Heparin is the most commonly used in vitro capacitation inducer in the bovine. However, hyaluronic acid (HA) has been recently used for capacitation induction as well as for other reproductive biotechnologies, such as sperm selection and in vitro fertilization (IVF). Our aim was to induce sperm capacitation with heparin or HA in order to study mAC and TK intracellular signals and their relation with cleavage and blastocyst rates after IVF as well as with the oxidative status of the potential bovine embryos. 2,5-dideoxyadenosine and genistein were used as mAC and TK inhibitors, respectively. Sperm capacitation was analyzed using CTC technique, sperm plasma membrane and acrosome integrity were determined using trypan blue stain and differential interference contrast, and mitochondrial activity was evaluated using fluorochrome JC-1. Cleavage rate was analyzed 48h and blastocyst production 7-8 days after IVF, while cytosolic oxidative activity was determined using RedoxSensor Red CC-1 fluorochrome 7h after IVF. When mAC and TK inhibitors were added to sperm samples, only capacitation decreased significantly both in HA and heparin treated samples (P < 0.05), but plasma membrane and acrosome integrity percentages were not affected in any of these groups (P > 0.05). Sperm mitochondrial membrane potential only decreased in heparin treated samples in the presence of both inhibitors (P < 0.05). Oocytes activated with HA sperm treated samples with the addition of 2,5-dideoxyadenosine and genistein presented a lower cytosolic oxidative status than those activated with sperm treated with HA alone (P < 0.05). On the other hand, oocytes activated with heparin treated sperm samples presented a lower cytosolic oxidative status only in the presence of 2,5-dideoxyadenosine (P < 0.05). Therefore, mAC and TK present a differential participation in heparin and HA sperm induced capacitation and mitochondrial function as well as in IVF.

中文翻译：

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透明质酸获能诱导细胞内信号由参与牛体外受精的膜相关腺苷酸环化酶和酪氨酸激酶调节

肝素是牛最常用的体外获能诱导剂。然而，透明质酸 (HA) 最近已用于诱导获能以及其他生殖生物技术，例如精子选择和体外受精 (IVF)。我们的目的是用肝素或 HA 诱导精子获能，以研究 mAC 和 TK 细胞内信号及其与 IVF 后卵裂和囊胚率以及潜在牛胚胎氧化状态的关系。2,5-二脱氧腺苷和染料木黄酮分别用作mAC和TK抑制剂。使用 CTC 技术分析精子获能，使用台盼蓝染色和微分干涉对比确定精子质膜和顶体完整性，使用荧光染料 JC-1 评估线粒体活性。在试管婴儿后 48 小时分析卵裂率和在试管婴儿后 7-8 天分析胚泡的产生，而在试管婴儿后 7 小时使用 RedoxSensor Red CC-1 荧光染料测定细胞溶质氧化活性。当在精子样本中加入 mAC 和 TK 抑制剂时，只有 HA 和肝素处理的样本中的获能显着降低（P < 0.05），但质膜和顶体完整性百分比在任何这些组中均未受到影响（P > 0.05）。在两种抑制剂存在的情况下，肝素处理的样品中精子线粒体膜电位仅降低（P < 0.05）。用添加了 2,5-双脱氧腺苷和染料木黄酮的 HA 精子处理的样品激活的卵母细胞呈现出比单独用 HA 处理的精子激活的卵母细胞更低的细胞溶质氧化状态（P < 0.05）。另一方面，仅在 2,5-二脱氧腺苷存在的情况下，用肝素处理的精子样本激活的卵母细胞呈现较低的细胞溶质氧化状态（P < 0.05）。因此，mAC 和 TK 在肝素和 HA 精子诱导的获能和线粒体功能以及 IVF 中表现出不同的参与。