不同月龄巴杜公猪精液品质分析

为研究不同月龄巴杜种公猪的精液品质差异,本实验选择8～12、13～18、19～24、25～30、31～36月龄各6头巴杜种公猪,进行精液品质检查。结果表明:随着月龄增加,巴杜种公猪精液的各项指标呈现规律性变化,19～24月龄的采精量、精子密度和精子活率显著高于其他月龄段,且精子畸形率显著低于其他月龄段;24月龄之后采精量、精子密度和精子活率缓慢下降,而畸形率则逐渐上升。本研究结果可为合理选择配种日龄、人工授精公猪的使用年限及人工授精的输精量提供数据支撑,为巴杜种公猪精液的合理应用及提高猪场经济效益提供参考依据。     

Effects of the Addition of Autologous Seminal Plasma to Highly Concentrated Stallion Semen After 48 Hours of Cooled Storage

Insemination with chilled transported semen has become distinctly important in the horse-breeding industry. To ensure cell survival during cooled storage, semen is diluted with an appropriate extender and the concentration of seminal plasma (SP) is reduced. Nevertheless, SP plays an important immunomodulatory role in the female genital tract and supports sperm fertility. The aim of the present study was to evaluate the effect of the addition of autologous SP after cooled storage to highly concentrated stallion semen. Therefore, SP was removed by simple centrifugation of extended semen, aspiration of the supernatant, and resuspension of the sperm pellet with semen extender. Motion characteristics were evaluated after cooled storage for 48 hours at concentrations of 333 × 10⁶ sperm/mL in comparison with stored samples at concentration of 25 × 10⁶ sperm/mL (control). The highly concentrated semen samples were diluted with an extender containing 0%, 5%, 20%, and 80% SP directly before motility analysis. Dilution of the cooled semen with a fresh semen extender without SP (0%) increased kinematic parameters (curvilinear velocity [VCL] 137.3 vs. 151.8; straight-line velocity [VSL] 49.0 vs. 57.5; average path velocity [VAP] 69.5 vs. 79.4 μm/second; amplitude of lateral head [ALH] 3.1 vs. 3.3 μm; beat cross frequency [BCF] 31.6 vs. 33.5 Hz; P < .05) but not total motility (51% vs. 43%) and progressive motility (46% vs. 36%) compared with controls. The addition of SP after storage for 48 hours decreased sperm total motility and progressive motility regardless of SP concentration: 5 (38% and 34%), 20 (37% and 33%), and 80% SP (27% and 22%; P < .05). In contrast, kinematic parameters were enhanced by extenders containing 5% and 20% SP (VCL: 148.0 and 155.6; VSL: 59.2 and 60.9; VAP: 78.7 and 81.9; BCF: 33.4 and 35.7; ALH: 3.4 and 3.4; P < .05). However, using an extender containing 80% SP was detrimental to kinematic parameters (VCL: 151.2; VSL: 52.2; VAP: 76.9; BCF: 34.8; P < .05) except for ALH, which increased (3.5; P < .05). In conclusion, cooled storage at concentrations of 333 × 10⁶ sperm/mL did not affect sperm motility. The addition of a fresh extender or an extender containing small concentrations of SP to highly concentrated ejaculated sperm increased kinematic values after storage; however, increasing concentrations of SP decreased sperm motility.     

Efficiency of three boar sperm enrichment techniques

The objective of the study was to investigate the efficiency of three enrichment methods to separate boar spermatozoa. Twenty-four ejaculates from 12 boars (2 ejaculates/boar) were extended (30 × 10⁶ spermatozoa/mL) in commercial Beltsville Thawing Solution. Each semen sample was processed with glass wool column (GW) and glass beads (GB) filtration and with the single-layer centrifugation (SLC) technique. Semen samples before (control; C) and after treatment were evaluated for sperm CASA motility/kinetics and concentration, viability, morphology and chromatin integrity. Data were analysed with mixed models. The concentration of total and motile spermatozoa was significantly decreased after treatment in groups GW and SLC, but not in group GB. Group GW showed increased values of WOB compared with both groups C and GB. Group GB showed greater values of rapid movement spermatozoa and lower values of slow movement spermatozoa compared with group C. In group SLC, higher values of VSL, LIN and STR were observed compared with group C. In conclusion, all techniques under examination enhanced various CASA variables. Based on our results, the GB method is a promising alternative separation technique for boar sperm and deserves further research regarding swine in vitro fertilization.     

稀释液溶质和渗透压对鸡精液低温保存效果的影响

观察15种物质不同渗透压（300mOsm、400mOsm、500mOsm）对鸡精子低温保存的精子活力、生存指数、存活时间和30h精子畸形率的影响。结果表明,对鸡精子低温保存生存指数大于50的物质由高到低依次为果糖、甘露醇、谷氨酸钠、山梨醇和氯化钾;精子总存活时间大于120h的物质由高到低依次是果糖、甘露醇、谷氨酸钠、山梨醇、氯化钾、磷檬酸钠和乙酸钾;30h精子畸形率小于0．2的物质由低到高依次为L-谷氨酸钠、甘露醇、蔗糖、乳糖。渗透压分析结果表明,各指标均随渗透压升高而降低。在300mOsm时,30h精子畸形率、精子生存指数和存活时间均极显著高。     

不同奶牛精子稀释液配方比较研究

牛的冻精稀释液是精液冷冻保存和人工授精最关键的环节。本文进行了两个方面的试验研究,一是把各个地区生产实践中应用的不同冷冻稀释液配方,进行综合筛选和比较;二是对没有条件冷冻精子的养殖场进行精子保鲜的稀释液配方进行综合筛选和比较。结果显示,当冷冻保存精液时,选择配方2为最佳配方;常温保存时选择配方2、3、4均可。     

人和哺乳动物精液的蛋白质组学研究进展

蛋白质组学技术是21世纪研究生命科学最有价值的技术之一,将其应用到精子和精浆蛋白质的研究后,已取得了许多成果。对蛋白质组、蛋白质组学、双向电泳、质谱、生物信息学等进行了简要介绍,并重点阐述了蛋白质组学技术在精子蛋白和精浆蛋白上的研究进展。     

Effects of Hyaluronan Supplementation on Cryopreserved Equine Spermatozoa Hyaluronan and Cryopreserved Equine Spermatozoa

Addition of hyaluronan, a nonsulfated glycosaminoglycan, to fresh and frozen thawed human semen results in substantial retention of motility over time. Hyaluronan also has been reported to preserve postthaw viability and maintain membrane stability of boar spermatozoa. Therefore, experiments were designed to investigate the use of a commercially available hyaluronan (Map-5, Bioniche Animal Health, Inc., Athens, GA) in freezing extender for cryopreservation of equine spermatozoa. In experiment 1, aliquots from ejaculates were supplemented before freezing with one of four levels of hyaluronan: 100 μg/mL, 200 μg/mL, 400 μg/mL, and 1000 μg/mL along with an untreated control. No differences in sperm motility, assessed by computer-assisted sperm motility analysis (CASA), were found for any treatment at times 0, 30, or 60 minutes postthaw. Decreases in motility were noted in the highest hyaluronan group (1,000 μg/mL) after 90 and 120 minutes of incubation. Sperm viability, as assessed using SYBR-14/propidium iodide staining, was decreased (P < .05) when treated with 1,000 μg/mL compared with the control (37.1% and 46.1%, respectively). Motility parameters tended to remain elevated in those ejaculates treated with 200 μg/mL at various time points. Experiment 2, therefore, further investigated the effects of hyaluronan at 200 μg/mL on motility parameters and acrosome integrity and zona pellucida binding. Total (TM) and progressive (PM) motility of treated sperm immediately after thawing and at 60 minutes post-thaw were higher compared with control (P < .05). A tendency (P < .1) to maintain TM at 90 and 120 minutes post-thaw also was noted. No differences were noted for the mean number of spermatozoa bound to bovine oocytes for control or treated sperm (22 ± 14 vs 25 ± 17, respectively). Acrosome integrity also was unchanged between the two groups based on fluorescein isothiocyanate (FITC)−peanut agglutinin (PNA)/propidium iodide staining. All samples contained <1% live acrosome-damaged spermatozoa. In the final experiment, the effects of hyaluronan supplementation post-thaw was investigated using hyaluronan concentrations of 100, 200, and 400 μg/mL. Motility parameters studied over an 8-hour period at 37°C yielded no consistent differences. In conclusion, addition of hyaluronan at a concentration of 200 μg/mL before freezing increased spermatozoal post-thaw motility. High concentration of hyaluronan (1,000 μg/mL) appeared to be detrimental to post-thaw motility. Effects of hyaluronan on fertility are beyond the scope of this study and have yet to be determined.     

Live foals produced from sperm-injected oocytes derived from pregnant mares

Recently, in vitro fertilization (IVF) in the horse has met with less than anticipated results. Various problems associated with equine IVF include: (1) the inability to collect large numbers of good quality oocytes, (2) the alteration of the zona pellucida associated with in vitro maturation of equine oocytes, and (3) the improper preparation of equine sperm cells for IVF of these oocytes. Therefore, this study was conducted to achieve fertilization via sperm injection of equine oocytes and to produce live offspring from this IVF procedure. Oocytes were collected by transvaginal ultrasound-guided oocyte retrieval procedures from early pregnant mares of mixed breeds (day 14 to day 70 of pregnancy) and were matured in vitro and subjected to intracytoplasmic sperm injection (ICSI). Injected oocytes were then cultured for 48 hours in either TCM-199 or P-1 medium (glucose and phosphate-free medium) supplemented with 15% fetal bovine serum. Cleavage rates for embryos cultured in the two culture media were different (47% vs. 63% in TCM-199 and P-1, respectively). Also, four Grade 1 embryos were surgically transferred into the oviducts of four recipient mares (one embryo/mare) at 48 hours post-ICSI, with three pregnancies (75%) developing as ultrasonically demonstrated by the presence of an embryonic vesicle in the uterine body by day 16 post-ICSI. On June 23rd one live filly was born after 328 days of gestation and subsequently, a second healthy filly was born after 319 days of gestation. To our knowledge, this is the first report of live foals resulting from in vitro fertilization (via ICSI) of in vitro matured oocytes recovered from pregnant mares using an efficient, repeatable transvaginal ultrasound-guided procedure.     

LDL对牛精子冷冻-解冻后运动特性的影响

在牛冷冻精液稀释液中,卵黄中的低密度脂蛋白(LDL)对精子起主要保护作用。本研究利用含有7%、8%、9%和10%(w/v)LDL配制成的稀释液制作牛细管冷冻精液(0.25mL/支),冷冻-解冻后,通过计算机辅助分析系统(CASA)对精子运动参数进行分析。结果表明:稀释液中含8%(w/v)LDL时冷冻效果最好,其冻后精子活力平均可达55.6%,显著高于其他组(P<0.01);精子直线速度平均为32.5μm/s,平均路径速度为35.7μm/s,与对照组相比差异显著(P<0.05);a,b级精子可占精子总数的54%。本研究为牛冷冻精液新型稀释液的开发与利用提供了一定的依据。