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精子功能与制备培养基的选择:Percoll和Accudenz不连续密度梯度的比较

Sperm function and choice of preparation media: comparison of Percoll and Accudenz discontinuous density gradients.

作者信息

Sbracia M, Sayme N, Grasso J, Vigue L, Huszar G

机构信息

Sperm Physiology Laboratory, Department of Obstetrics and Gynecology, Yale University School of Medicine, New Haven, Connecticut, USA.

出版信息

J Androl. 1996 Jan-Feb;17(1):61-7.

PMID:8833742
Abstract

We compared the sperm populations prepared by Accudenz (35-65%) and Percoll (40-80%) density gradients in 21 normospermic specimens (concentration, 53.6 +/- 3.8 x 10(6) sperm/ml; motility, 44.5 +/- 3.5%). Accudenz facilitated a higher recovery of sperm and motile sperm (68.4 +/- 6.6% vs. 49.3 +/- 4.9%, P < 0.001, and 87.8 +/- 4.1% vs. 77.8 +/- 3.7%, P < 0.01, respectively). Sperm motility was lower in the Accudenz compared to the Percoll pellets; thus the values of total motile sperm recovered were not different (17.1 +/- 2.4 vs. 15 + /- 1 2.2 x 10(6) sperm/ml). The long term retention of sperm motility was substantially improved in Accudenz (at 24 hours, 34.9 +/- 2.8% vs. 26.3 +/- 1.5%; 60% vs. 40% of the initial motility, P < 0.001), and the Accudenz vs. Percoll samples also exhibited a higher retention of total motile sperm (at 24 hours, 9.8 +/-.2 vs. 6.1 = 0.5 x 10(6) motile sperm/ml, P < 0.05). The sperm motility index, a multiple of velocity and motility in the sample that reflects the efficiency of the sperm population in sperm-oocyte interaction, was 75% higher in the Accudenz samples at 24 hours (3.6 +/- 0.4 vs. 2.1 +/- 0.2, mu m/second, P < 0.01). Sperm cellular maturity by the creatine phosphokinase (CK) activity and CK-M to CK-B isoform ratio parameters (in the original samples 0.14 +/- 0.02 lU CK/100 x 10(6) sperm and 57.9 +/- 3.7%, respectively) were improved in both the Accudenz and Percoll pellets (P < 0.001), with no difference between the two sperm fractions. Sperm activation status monitored by chlortetracycline fluorescence indicated that after 4 hours of incubation the incidence of fully acrosome-reacted spermatozoa in the Accudenz versus Percoll pellets was 6.2 +/- 0.3% versus 13.1 +/- 1.0% (P < 0.001), a 100% increase in Percoll. We can conclude that Accudenz yields a higher concentration of motile spermatozoa, with improved retention of motility, velocity, and acrosomal integrity and without an increase of sperm with diminished cellular maturity. Thus, in sperm preparation for intrauterine insemination, in which the timing of ovulation and insemination frequently do not correspond, Accudenz-prepared sperm, with a better retention of motility/velocity and acrosomal integrity and with a consequential higher resistance to activation by the female reproductive tract, are expected to be more effective.

摘要

我们比较了在21份正常精子标本(浓度为53.6±3.8×10⁶精子/ml;活力为44.5±3.5%)中,通过Accudenz(35 - 65%)和Percoll(40 - 80%)密度梯度制备的精子群体。Accudenz能使精子和活动精子的回收率更高(分别为68.4±6.6%对49.3±4.9%,P<0.001;以及87.8±4.1%对77.8±3.7%,P<0.01)。与Percoll沉淀相比,Accudenz中的精子活力较低;因此,回收的总活动精子数量没有差异(分别为17.1±2.4对15±1.2×10⁶精子/ml)。Accudenz中精子活力的长期保持有显著改善(24小时时,分别为34.9±2.8%对26.3±1.5%;分别为初始活力的60%对40%,P<0.001),并且Accudenz与Percoll样本相比,总活动精子的保留率也更高(24小时时,分别为9.8±0.2对6.1±0.5×10⁶活动精子/ml,P<0.05)。精子活力指数是样本中速度和活力的倍数,反映精子群体在精子 - 卵母细胞相互作用中的效率,在24小时时Accudenz样本中的精子活力指数高75%(分别为3.6±0.4对2.1±0.2,μm/秒,P<0.01)。通过肌酸磷酸激酶(CK)活性和CK - M与CK - B同工型比例参数(原始样本中分别为0.14±0.02 lU CK/100×10⁶精子和57.9±3.7%)评估的精子细胞成熟度,在Accudenz和Percoll沉淀中均得到改善(P<0.001),两种精子组分之间没有差异。通过金霉素荧光监测的精子激活状态表明,孵育4小时后,Accudenz沉淀与Percoll沉淀中完全顶体反应精子的发生率分别为6.2±0.3%对13.1±1.0%(P<0.001),Percoll中的发生率增加了100%。我们可以得出结论,Accudenz能产生更高浓度的活动精子,活力、速度和顶体完整性的保留得到改善,且细胞成熟度降低的精子没有增加。因此,在用于宫内授精的精子制备中,由于排卵和授精时间常常不一致,Accudenz制备的精子具有更好的活力/速度和顶体完整性保留,以及因此对女性生殖道激活的更高抗性,预计会更有效。

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