摘要
The objective of this experiment was to determine the effects of flow cytometric sorting and freezing on stallion sperm fertility. A 2 × 2 factorial design was used to delineate effects of flow sorting and freezing spermatozoa. Oestrus was synchronised (July-August) in 41 mares by administering 10 ml altrenogest (2.2 mg/ml) per os for 10 consecutive days, followed by 250 μg cloprostenol i.m. on Day 11. Ovulation was induced by administering 3000 iu hCG i.v. either 6 h (fresh spermatozoa) or 30 h (frozen/thawed spermatozoa) prior to insemination. Mares were assigned randomly to one of 4 sperm treatment groups. Semen was collected from 2 stallions with an artificial vagina and processed for each treatment. Treatment 1 (n = 10 mare cycles) consisted of fresh, nonsorted spermatozoa and Treatment 2 (n = 16 mare cycles) of fresh, flow sorted spermatozoa. Spermatozoa to be sorted were stained with Hoechst 33342 and sorted into X- and Y-chromosome-bearing populations based on DNA content using an SX MoFlo sperm sorter. Treatment 3 (n = 16 mare cycles) consisted of frozen/thawed nonsorted spermatozoa (frozen at 33.5 × 106 sperm/ml in 0.25 ml straws) and Treatment 4 (n = 15 mare cycles) of flow sorted frozen/thawed spermatozoa (frozen at 64.4 × 106 sperm/ml). Concentrations of sperm in both cryopreserved treatments were adjusted, based on predetermined average post-thaw motilities, so that each insemination contained approximately 5 × 106 motile spermatozoa. Hysteroscopic insemination of 5 × 106 motile spermatozoa in a volume of 230 μl was used for all treatments. Pregnancy was determined ultrasonographically 16 days postovulation.
No differences were found (P>0.1) in the pregnancy rates for mares inseminated with fresh nonsorted (4/10 = 40.0%), fresh flow sorted (6/16 = 37.5%), frozen/thawed nonsorted (6/16 = 37.5%) and flow sorted frozen/thawed spermatozoa (2/15 = 13.3%). Pregnancy rates tended (P = 0.12) to be lower following insemination of frozen/thawed flow sorted spermatozoa. Further studies are needed with a larger number of mares to determine if fertility of flow sorted frozen/thawed spermatozoa can be improved.
摘要译文
该实验的目的是确定流式细胞分选和冻结种马精子生育的影响。A 2×2析因设计用来划定的流量分类和冷冻精子的影响。发情是同步的（月月）的41匹母马通过给予10毫升烯丙孕素（2。2毫克/毫升）经口，连续10天，随后250无论是6小时（鲜精子）或之前，受精30小时（冷冻/解冻精子）。母被随机分配到4个精子治疗组之一。精液从2种马收集用人造阴道和处理用于每个处理。处理1（N \x3d 10马的周期）由新鲜，nonsorted精子和新鲜的处理2（N \x3d 16马的周期），流分类的精子。精子进行排序进行染色用Hoechst 33342和分类到X轴和基于使用一个SX MoFlo精子分拣DNA含量Y-染色体轴承种群。治疗3（N \x3d 16马的周期）由冻结/解冻nonsorted精子（冷冻在33.5×10 SUP 6 / SUP精子/毫升0。25毫升秸秆）和流量的处理4（N \x3d 15马的周期）进行分类冷冻/解冻的精子（冻结在64.4×10 SUP 6 / SUP精子/毫升）。精子在两个冷冻治疗浓度进行了调整，基于预定平均解冻后motilities，使每个受精包含约5×10 SUP 6 / SUP活动精子。为5×10 SUP 6 / SUP活动精子中的230的体积宫腔镜授精升被用于所有的治疗方法。怀孕被确定超音波16天postovulation。没有差异（P1）在妊娠率为母马授精与新鲜nonsorted（4/10 \x3d 40.0％），新鲜流量排序（6/16 \x3d 37.5％），冷冻/解冻nonsorted（6/16 \x3d 37。5％）和流分类冷冻/解冻的精子（2/15 \x3d 13.3％）。怀孕率趋向（P \x3d 0.12）要低以下冷冻/解冻流程人工授精的精子进行排序。需要进一步的研究以母马的更大数目，以确定是否流生育排序冷冻/解冻精子可以得到改善。
A. C. LINDSEY, J. L. SCHENK, J. K. GRAHAM, J. E. BRUEMMER and E. L. SQUIRES. Hysteroscopic insemination of low numbers of flow sorted fresh and frozen/thawed stallion spermatozoa[J]. Equine Veterinary Journal, 2002,34(2): 121-127