Jakab Attila, Kovacs Tamas, Zavaczki Zoltan, Borsos Antal, Bray-Ward Patricia, Ward David, Huszar Gabor
The Sperm Physiology Laboratory, Department of Obstetrics and Gynecology, New Haven, CT, USA.
Hum Reprod. 2003 Jul;18(7):1481-8. doi: 10.1093/humrep/deg288.
We have previously shown that after 80% Percoll centrifugation there is an overall 2.7-fold reduction of sperm with chromosomal disomies and diploidies (3.2-fold and 2.0-fold respectively), and of sperm with diminished maturity as detected by cytoplasmic retention. The relationship between disomies and immature sperm was r = 0.7, suggesting that disomy primarily originates in immature sperm. In the present work we studied the efficacy of the swim-up method in elimination of sperm with diminished maturity and with chromosomal aberrations in the swim-up sperm fractions of 10 patients (sperm concentration: 20 +/- 3.9 x 10(6)/ml, range 8.9-45.5; sperm motility: 45.2 +/- 2.4, all mean +/- SEM).
The validity of the study was enhanced by assessing each sperm fraction with three-colour (X, Y and 17; 5000 sperm) and two-colour (10 and 11; 5000 sperm) chromosome probes using fluorescence in-situ hybridization (FISH). Thus, in each sample 10 000 sperm were evaluated. The incidence of diminished maturity sperm was assessed with creatine kinase immunocytochemistry.
In the swim-up fractions there was a reduction in the frequencies of disomic sperm, whether considering the sex chromosomes (1.4-fold) or the three autosomal chromosomes (1.5-fold based on the aggregate frequencies of disomy 10, 11 and 17). There was also a 1.5-fold reduction in diminished maturity sperm, indicating a relationship between the proportion of immature sperm and chromosomal aneuploidies (r = 0.46, P < 0.05, n = 20). Diploid sperm were reduced at a 2.7-fold rate, whether assessed with two- or three-colour FISH. There was a slight increase in the X/Y ratios.
Swim-up reduces the proportion of sperm with chromosomal aberrations and of sperm with diminished maturity. When compared with the results of the previous study with gradient centrifugation performed on semen samples with similar quality, the efficacy after swim-up is lower for disomies and higher for diploidies than that of gradient centrifugation.
我们之前已经表明,经过80%的 Percoll 离心后,染色体二体性和二倍体精子(分别为3.2倍和2.0倍)以及通过细胞质滞留检测出的成熟度降低的精子总体减少了2.7倍。二体性与未成熟精子之间的关系为r = 0.7,表明二体性主要起源于未成熟精子。在本研究中,我们研究了上浮法在10例患者的上浮精子部分中消除成熟度降低和染色体畸变精子的效果(精子浓度:20±3.9×10⁶/ml,范围8.9 - 45.5;精子活力:45.2±2.4,均为平均值±标准误)。
通过使用荧光原位杂交(FISH)用三色(X、Y和17;5000个精子)和双色(10和11;5000个精子)染色体探针评估每个精子部分,提高了研究的有效性。因此每个样本评估了10000个精子。用肌酸激酶免疫细胞化学评估成熟度降低精子的发生率。
在上浮部分,无论是考虑性染色体(1.4倍)还是三条常染色体(基于二体性10、11和17的总频率为1.5倍),二体性精子的频率都有所降低。成熟度降低的精子也减少了1.5倍,表明未成熟精子比例与染色体非整倍性之间存在关系(r = 0.46,P < 0.05,n = 20)。无论用双色还是三色FISH评估,二倍体精子都以2.7倍的速率减少。X/Y比值略有增加。
上浮法降低了染色体畸变精子和成熟度降低精子的比例。与之前对质量相似的精液样本进行梯度离心的研究结果相比,上浮后对于二体性的效果低于梯度离心,而对于二倍体的效果高于梯度离心。