Mechanism of mammalian sperm protection by extender components and development of novel extenders free of products of animal origin

加拿大农业部报告称，2013年，加拿大向100多个国家出口了价值超过1亿美元的公牛精液。这占全球公牛精液出口的20%以上。保存哺乳动物精子最常用的保护剂包括20%的卵黄或牛奶和6-7%的甘油。自从大约75年前引入人工授精以来，精子保存介质(通常称为稀释剂或扩充剂)的基本成分一直保持不变。蛋黄和牛奶是动物来源的产品，具有传播疾病的潜在风险。因此，迫切需要开发不含动物源性产品的新型填充剂。然而，由于缺乏关于卵黄和牛奶如何在液态和/或冷冻状态保存过程中保护精子的知识，新型延长剂的开发一直受到阻碍。因此，在过去的15年里，我们集中精力了解精子延长器在精子保存过程中保护精子的分子机制。我们发现大多数有蹄动物(公牛、种马、公羊、山羊、野牛等)的精液中含有有害的蛋白质，称为精子结合蛋白(BSP)。这些BSP由精囊分泌，在射精时添加到精子中，与精子膜结合并提取脂类(胆固醇和磷脂)，使精子在冷却和冷冻保存期间对储存敏感。有趣的是，BSP与低密度脂蛋白相互作用，低密度脂蛋白是蛋黄增强剂的主要成分。虽然这些BSP结合在精子膜上，但它们对精子产生有害影响，但当它们与低密度蛋白结合时，对精子的有害影响就会减弱，精子得到保护。在此基础上，我们得出结论：延长体组分对BSPs的隔离是精子保护的一般机制。近年来，我们还发现右旋糖苷与二乙氨基乙基偶联，与BSP结合，这种相互作用保留了精子的功能，就像卵黄一样。因此，本计划的短期目标是测试公牛精子在合成稀释液中的保存情况，以开发不含动物产品的新型稀释液用于人工授精。该计划的长期目标是开发用于哺乳动物精子保存(其他农场动物、家畜、动物园动物和濒危物种)的新型扩充剂。加拿大育种者将从获得安全的精液稀释器中受益，并可能在国际上声明这一主张，从而促进他们自己的牛群。该程序还将生成 高素质人员(HQP)，他们将在人工授精行业和/或生育诊所等高度专业化的领域获得工作机会。

Agriculture Canada reported that Canada exported more than 100 million dollars worth of bull semen to over 100 countries in 2013. This represents more than 20% of world export of bull semen. The most commonly used protective agents to conserve mammalian sperm contain 20% egg yolk or milk and 6-7% glycerol in buffers. This basic constituent of sperm preservation media (commonly called diluters or extenders) has remained the same since the practice of artificial insemination was introduced some 75 years ago. Egg yolk and milk being products of animal origin represent a potential risk of transmission of diseases. Therefore, there is an urgent need for developing novel extenders, which are free of products of animal origin. However, in view of the lack of knowledge on “how Egg yolk and milk protect sperm during conservation in liquid and/or frozen state”, the development of novel extenders has been hindered. Consequently, we focused our effort in the past 15 years to understand the molecular mechanism of sperm protection by extenders during conservation. We discovered that most hoofed animals (bull, stallion, ram, goat, bison, etc) semen contains detrimental proteins called Binder of Sperm (BSP) proteins. These BSPs, secreted by seminal vesicles and added to sperm at ejaculation, bind to sperm membrane and extract lipids (cholesterol and phospholipids), which make sperm sensitive to storage during cooling and cryopreservation. Interestingly, BSPs interact with low-density lipoproteins, the main constituents of egg yolk extenders. While these BSPs bind to sperm membrane, they exert a detrimental effect on sperm but when they associate with low-density proteins, the detrimental effect on sperm is subdued and sperm are protected. Based on the above, we concluded that the sequestration of BSPs by extender components is the general mechanism of sperm protection. In recent years, we also discovered that dextrans coupled with diethyl aminoethyl groups, bind BSPs, and this interaction preserves sperm functions just like egg yolk. Thus, the short-term objectives of the present program are to test bull sperm conservation in synthetic extenders, in order to develop novel extenders free of animal products for artificial insemination application. The long-term goal of this program is to develop novel extenders for mammalian sperm conservation (other farm animals, domestic animals, zoo animals, and endangered species). Canadian breeders would benefit from having access to safe semen extenders and might state this claim internationally, thus promoting their own herd. This program would also generate highly qualified personnel (HQPs), who would have the job opportunity in highly specialized areas such as Artificial Insemination Industries and/or Fertility Clinics.