Association Between Anogenital Distance, Sperm Phenotype and Epigenetics in Infertile Men.
试验速览
- 阶段
- 不适用
- 状态
- 已完成
- 入组人数
- 60
- 试验地点
- 1
- 主要终点
- anogenital distance and epigenetics
研究概览
简要总结
Testicular dysgenesis syndrome (TDS) is known to cause epigenetic abnormalities in spermatozoa. Anogenital distance (AGD) is considered to be a suitable clinical marker of TDS, but the direct link between AGD and epigenetic abnormalities is still missing.
Infertile men (n=10) presenting with shortened AGD and a control group of normal semen donors (n=10) with normal AGD will then be asked to provide one semen sample each. Using a flow cytometer and sorter (FACS) their spermatozoa will be sorted into populations of spermatozoa with/without DNA fragmentation or with/without chromatin decondensation. These sorted populations of spermatozoa will then be examined for differences in epigenetic imprinting differences using whole genome expression analysis. Whereas the sorting of spermatozoa will be carried out in Basel, the epigenetic analysis will be carried at the University of Geneva.
详细描述
A subset of 10 men with shortened AGD (together with a control group of 10 fertile donors with normal AGD) will be asked to provide up to three semen samples, each of which then will be sorted with FACS into subpopulations with/without DNA fragmentation and into subpopulations with/without chromatin decondensation.
Spermatozoa with fragmented DNA will be separated through FACS-sorting of spermatozoa with intact DNA using the YoPro 1-dye, which has been shown to correlate significantly with the degree of DNA fragmentation in the nuclei of sperm.
In addition, spermatozoa with abnormal chromatin remodelling will be separated through sorting from spermatozoa with condensed chromatin using the fluorochrome chromomycin A3 (CMA3), which competes for protamin for binding to the minor groove of DNA thereby correlating with the persistence of histones in the sperm nuclei. Pilot experiments have demonstrated the highly significant and close correlation of CMA3 with anilin blue staining. Anilin blue staining is not suitable for the sorting experiment, because it requires fixation of the spermatozoa. Sorting based on CMA3 can be carried out with living spermatozoa.
The sorted and anonymized samples will then be sent frozen in dry ice to a laboratory at the University of Geneva for the assessment of differences in the epigenetic imprinting of the DNA using whole genome expression studies.
研究设计
- 研究类型
- Observational
- 观察模型
- Case Control
- 时间视角
- Prospective
入排标准
- 年龄范围
- 20 Years 至 55 Years（Adult）
- 性别
- Male
- 接受健康志愿者
- 否
入选标准
- •infertile men with known anogenital distance
排除标准
- •sperm concentration must be more than 15 million/ml to allow appropriate sorting with flow cytometry...
结局指标
主要结局
anogenital distance and epigenetics
时间窗: 6 months
number of differentially methylated transposable regulatory sequences in the genome of sorted spermatozoa.
次要结局
- sperm phenotype 2(6 months)
- sperm phenotype 3(6 months)
- sperm phenotype 4(6 months)
- sperm phenotype 5(6 months)
- sperm phenotype 1(6 months)
研究者
Christian De Geyter
Prof. Dr.med.
University of Basel