Magargee S F, Cramer P G, Hammerstedt R H
Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania, USA.
Mol Reprod Dev. 2000 Dec;57(4):406-11. doi: 10.1002/1098-2795(200012)57:4<406::AID-MRD13>3.0.CO;2-8.
We report use of an in vitro assay (Barbato et al., 1998: Biol Reprod 58:686-699) to assess binding ability of cauda epididymal mouse sperm to a surrogate zona pellucida and effect of a synthetic peptide (Amann et al., 1999: J Androl 20: 42-46) on fertilization ability in in vitro fertilization (IVF) tests. Sperm from C57Bl/6, CD1, and CF1 mice (4 replicates each) were evaluated for binding ability after exposure to 0 (control) and 80-1280 pM peptide. For control sperm, endogenous binding was C57Bl/6 < CD1 = CF1 (P < 0.05, 1-way ANOVA). Across all three strains, exposure to > 320 pM peptide increased relative binding of sperm (P < 0.05; 2-way general linear model; GLM). Strains differed both in basal binding ability and in response to synthetic peptide. To determine if IVF rate increased after exposure of sperm to peptide, ova from B6C3 mice (four replicate pools) were collected after eCG and hCG stimulation. Cumulus-oocyte complexes (COC; 8-15 ova in each of 3-6 drops/treatment) were incubated with hyperactivated C57Bl/6 sperm at approximately 1500 sperm per ovum. Data for incubations were corrected for false-positive classification to yield a better estimate of true cleavage rate, and then related to results observed with a tenfold greater sperm concentration. Relative cleavage rates were 0 peptide (0.48); 420 pM (0.78, P < 0.05); and 840 pM (0.90, P < 0.01; GLM and Tukey tests). IVF rate was increased by exposure of mouse sperm to peptide at concentrations effective in the in vitro assay, and use of peptide allowed use of 1/10 as many sperm.
我们报告了使用一种体外试验（Barbato等人，1998年：《生物繁殖》58:686 - 699）来评估附睾尾部小鼠精子与替代透明带的结合能力，以及一种合成肽（Amann等人，1999年：《雄性学杂志》20:42 - 46）在体外受精（IVF）试验中对受精能力的影响。将来自C57Bl/6、CD1和CF1小鼠的精子（各4个重复样本）在暴露于0（对照）和80 - 1280 pM的肽后评估其结合能力。对于对照精子，内源性结合能力为C57Bl/6 < CD1 = CF1（P < 0.05，单因素方差分析）。在所有三个品系中，暴露于> 320 pM的肽会增加精子的相对结合能力（P < 0.05；双因素一般线性模型；GLM）。品系在基础结合能力和对合成肽的反应方面均存在差异。为了确定精子暴露于肽后体外受精率是否增加，在促卵泡素（eCG）和人绒毛膜促性腺激素（hCG）刺激后收集B6C3小鼠的卵子（四个重复样本池）。卵丘 - 卵母细胞复合体（COC；每个处理的3 - 6滴中各有8 - 15个卵子）与超活化的C57Bl/6精子一起孵育，每个卵子约1500个精子。对孵育数据进行假阳性分类校正，以更好地估计真实的卵裂率，然后将其与精子浓度高十倍时观察到的结果相关联。相对卵裂率分别为：0肽（0.48）；420 pM（0.78，P < 0.05）；840 pM（0.90，P < 0.01；GLM和Tukey检验）。小鼠精子暴露于在体外试验中有效的浓度的肽后，体外受精率增加，并且使用肽可使精子用量减少至十分之一。
