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题名	灵长类动物精子超低温冷冻保存的研究
作者	李亚辉
学位类别	博士
答辩日期	2005-01
授予单位	中国科学院研究生院
授予地点	北京
导师	季维智
关键词	精子 食蟹猴 冷冻 灵长类动物 防冻剂
其他题名	Cryopreservation of Primate Animal Spermatozoa
中文摘要	精子的超低温冷冻保存为珍稀、濒危动物种质资源的保存、人类不育症的治疗、家畜繁殖以及辅助生殖技术（如人工授精、体外受精和胚胎移植）的进一步发展奠定了基础。灵长类动物精子的冷冻保存距今已有30多年的历史，到目前为止，全世界两百余种灵长类动物中仅有约15个物种的精子被冷冻保存的报道，而用冷冻精液人工授精产下后代的报道仅见于大猩猩、黑猩猩、食蟹猴、械猴和称猴等5种灵长类动物。冻精人工授精率如此之低主要是因为人们对灵长类动物精子的低温生物学知识、精子的冷冻损伤机制以及某些物质的冷冻保护作用机理缺乏了解，导致了冷冻复苏精子的存活率降低，受精力急剧下降。为了探讨灵长类动物精子冷冻损伤的机理，提高精子的冷冻保存效率，本论文以食蟹猴和称猴为动物模型，以冷冻复苏精子的运动度、质膜完整率和顶体完整率为检测指标，对．影响灵长类动物精子冷冻保存效果的因素作了积极的探讨和研究：1）研究了脯氨酸、谷氨酞胺和甘氨酸对食蟹猴精子冷冻保存的影响，发现一定浓度的氨基酸有利于食蟹猴精子的冷冻保存，但高浓度的氨基酸却会对冷冻复苏精子产生负面影响；2）首次建立了灵长类动物精子成分确定防冻液的冷冻保存方法，为准确分析研究精子冷冻损伤机制和某些物质的冷冻保护机理莫定了基础；3）对比研究了不同类型冷冻保存稀释液及不同渗透性防冻剂在食蟹猴精子冷冻保存中的作用以及对冷冻复苏精子功能的影响，筛选、优化出了凡种适合于食蟹猴精子冷冻保存的稀释液和渗透性防冻剂；4）比较研究了几种不同类型防冻液对食蟹猴和称猴精子冷冻保存的效果，发现适合食蟹猴精子冻存的防冻液也同样适合于称猴精子。本研究的结果既有利于改进现有的灵长类动物精子冷冻保存方法，又有利于对精子的冷冻损伤机制和保护性物质的作用机理作进一步的研究，为低温生物学和生物多样性的保护作贡献。
英文摘要	animals, treating human infertility, and further developing reproductive techniques, such as artificial insemination(AI), in vitro fertilization(IVF) and embryo transplantation(ET). The first attempt to cryopreserve spermatozoa of nonhuman primate (NHP) was made more than 30 years ago, and up to now sperm of 15 primate species out of more than 200 NHP in the world have been frozen and only five species (i.e. gorillar, chimpanzee, cynomolgus monkey, marmoset and rhesus monkey) have been documented to produce offspring when frozen-thawed spermatozoa were artificially inseminated so far. As few knowledge has been obtained on sperm cryobiology of NHP and the mechanisms of sperm cryo-injury as well as cryoprotection by some compounds are lacking, post-thaw sperm survival and fertility decrease dramatically, which will in turn cause a low rate of artificial insemination in frozen-thawed spermatozoa. In order to probe into the mechanisms of sperm cryo-injury and improve the fertility of cryopreserved sperm, factors influencing the efficiency of sperm cryopreservation were studied using cynomolgus and rhesus monkey as experimental models. Sperm motility, plasma membrane and acrosomal integrity were examined to evaluate the cryopreserved sperm function. The present study included:!) Effect of amino acids on cryopreservation of cynomolgus monkey sperm; 2) Cryopreservation of cynomolgus monkey spermatozoa in a chemically defined extender; 3) Effects of various extenders and permeating cryoprotectants on cryopreservation of cynomolgus monkey spermatozoa; 4) Comparative studies with six extenders for sperm cryopreservation in cynomolgus monkey and rhesus monkey. The results showed that: 1) Adding a limited amount of amino acids to the extenders is beneficial for freezing cynomolgus monkey sperm but higher concentration of amino acid had negative effects on sperm freezing; 2) cynomolgus monkey spermatozoa can be successfully cryopreserved in a chemically defined extender. 5% glycerol and 30 min equilibration yielded the highest post-thaw sperm motility and head membrane integrity; 3) TTE, DM, mDM, LG-DM, G-DM and TCG are excellent extenders and suitable for cynomolgus monkey sperm cryopreservation. The mechanism of action of permeating cryoprotectants are not affected by extender composition. Ethylene glycol has a similar cryoprotective efficacy to glycerol; 4) TTE, DM, mDM, LG-DM, G-DM and TCG are equally suitable for cryopreservation of spermatozoa from cynomolgus and rhesus monkeys. These results will be beneficial for increasing the efficiency of sperm cryopreservation in NHP by improving sperm freezing procedures. Additionally, the results will also be helpful for researchers to understand and study the mechanisms of sperm cryo-injury as well as cryoprotection given by some compounds. This work will contribute some information to the area of cryobiology and bio-diversity preservation.
语种	中文
公开日期	2010-10-15
内容类型	学位论文
源URL	[http://159.226.149.42:8088/handle/152453/6194]
专题	昆明动物研究所_生殖与发育生物学
推荐引用方式 GB/T 7714	李亚辉. 灵长类动物精子超低温冷冻保存的研究[D]. 北京. 中国科学院研究生院. 2005.

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