Whereas studies have revealed that the cryopreservation of human semen increases sperm DNA fragmentation, the mechanisms involved in this type of cryo-injury are largely unknown. Elucidation of these mechanisms may provide insight into preventing such injury. We obtained 60 semen samples from 60 men and conducted experiments to determine the cause of cryopreservation-induced DNA fragmentation using 8-oxo-7,8-dihydro-2′deoxyguanosine (8OHdG) as a biomarker of oxidative stress, percentage caspase positive cells as an indicator of apoptosis, the potential antioxidant genistein and the caspase inhibitor Z-VAD(OMe)-FMK. Cryopreservation led to a significant increase in percentage DNA fragmentation, percentage 8OHdG and percentage caspase positive cells (P < 0.001). Percentage DNA fragmentation was positively correlated with percentage 8OHdG before (r = 0.756, P < 0.001) and after cryopreservation (r = 0.528, P = 0.017). The addition of 50 and 100 µM genistein to the cryoprotectant had a significant protective effect on sperm DNA (P < 0.001) although the caspase inhibitor demonstrated no difference to the control. Human sperm DNA fragmentation is associated with an increase in oxidative stress during cryopreservation, rather than the activation of caspases and apoptosis. The estrogenic compound genistein may be useful in reducing this effect but larger trials are needed to confirm this.

Integrins are heterodimeric glycoproteins that have been found to undergo dynamic temporal and spatial changes in distribution in the endometrium during the menstrual cycle in women. Likewise the extracellular matrix (ECM) ligands for these receptors are likely to play a role in the establishment of a receptive endometrium. To develop primate models to study the role of these molecules in the cascade of molecular events leading to implantation, integrin expression and associated changes in ECM were investigated during the menstrual cycle and in early pregnancy in the baboon. Antibodies specific for the integrins (alpha(1-6) and alpha(v); beta1, beta3, and beta4) and ECM (laminin, collagen IV, fibronectin) were utilized. In addition, cytokeratin and alpha-smooth muscle actin were used as epithelial, stromal, and smooth muscle cell markers, respectively. Endometrium was obtained in duplicate or triplicate during the menstrual cycle and early pregnancy. Changes observed during the natural menstrual cycle were confirmed using ovariectomized, steroid-treated animals. Constitutively expressed integrins on the endometrial epithelium included the collagen/laminin receptors: alpha2, alpha3, alpha6, and beta4. The pattern of expression correlated well with the distribution of ECM in this tissue. Collagen IV was confined to the basement membrane of glandular epithelium and blood vessels. Laminin immunostaining was found in the basement membrane, mostly in the stroma of the basal region, in the glandular endometrium and vasculature. Fibronectin was present throughout the stroma but not in the basement membrane. The collagen receptor alpha1 beta1 and fibronectin receptor alpha4 beta1 appeared in the glandular epithelium in the luteal phase. As in the human, alpha1 and alpha4 disappeared from the glandular epithelium with the establishment of pregnancy. In contrast, the alpha4 beta3 vitronectin receptor appeared in the glandular epithelium only in pregnancy or following long-term steroid treatment with estrogen and progesterone but not during the time of uterine receptivity associated with the initial period of embryo attachment. Osteopontin, an ECM ligand for alpha(v) beta3, was coexpressed with this integrin in invading cytotrophoblasts, glandular epithelium, and decidualizing stromal cells. Decidualization in the baboon was associated with changes in integrin expression similar to those found in humans: there was an increase in alpha1, alpha3, alpha6, beta1, and alpha(v) beta3 in the decidualized stromal cells. Laminin and collagen IV expression also increased at the implantation site and throughout the endometrium. In contrast, fibronectin expression was most evident at the implantation site and corresponded to alpha5 expression on the invading cytotrophoblasts. In summary, marked similarities were found in the expression of ECM and the integrin receptors between the baboon and the human endometrium throughout the menstrual cycle and in pregnancy. Cycle-specific integrins, alpha1, and alpha4, were present on epithelial cells during the secretory phase. Delayed expression of alpha(v) beta3 in baboon endometrial glands correlated closely with the time of enhanced glandular secretory activity in this primate. The baboon appears to be an excellent model for the investigation of the role of integrins and ECM leading to successful implantation.

A membrane-based electrophoretic filtration system, known as the Cell Sorter-10 (CS-10), that preferentially isolates spermatozoa with very low levels of DNA damage has recently been developed. However, it remains to be proven whether spermatozoa prepared in this way are capable of achieving fertilization in assisted conception. Therefore, this clinical trial was designed to answer this question. A split-sample split-cohort study design was employed to control for differences in semen and oocyte quality between 28 couples undergoing either intracytoplasmic sperm injection (ICSI) or IVF in this clinical trial. Each semen sample was split between preparation using the CS-10 and preparation by standard density gradient centrifugation (DGC) and each cohort of oocytes was split for insemination using either CS-10 (n = 197) or DGC (n = 195) prepared spermatozoa. Both methods of sperm preparation yielded comparable rates of sperm recovery, motility and DNA fragmentation. There was no significant difference between the ability of CS-10 and DGC prepared spermatozoa to produce fertilization (62.4% versus 63.6%), cleavage (99.0% versus 88.5%) and high-quality embryos (27.4% versus 26.1%). This pilot study demonstrates that membrane-based electrophoresis is as effective as DGC in preparing sperm for IVF and ICSI, although it takes only a fraction of the time.

Semen quality represents a compelling factor for fertility, and delineating the normal values has proven difficult. In the last four decades, several authors have reported a noticeable decline in sperm parameters. Also, studies investigating ‘time to pregnancy’ have shown that fecundity begins to be reduced when sperm numbers decrease below 30 million, even though according to the 6th edition of the WHO manual, the normal value is currently 16 million/mL or 39 million per ejaculate. There exists sufficient data to suggest a decline in sperm counts over time, even though the clear reason for this adverse trend is not well established, but some associations have been hypothesised, such as maternal smoking during pregnancy. Additional potential factors have yet to be fully illustrated but involve poor diet, increased obesity, and exposure to environmental toxins. Moreover, the change in environmental conditions and more common exposure to endocrine-disrupting chemicals (EDCs), such as pesticides and herbicides, as well as bisphenol A, phthalates, polychlorinated biphenyls, and heavy metals, starting from prenatal life and continuing into adulthood, may exhibit probable features explaining the reduction in sperm parameters. Therefore, the main goal of this narrative review is to furnish an overview of the possible effects of exposure to EDCs on testicular function and spermatogenesis and, also, to summarise the evidence regarding a decrease in sperm quality and examine its potential consequences.

The protein ubiquitin (Ub) appears to be present in all eukaryotic cells. Its widespread presence and extremely conserved structure indicate that it may play a vital role in cell metabolism. The roles of Ub are mediated by its covalent attachment to target proteins, a process known as ubiquitylation, a form of protein modification which may lead to degradation of the modified protein. A number of proteins with similar structure to Ub but varying in function have been isolated. Recently, there has been much interest in the role of Ub and its related proteins in reproductive processes. Ub and Ub-related proteins may be involved in gametogenesis, modulation of steroid receptor concentrations, placental development and endometrial modification at the beginning of pregnancy. These wide-ranging effects have led to extensive research which will be reviewed in this article.