黑猩猩精液冷冻和优选研究

对精液进行有效的冷冻保存和解冻优选是人工授精顺利开展的重要步骤。本研究在黑猩猩上做了尝试,分别用人工按摩采精、电刺激采精、附睾采精进行6次精液采集,所测精子活率为0.65～0.90,密度为1.69×10~（10）～6.64×10~（11）（个/L）。用自配精液冷冻液,采用CL-8800程序降温仪成功对所采集的精液进行冷冻保存。在冷冻后的10～400 d,分别对6次冻精解冻,并用人用精液优选试剂盒对其后3次进行解冻后优选。结果显示,解冻后精液活率为0.35～0.7;优选后精液活率为0.85～0.9,密度为4.5×10~（11）～5.0×10~（11）（个/L）,基本可以满足人工授精的需要。     

Sperm Mitochondrial Function is Affected by Stallion Age and Predicts Post-Thaw Motility

Sperm motility and quality decline with stallion age and sperm preservation, but the mechanisms of functional deficit have not been explained. We tested the hypothesis that mitochondrial deficits underlie age- and cryopreservation-related deficits in stallion fertility by measuring mitochondrial function, motility, and reactive oxygen species (ROS) in 88 frozen-thawed ejaculates from 19 stallions of varying ages. As expected with increasing age, total sperm motility, progressive motility, and average path velocity decreased, and ROS production increased. For every unit increase in oxygen consumption, there was a 77% increase in the odds of sperm movement (P < .05), confirming the link between mitochondrial functionality and motility. In addition, the rate of mitochondrial oxygen consumption increased from 4 years of age to a peak at 12 years of age and decreased steadily thereafter (P < .05). This confirms the importance of mitochondrial functionality for overall sperm health and motility, implicating mitochondrial dysfunction as a major contributor to sperm aging and cryopreservation damage.     

家兔精液品质评定方法的研究进展

家兔精子质量的评定,对于人工授精技术和体外受精技术的应用具有重要意义。用于反映精子质量的主要指标有：精液量、精子密度和形态、精子运动力、活率、质膜完整性、顶体完整性、精子染色质状态、线粒体活性和受精能力的检测。目前,多指标联合检测是生产中评定精子质量的有效方法。本文对上述指标的评定方法作一综述,并对其应用前景进行了展望。     

Sperm of the giant grouper: cryopreservation,physiological and morphological analysis and application in hybridizations with red-spotted grouper

In order to develop excellent germplasm resources for giant grouper (Epinephelus lanceolatus), cryopreservation of giant grouper sperm was examined in the present study. Firstly, 13 kinds of sperm dilution (ELS1-3, EM1-2, TS-2, MPRS, ELRS0-6) were prepared with physiological salt, sucrose, glucose and fetal bovine serum. The physiological parameters of ELRS3 (ratio of fast motion, ratio of slow motion, time of fast motion, time of slow motion, lifespan and motility) and ELS3 (sperm ratio of slow motion, time of slow motion and motility) were significantly higher than those of the other dilutions (P < 0.05). Secondly, after adding 15% DMSO and 10% FBS to ELRS3 and ELS3, most physiological parameters of frozen sperm were also significantly higher than the other gradients (P < 0.05), and sperm motility was as high as 63.68 ± 4.16% to74.75 ± 12.71% (fresh sperm motility, 80.70 ± 1.37% to 80.71 ± 1.49%). Mixed with the above dilutions, a final volume of 105 ml semen was cryopreserved. Finally, the sperm of giant grouper cryopreserved with cryoprotectants (ELRS3 + 15% DMSO + 10% FBS) was used for electron-microscopic observation and crossbreeding with red-spotted groupers (Epinephelus akaara). The electron-microscopic observation revealed that part of the frozen-thawed sperm was cryodamaged, e.g., flagellum fracturing and mitochondria falling out, while the ultrastructure of sperm membrane, mitochondria and flagellum remained intact. Also, the fertilization and hatchability rates of giant grouper frozen sperm and red-spotted grouper eggs were as high as 94.56% and 75.56%, respectively. Thus, a technique for cryopreservation of giant grouper sperm was successfully developed and applied to crossbreeding with red-spotted grouper eggs.     

The porcine sperm reservoir in relation to the function of hyaluronan

The oviduct plays a role in successful animal reproduction not only in spermatozoa and ova transport to the fertilization site but also by affording a microenvironment for fertilization and early embryonic development. The sperm reservoir (SR) is restricted in the uterotubal junction (UTJ) and caudal isthmus. Billions of porcine spermatozoa are distributed to the female reproductive tract during/after insemination, and small amounts of them are stored for about 36–40 hours in the SR, which maintains sperm viability in the pre-ovulation period through its surface epithelium and production of fluid. The SR regulates the release of spermatozoa so that only a small population moves towards the fertilization site (ampulla) to decrease polyspermy. This review attempts to provide information about the structure and function of the porcine SR, its intraluminal content (hyaluronan, HA), and the influences of HA on porcine spermatozoa in vivo. In pigs, the spermatozoa are stored in a mucous-like fluid within the UTJ and caudal isthmus in the pre-ovulation period. The oviduct fluid contains sulfated glycosaminoglycans (GAGs) and non-sulfated GAGs, i.e., HA. It is interesting to note that HA is synthesized by hyaluronan synthase-3 (HAS-3), and its receptor, CD44, is found in the epithelium of the porcine SR site. Additionally, sperm capacitation does not occur in vivo in the SR during the pre- and peri-ovulation periods, but spermatozoa in the SR will attempt to capacitate if exposed to bicarbonate. However, capacitation in the SR will rise in the post-ovulation period, indicating the role of HA in modulating sperm capacitation after ovulation. All data support the understanding that the porcine SR ensures the viability of fertile spermatozoa and maintains the non-capacitated status during the pre-ovulation period. This basic knowledge about the SR is believed to be useful to advance sperm preparation procedures for in vitro fertilization (IVF) and improve the preservation process of porcine semen.     

Effects of exposure to methylglyoxal on sperm motility and embryonic development after fertilization in mice

Methylglyoxal (MG) is a precursor for the generation of endogenous advanced glycation end-products involved in various diseases, including infertility. The present study evaluated the motility and developmental competence after in vitro fertilization of mouse sperm which were exposed to MG in the capacitation medium for 1.5 h. Sperm motility was analyzed using an SQA-V automated sperm quality analyzer. Intracellular reactive oxygen species (ROS), membrane integrity, mitochondrial membrane potential, and DNA damage were assessed using flow cytometry. The matured oocytes were inseminated with MG-exposed sperm, and subsequently, the fertilization and embryonic development in vitro were evaluated in vitro. The exposure of sperm to MG did not considerably affect the swim-up of sperm but resulted in a deteriorated sperm motility in a concentration-dependent manner, which was associated with a decreased mitochondrial activity. However, these effects was not accompanied by obvious ROS accumulation or DNA damage. Furthermore, MG diminished the fertilization rate and developmental competence, even after normal fertilization. Collectively, a short-term exposure to MG during sperm capacitation had a critical impact on sperm motility and subsequent embryonic development after fertilization. Considering that sperm would remain in vivo for up to 3 days until fertilization, our findings suggest that sperm can be affected by MG in the female reproductive organs, which may be associated with infertility.     

兔精子微注射受精研究Ⅰ——精子表型对精子微注射的影响

将不同表型和数目的精子注射到兔卵胞质或卵周隙中,结果表明,精子的表型对精子注射有影响。卵胞质中注射单个已获能死精子的受精率和发育率分别为53.26％和39.13％,注射单个未获能死精子的受精率和发育率为43.21％和24.69％,而注射单个未获能精子头核的受精率和发育率为18.18％和11.04％。前二者之间没有显著的差异。但与后者比较,受精率差异极显著,发育率差异显著。卵周隙中注射1～4个已获能运动精子的受精率和发育率均为23.08％,注射1～4个未获能运动精于的受精率和发育率为29.03％和25.81％,而注射1～4个已获能不运动精子的受精率和发育率均为5％。前二者差异不显著,但与后者比较,差异极显著。     

锰对公鸡繁殖性能和血清LH睾酮含量的影响

本文用组织学和放射免疫分析法就饲粮锰对公鸡睾丸组织学和血清睾酮、促黄体生成激素(LH)含量的影响进行了研究,并对公鸡的繁殖性琵进行了观察。结果表明,与Ⅲ组(添加锰100ppm)相比,缺锰的Ⅰ组(添加锰0ppm)或高锰的Ⅳ组(添加锰300ppm)辜丸生长发育受阻,曲细精管管径变小,精子数减少,种蛋孵化率下降,血清睾酮含量降低,LH含量略有上升。结果提示:缺锰或高锰日粮引起公鸡繁殖性能下降的重要原因之一是由于锰影响了睾丸间质细胞睾酮的分泌。     

奶牛卵胞质内单精注射(ICSI)的研究

[目的]探讨不同状态奶牛精子卵胞质内单精注射（ICSI）的效果。[方法]体外成熟培养24h的奶牛卵母细胞注入不同状态精子，观察其体外发育情况。[结果]结果表明，试验1，研究精子用5mmol DTT（二硫苏糖醇）预处理与精子不经过预处理的对照组的效果。经DTT处理后。精子解聚率和雄原核率显著高于对照组（30．4％和11．6％，47．8％和27．9％，P〈0．05），两组的卵裂率差异不显著（85．0％和80．0％）。囊胚率和孵化率显著高于对照组（45．8％和26．O％，31．3％和14．0％，P〈0．05）。试验2，比较死、活精子的卵胞质内单精注射的效果，两者之间卵裂率（75．0％和79．4％）、囊胚率（27．9％和30．9％）和孵化率（13．2％和16．2％）均无显著性差异（P〉0．05）。试验3，比较睾丸或附睾（头、体、尾）精子卵胞质内单精注射的效果，卵裂率（79．1％、81．8％、73．2％和75．0％）、囊胚率（23．3％、20．4％、19．5％和18．2％）和孵化率（14．O％、13．4％、12．2％和9．1％）均无显著性差异（P〉0．05）。[结论]DTF预处理奶牛精子有助于提高ICSI的效率；死精子能用于奶牛卵母细胞的ICSI；从睾丸精子到附睾尾精子，虽其成熟程度有很大不同，但对ICSI受精不产生显著影响。