing and selection of surgically-retrieved sperm for ICSI: A review. Basic Clin Androl 2017; 27: 6.
 Bonte D, Ferrer-Buitrago M, Dhaenens L, Popovic M, Thys V, De Croo I, et al. Assisted oocyte activation significantly increases fertilization and pregnancy outcome in patients with low and total failed fertilization after intracytoplasmic sperm injection: A 17-year retrospective study. Fertil Steril 2019; 112: 266–274.
 Nomikos M, Yu Y, Elgmati Kh, Theodoridou M, Campbell K, Vassilakopoulou V, et al. Phospholipase C-zeta rescues failed oocyte activation in a prototype of male factor infertility. Fertil Steril 2013; 99: 76–85.
 Hachem A, Godwin J, Ruas M, Lee HC, Buitrago MF, Ardestani G, et al. PLC? is the physiological trigger of the Ca2+ oscillations that induce embryogenesis in mammals but conception can occur in its absence. Development 2017; 144: 2914–2924.
 Ferrer-Buitrago M, Dhaenens L, Lu Y, Bonte D, Vanden Meerschaut F, De Sutter P, et al. Human oocyte calcium analysis predicts the response to assisted oocyte activation in patients experiencing fertilization failure after ICSI. Hum Reprod 2018; 33: 416–425.
 Kashir J, Nomikos M, Swann K, Lai FA. PLC? or PAWP: Revisiting the putative mammalian sperm factor that triggers egg activation and embryogenesis. Mol Hum Reprod 2015; 21: 383–388.
 Karabulut S, Aksünger Ö, Ata C, Sagfiroglu Y, Keskin I. Artificial oocyte activation with calcium ionophore for frozen sperm cycles. Syst Biol Reprod Med 2018; 64: 381– 388.
 Li B, Zhou Y, Yan Zh, Li M, Xue S, Cai R, et al. Pregnancy and neonatal outcomes of artificial oocyte activation in patients undergoing frozen-thawed embryo transfer: A 6- year population-based retrospective study. Arch Gynecol Obstet 2019; 300: 1083–1092.
 Norozi-Hafshejani M, Tavalaee M, Azadi L, Bahadorani M, Nasr-Esfahani MH. Effects of assisted oocyte activation with calcium-ionophore and strontium chloride on in vitro ICSI outcomes. Iran J Basic Med Sci 2018; 21: 1109–1117.
 Ebner Th, Montag M. Live birth after artificial oocyte activation using a ready-to-use ionophore: A prospective multicentre study. Reprod Biomed Online 2015; 30: 359– 365.
 Kochhar PK, Ghosh P. Intracytoplasmic sperm injection with assisted oocyte activation resulting in successful pregnancies and live birth in couples with globozoospermia: A report of two cases. J Hum Reprod Sci 2018; 11: 72–74.
 Eldar-Geva T, Brooks B, Margalioth EJ, Zylber-Haran E, Gal M, Silber ShJ. Successful pregnancy and delivery after calcium ionophore oocyte activation in a normozoospermic patient with previous repeated failed fertilization after intracytoplasmic sperm injection. Fertil Steril 2003; 79 (Suppl.): 1656–1658.
 Hebisha SA, Aboelazm BA, Adel HM, Aboali AA, Ahmed AL. Effect of oocyte activation with calcium ionophore on ICSI outcomes in patients with previous fertilization failure. Fertil Steril 2016; 106: e186–e187.
 Ebner Th, Köster M, Shebl O, Moser M, Van der Ven H, Tews G, et al. Application of a ready-to-use calcium ionophore increases rates of fertilization and pregnancy in severe male factor infertility. Fertil Steril 2012; 98: 1432– 1437.
 Fesahat F, Montazeri F, Sheikhha MH, Saeedi H, Dehghani Firouzabadi R, Kalantar SM. Frequency of chromosomal aneuploidy in high quality embryos from young couples using preimplantation genetic screening. Int J Reprod Biomed 2017; 15: 297–304.
 Tavalaee M, Kiani-Esfahani A, Nasr-Esfahani MH. Relationship between phospholipase C-zeta, semen parameters, and chromatin status. Syst Biol Reprod Med 2017; 63: 259–268.
 Rahimizadeh P, Rezaei Topraggaleh T, Nasr-Esfahani MH, Ziarati N, Mirshahvaladi Sh, Esmaeili V, et al. The alteration of PLCζ protein expression in unexplained infertile and asthenoteratozoospermic patients: A potential effect on sperm fertilization ability. Mol Reprod Dev 2020; 87: 115– 123.
 Lu Q, Zhao Y, Gao X, Li Y, Ma S, Mullen S, et al. Combination of calcium ionophore A23187 with puromycin salvages human unfertilized oocytes after ICSI. Eur J Obstet Gynecol Reprod Biol 2006; 126: 72–76.
 Long R, Wang M, Yang QY, Hu SQ, Zhu LX, Jin L. Risk of birth defects in children conceived by artificial oocyte activation and intracytoplasmic sperm injection: A metaanalysis. Reprod Biol Endocrinol 2020; 18: 123.
 Kim ST, Cha YB, Park JM, Gye MC. Successful pregnancy and delivery from frozen-thawed embryos after intracytoplasmic sperm injection using roundheaded spermatozoa and assisted oocyte activation in a globozoospermic patient with mosaic Down syndrome. Fertil Steril 2001; 75: 445–447.
 Lv M, Zhang D, He X, Chen B, Li Q, Ding D, et al. Artificial oocyte activation to improve reproductive outcomes in couples with various causes of infertility: A retrospective cohort study. Reprod Biomed Online 2020; 40: 501–509.
 Nasr-Esfahani MH, Razavi Sh, Mardani M, Shirazi R, Javanmardi S. Effects of failed oocyte activation and sperm protamine deficiency on fertilization post-ICSI. Reprod Biomed Online 2007; 14: 422–429.
 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: 469–475.
 Vanden Meerschaut F, Nikiforaki D, Heindryckx B, De Sutter P. Assisted oocyte activation following ICSI fertilization failure. Reprod Biomed Online 2014; 28: 560– 571.
 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: 588–596.
 Aytac PC, Kilicdag EB, Haydardedeoglu B, Simsek E, Cok T, Parlakgumus HA. Can calcium ionophore “use” in patients with diminished ovarian reserve increase fertilization and pregnancy rates? A randomized, controlled study. Fertil Steril 2015; 104: 1168–1174.
 Yanagida K, Morozumi K, Katayose H, Hayashi S, Sato A. Successful pregnancy after ICSI with strontium oocyte activation in low rates of fertilization. Reprod Biomed Online 2006; 13: 801–806.
 Moaz MN, Khattab Sh, Aboul Foutouh I, Abdel Mohsen E. Chemical activation of oocytes in different types of sperm abnormalities in cases of low or failed fertilization after ICSI: A prospective pilot study. Reprod Biomed Online 2006; 13: 791–794.
 Murugesu S, Saso S, Jones BP, Bracewell-Milnes T, Athanasiou T, Mania A, et al. Does the use of calcium ionophore during artificial oocyte activation demonstrate an effect on pregnancy rate? A meta-analysis. Fertil Steril 2017; 108: 468–482.
 Niu X, Ruan Q, Witz CA, Wang W. Comparison of human oocyte activation between round-headed sperm injection followed by calcium ionophore treatment and normal sperm injection in a patient with globozoospermia. Front Endocrinol 2020; 11: 183.