Importance of primary culture conditions for the development of rat ICSI embryos and long-term preservation of freeze-dried sperm
Importance of primary culture conditions for the development of rat ICSI embryos and long-term preservation of freeze-dried sperm
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DOI：
10.1016/j.cryobiol.2009.02.004
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发表时间：
2009-06-01
期刊：
影响因子：
2.7
通讯作者：
Nakagata, Naomi
中科院分区：
文献类型：
--
作者：
Kaneko, Takehito;Kimura, Shinya;Nakagata, Naomi
在含有Tris和乙二胺四乙酸（EDTA） （TE缓冲液）的溶液中冷冻干燥的大鼠精子可以在4℃下保存，注射这些精子的卵母细胞虽然发育能力较低，但仍能发育成后代。研究了冻干精子注射后提高卵母细胞发育能力的培养条件。将受精卵注入新鲜精子后，分别在改性的克雷布-林格碳酸氢盐（mKRB）、改性的大鼠1细胞胚胎培养基(mR1ECM)/BSA和不同渗透压的mR1ECM中培养，然后在mR1ECM中培养。与仅用mR1ECM培养的受精卵相比，在mR1ECM培养前用mKRB培养的受精卵（270 mOsm）成囊胚的比例较高（50%比28%，P < 0.05）。与仅用mR1ECM培养的受精卵相比，在mKRB中培养的受精卵在注射冻干精子后发育成囊胚的比例也更高（32%比15%，P < 0.05）。将19个卵母细胞的2细胞胚胎注入在4℃下保存1年的冻干精子后移植，获得后代（16%）。本研究表明，注射精子后不久的培养条件显著影响胚胎的后续发育。在TE缓冲液中冷冻干燥后的大鼠精子在4℃下长期保存而不变质。(C) 2009爱思唯尔公司版权所有。
Rat sperm freeze-dried in a solution containing Tris and ethylenediaminetetraacetic acid (EDTA) (TE buffer) can be preserved at 4 degrees C, and oocytes injected with these sperm developed into offspring though developmental ability was low. We studied the culture conditions to improve the developmental ability of oocytes injected with freeze-dried sperm. After being injected with fresh sperm, the zygotes were cultured in modified Krebs-Ringer bicarbonate (mKRB), modified rat 1-cell embryo culture medium (mR1ECM)/BSA, and mR1ECM with different osmolality, before being cultured in mR1ECM. High proportion of zygotes cultured in mKRB (270 mOsm) before being cultured in mR1ECM developed into blastocysts compared to zygotes cultured only with mR1ECM (50% vs. 28%, P < 0.05). Culturing in mKRB also led to a high proportion of zygotes developing into blastocysts after the injection of freeze-dried sperm than zygotes cultured only with mR1ECM (32% vs. 15%, P < 0.05). Offspring (16%) were obtained when 19 2-cell embryos derived from oocytes that had been injected with freeze-dried sperm preserved at 4 degrees C for I year were transferred. This study demonstrated that the culture conditions soon after the injection of sperm markedly influenced the subsequent development of embryos. Also, rat sperm after freeze-drying in TE buffer were preserved at 4 degrees C for long term without their deterioration. (C) 2009 Elsevier Inc. All rights reserved.