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Effect of MnTBAP on in vitro capacitation of frozen-thawed stallion sperm.
Animal Reproduction Science ( IF 2.2 ) Pub Date : 2020-08-05 , DOI: 10.1016/j.anireprosci.2020.106570
Favián Treulen 1 , Luis Aguila 2 , María Elena Arias 3 , Ignacio Jofré 4 , Ricardo Felmer 5
Affiliation  

In vitro manipulation of spermatozoa leads to deleterious changes of structure and function that occur mainly due to oxidative stress, therefore, prevention or treatment is a strategy to improve the functions of processed sperm. In the present study, the aim was to evaluate the effects of MnTBAP supplementation, a compound with antioxidant activity, on in vitro capacitation conditions of thawed equine sperm. For this purpose, stallion spermatozoa (2 × 106 cells/mL) were incubated in the sperm-TLP base medium for 4 h in which there were three different conditions: non-capacitating, capacitating, and capacitating plus 150 mM MnTBAP. There were incubations for 4 h at 37.5 °C in a humidified air atmosphere. Sample analysis was performed immediately after thawing (0 h), and at the end of the incubation period (4 h), unless otherwise indicated. The following variables were evaluated for spermatozoa: plasma membrane integrity and fluidity, acrosome integrity, intracellular calcium concentrations, intracellular pH, tyrosine phosphorylation, ATP concentrations, motility and heterologous zona-binding assay, using flow cytometry, fluorescent microscopy and/or chemiluminescence, depending on the most appropriate procedure for the variable being evaluated. Results indicated that capacitation-like changes were synergistically induced by the cAMP agonists, phosphodiesterase inhibitor and bicarbonate. The presence of bovine serum albumin was harmful to the plasma membrane. The MnTBAP supplementation had a positive effect on viability-related markers (plasma membrane integrity, membrane fluidity, associated with greater intracellular pH) when there were capacitating conditions. In conclusion, the activity of MnTBAP contributes to improving the in vitro incubation conditions of frozen-thawed stallion sperm.



中文翻译:

MnTBAP对冻融种马精子体外获能的影响。

精子的体外操作导致主要由于氧化应激而发生的结构和功能的有害变化,因此,预防或治疗是改善已加工精子功能的策略。在本研究中,目的是评估补充 MnTBAP(一种具有抗氧化活性的化合物)对解冻马精子体外获能条件的影响。为此,种马精子 (2 × 10 6细胞/mL)在精子-TLP 基础培养基中培养 4 小时,其中存在三种不同的条件:非获能、获能和获能加 150 mM MnTBAP。在 37.5 °C 的加湿空气环境中孵育 4 小时。除非另有说明,否则在解冻后(0 小时)和孵育期结束时(4 小时)立即进行样品分析。评估了精子的以下变量:质膜完整性和流动性、顶体完整性、细胞内钙浓度、细胞内 pH 值、酪氨酸磷酸化、ATP 浓度、运动性和异源带结合测定,使用流式细胞术、荧光显微镜和/或化学发光,取决于评估变量的最合适程序。结果表明,cAMP 激动剂、磷酸二酯酶抑制剂和碳酸氢盐协同诱导了获能样变化。牛血清白蛋白的存在对质膜有害。当存在获能条件时,MnTBAP 补充剂对活力相关标志物(质膜完整性、膜流动性、与更高的细胞内 pH 值相关)具有积极影响。总之,MnTBAP 的活性有助于提高 与更大的细胞内 pH 值相关)。总之,MnTBAP 的活性有助于提高 与更大的细胞内 pH 值相关)。总之,MnTBAP 的活性有助于提高冻融种马精子的体外培养条件。

更新日期:2020-08-08
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