Live rats resulting from injection of oocytes with spermatozoa freeze-dried and stored for one year
Live rats resulting from injection of oocytes with spermatozoa freeze-dried and stored for one year
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DOI：
10.1002/mrd.20825
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发表时间：
2008-05-01
影响因子：
2.5
通讯作者：
Hirabayashi, Masumi
中科院分区：
文献类型：
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作者：
Hochi, Shinichi;Watanabe, Kaori;Hirabayashi, Masumi
本研究旨在检测冷冻干燥和保存1年后的大鼠精子是否可以参与卵胞浆内单精子注射(ICSI)后的完全发育。雄性Wistar大鼠附睾尾精子在液氮中冷冻，先在0.37hPa下干燥14h，然后在0.001 hPa下干燥3h。干燥的精子在+25摄氏度的干燥器中保存1年，或在+4摄氏度的冰箱中保存1年，或在-196摄氏度的液氮中保存1年。对照组包括只冷冻并保存在液氮中的精子。精子保存后，超声分离精子尾部，注入超排SLC：SD大鼠卵母细胞。冷冻精子和冷冻干燥精子在+25℃、+4℃和-196℃保存的受精率分别为79%、75%、70%和73%。而注射后的卵母细胞的卵裂率分别为63%、1%、38%和36%。在将每组80个受精卵移植到受体体内后，冷冻精子和冷冻干燥精子在+25℃、+4℃和-196℃保存的完全正常后代的百分比分别为36%、0%、7%和14%。这些结果表明，保存温度对注射冷冻干燥精子的卵母细胞产生的大鼠足月发育的可能性有显著影响。对ICSI卵母细胞中大鼠精子的染色体分析表明，+25℃冷冻干燥精子的染色体畸变率(100%)高于冷冻对照精子(41%)和-196℃冷冻干燥精子(35%)，其中+4℃冷冻干燥精子染色体畸变率居中(65%)。综上所述，冷冻干燥并在+4℃保存1年的大鼠精子能够参与ICSI后的完全发育。
This study was designed to examine whether rat spermatozoa after freeze-drying and 1-year storage can participate in full-term development following intracytoplasmic sperm injection (ICSI). Cauda epididymal spermatozoa from Crlj:Wistar rats were frozen in liquid nitrogen (LN2), first dried for 14 hr at 0.37 hPa and then for 3 hr at 0.001 hPa. The dried spermatozoa were stored for 1 year in a desiccator at +25 degrees C, or in a refrigerator at +4 degrees C, or in LN2 at -196 degrees C. Controls consisted of sperm that had only been frozen and stored in LN2. After being stored, spermatozoa were sonicated to dissociate the sperm tail and were injected into oocytes from superovulated Slc:SD rats. The respective fertilization rates of oocytes injected with frozen sperm, or with freeze-dried sperm stored at +25, +4, and -196 degrees C were 79%, 75%, 70%, and 73%. However, the corresponding cleavage rates of injected oocytes were 63%, 1%, 38%, and 36%. After transfer of >80 zygotes of each group into recipients, the respective percentages of full-term normal offspring resulting from frozen sperm or from freeze-dried sperm stored at +25, +4, and -196 degrees C were 36%, 0%, 7%, and 14%. These results demonstrate that the storage temperature significantly influenced the likelihood of term development of rats produced by injection of oocytes with freeze-dried spermatozoa. Chromosomal analysis of the rat spermatozoa in the ICSI oocytes indicated that chromosomal aberration in freeze-dried spermatozoa stored at +25 degrees C (100%) occurred more frequently than in frozen control spermatozoa (41%) and freeze-dried spermatozoa stored at -196 degrees C (35%), and the frequency of chromosomal aberrations in freeze-dried spermatozoa stored at +4 degrees C (65%) was the intermediate. In conclusion, rat spermatozoa freeze-dried and stored at +4 degrees C for 1 year are capable of participating in full-term development after ICSI.