Production of bovine embryos by piezo-ICSI using capacitated spermatozoa selected by fluorescence-activated cell sorting (FACS-piezo-ICSI).

Autor: Castro M; Laboratory of Reproduction, Center of Reproductive Biotechnology (CEBIOR-BIOREN), Faculty of Medicine, Universidad de la Frontera, Temuco, Chile; Master of Science Program specializing in Biology of Reproduction, Universidad de La Frontera, Temuco, Chile., Aguila L; Laboratory of Reproduction, Center of Reproductive Biotechnology (CEBIOR-BIOREN), Faculty of Medicine, Universidad de la Frontera, Temuco, Chile., Arias ME; Laboratory of Reproduction, Center of Reproductive Biotechnology (CEBIOR-BIOREN), Faculty of Medicine, Universidad de la Frontera, Temuco, Chile; Department of Agricultural Production, Faculty of Agriculture and Environmental Sciences, Universidad de La Frontera, Temuco, Chile., Felmer R; Laboratory of Reproduction, Center of Reproductive Biotechnology (CEBIOR-BIOREN), Faculty of Medicine, Universidad de la Frontera, Temuco, Chile; Department of Agricultural Sciences and Natural Resources, Faculty of Agricultural and Environmental Sciences, Universidad de La Frontera, Temuco, Chile. Electronic address: ricardo.felmer@ufrontera.cl.
Jazyk: angličtina
Zdroj: Animal reproduction science [Anim Reprod Sci] 2024 Sep; Vol. 268, pp. 107560. Date of Electronic Publication: 2024 Jul 08.
DOI: 10.1016/j.anireprosci.2024.107560
Abstrakt: Intracytoplasmic sperm injection (ICSI) remains inefficient in cattle. One reason could lie in the injection of oocytes with sperm that have not undergone molecular changes associated with in vivo capacitation and fertilizing ability. This study aimed to enhance the efficiency of bovine intracytoplasmic sperm injection (piezo-ICSI) by employing fluorescent-activated cell sorting (FACS) to select the sperm population before injection based on capacitation markers. First, we evaluated the effects of incubating thawed sperm for 2 hours with different capacitating inductors: heparin, methyl-beta-cyclodextrin (MβCD), and dibutyryl cyclic AMP (dbcAMP), alone or in combinations in a basal capacitating (C) medium (Sp-TALP). Sperm capacitation and quality markers were evaluated by flow cytometry, revealing heparin as the most effective inducer of sperm capacitation changes. It, therefore, this treatment was chosen as the sperm pretreatment for FACS-piezo-ICSI. Two cell populations showing high capacitating levels (Heparin-HCL) and low capacitating levels (Heparin-LCL) of the markers associated with sperm capacitation i(Ca 2+ ) levels and acrosome integrity were selected by FACS and used for sperm injection. Pronuclear formation was significantly higher when ICSI was performed with Heparin-HCL sperm than with Heparin-LCL and the control group (Heparin unsorted) groups (50 %, 10 %, and 20 %, respectively). Furthermore, injecting Heparin-HCL sperm resulted in a higher blastocyst rate (22.5 %) than Heparin-LCL (10 %) and the control group (15.2 %). In conclusion, heparin treatment effectively induced changes associated with sperm capacitation. The combination of Heparin-HCL treatment and FACS enabled precise selection of capacitated sperm before ICSI, enhancing the efficiency of this technology in the bovine species.
Competing Interests: Declaration of Competing Interest The authors declare to have no conflicts of interest to disclose.
(Copyright © 2024 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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