No abstract

Despite the clinical importance of aneuploidy, surprisingly little is known concerning its impact during the earliest stages of human development. This study aimed to shed light on the genesis, progression, and survival of different types of chromosome anomaly from the fertilized oocyte through the final stage of preimplantation development (blastocyst). 2,204 oocytes and embryos were examined using comprehensive cytogenetic methodology. A diverse array of chromosome abnormalities was detected, including many forms never recorded later in development. Advancing female age was associated with dramatic increase in aneuploidy rate and complex chromosomal abnormalities. Anaphase lag and congression failure were found to be important malsegregation causing mechanisms in oogenesis and during the first few mitotic divisions. All abnormalities appeared to be tolerated until activation of the embryonic genome, after which some forms started to decline in frequency. However, many aneuploidies continued to have little impact, with affected embryos successfully reaching the blastocyst stage. Results from the direct analyses of female meiotic divisions and early embryonic stages suggest that chromosome errors present during preimplantation development have origins that are more varied than those seen in later pregnancy, raising the intriguing possibility that the source of aneuploidy might modulate impact on embryo viability. The results of this study also narrow the window of time for selection against aneuploid embryos, indicating that most survive until the blastocyst stage and, since they are not detected in clinical pregnancies, must be lost around the time of implantation or shortly thereafter.

DNA fragmentation is considered an important parameter of semen quality, and of significant value as a predictor of male fertility. Poor quality chromatin is closely associated with, and highly indicative of, some fertility problems. Many methodologies to assess DNA fragmentation in spermatozoa are available, but they are all unable to differentiate between single-stranded DNA breaks (SSB) and double-stranded DNA breaks (DSB) in the same sperm cell. The two-tailed Comet assay (2T-Comet) protocol overcomes this limitation. A modification of the original Comet assay was developed for the simultaneous evaluation of DNA SSB and DSB in human spermatozoa. The 2T-Comet assay is a fast, sensitive, and reliable procedure for the quantification and characterization of DNA damage in spermatozoa. It is an innovative method for assessing sperm DNA integrity, which has important implications for human fertility and andrological pathology.

MTHFR is an important enzyme in the metabolism of folic acid and is crucial for reproductive function. Variation in the sequence of MTHFR has been implicated in subfertility, but definitive data are lacking. In the present study, a detailed analysis of two common MTHFR polymorphisms (c.677C>T and c.1298A>C) was performed. Additionally, for the first time, the frequencies of different MTHFR alleles were assessed in preimplantation embryos. Several striking discoveries were made. Firstly, results demonstrated that maternal MTHFR c.1298A>C genotype strongly influences the likelihood of a pregnancy occurring, with the 1298C allele being significantly overrepresented amongst women who have undergone several unsuccessful assisted reproductive treatments. Secondly, parental MTHFR genotypes were shown to affect the production of aneuploid embryos, indicating that MTHFR is one of the few known human genes with the capacity to modulate rates of chromosome abnormality. Thirdly, an unusual deviation from Hardy-Weinberg equilibrium was noted for the c.677C>T polymorphism in subfertile patients, especially those who had experienced recurrent failure of embryo implantation or miscarriage, potentially explained by a rare case of heterozygote disadvantage. Finally, a dramatic impact of the MTHFR 677T allele on the capacity of chromosomally normal embryos to implant is described. Not only do these findings raise a series of interesting biological questions, but they also argue that testing of MTHFR could be of great clinical value, identifying patients at high risk of implantation failure and revealing the most viable embryos during in vitro fertilisation (IVF) cycles.

ABSTRACT:The frequency of sperm cells with fragmented DNA was studied in a group of 18 infertile patients with varicocele and compared with those obtained in a group of 51 normozoospermic patients, 103 patients with abnormal standard semen parameters, and 22 fertile men. The spermatozoa were processed to discriminate different levels of DNA fragmentation using the Halospermா kit, an improved Sperm Chromatin Dispersion (SCD) test. In this technique, after an acid incubation and subsequent lysis, those sperm cells without DNA fragmentation show big or medium-sized halos of dispersion of DNA loops from the central nuclear core. Otherwise, those spermatozoa containing fragmented DNA either show a small halo, exhibit no halo with solid staining of the core, or show no halo and irregular or faint stain of the remaining core. The latter, that is, degraded type, corresponds to a much higher level of DNA-nuclear damage. The varicocele patients showed 32.4% Ϯ 22.3% of spermatozoa with fragmented DNA, significantly different from the group of fertile subjects (12.6% Ϯ 5.0%). Nevertheless, this was not different from that of normozoospermic patients (31.3% Ϯ 16.6%) (P ϭ .83) and with abnormal semen parameters (36.6% Ϯ 15.5%) (P ϭ .31). No significant differences were found between the normozoospermic patients and the patients with abnormal semen parameters. Strikingly, the proportion of the degraded cells in the total of sperm cells with fragmented DNA was 1 out of 4.2 (23.9% Ϯ 12.9%) in the case of varicocele patients, whereas it was 1 out of 8.2 to 9.7 in the normozoospermic patients (11.1% Ϯ 9.9%) in the patients with abnormal sperm parameters (12.2% Ϯ 8.3%) and in the fertile group (10.3% Ϯ 7.2%). Thus, whereas no differences in the percentage of sperm cells with fragmented DNA were evident with respect to other infertile patients, individuals with varicocele exhibit a higher yield of sperm cells with the greatest nuclear DNA damage level in the population with fragmented DNA. This finding illustrates the value of assessing different patterns of DNA-nuclear damage within each sperm cell and the particular ability of the Halospermா kit to reveal them.Key words: Human sperm, DNA fragmentation. J Androl 2006;27:106-111 V aricocele, that is, dilation of the pampiniform venous plexus above and around the testicle, occurs in approximately 15% to 20% of the general male population, especially in adolescents. Varicocele occurs in 19% to 41% of men seeking infertility treatment and in around 80% of men with secondary infertility. Thus, this anatomical abnormality is perhaps one of the most common causes of poor sperm production and decreased semen quality.