Mariangela Primiterra

Differences in blood and semen oxidative status in fertile and infertile men, and their relationship with sperm quality.

Oxidative stress plays a fundamental role in the aetiology of male infertility by negatively affecting sperm quality and function. Assessment of blood and seminal plasma oxidative profiles might be a valuable tool to improve evaluation of sperm reproductive capacity and functional competence. This study examined the lipid-soluble antioxidant profile and levels of lipid peroxidation both in blood and seminal plasma samples of infertile and fertile males, in relation to semen parameters. Total antioxidant capacity (TAC) and vitamin E concentrations were significantly (P<0.05) lower in seminal plasma of infertile men compared with fertile subjects; concurrently, a significant accumulation of malondialdehyde was found in infertile patients (P=0.032 compared with controls), which was negatively correlated with sperm motility and morphology. In blood samples, infertile men presented lower concentrations of TAC, carotenoids and vitamin E than fertile subjects; TAC and carotenoids were positively correlated with sperm motility, morphology and concentration. Finally, blood TAC and vitamin E concentrations were positively correlated with the corresponding seminal values, confirming the close relationship between blood and semen antioxidants. All these results indicated the possibility of using not only seminal antioxidants but also blood antioxidants as biochemical markers to support sperm quality evaluation. Oxidative stress induced by reactive oxygen species (ROS) has been widely recognized as one of the major causes of male infertility; indeed, excessive ROS production can negatively impact sperm quality and function. The assessment of blood and seminal plasma oxidative profiles has been suggested as a valuable tool to improve the evaluation of sperm reproductive capacity and functional competence in infertile men. With this in mind, in the present study we examined the lipid soluble antioxidant profile (carotenoids and vitamins A and E) and the levels of lipid peroxidation (malondialdehyde; MDA) both in blood and seminal plasma samples of infertile and fertile males, in correlation with semen parameters namely motility, morphology and concentration. As a result, we obtained evidence that the total antioxidant capacity (TAC) and the concentrations of vitamin E of seminal plasma samples were significantly lower in infertile men than in fertile subjects; at the same time, a significant accumulation of MDA was found in infertile patients. MDA, in turn, negatively correlated with sperm motility and morphology, thus confirming that oxidative damage to lipids impairs sperm quality. In blood samples, infertile men presented lower TAC and lower concentrations of carotenoids and vitamin E than fertile subjects; interestingly, TAC and carotenoid concentrations were positively correlated with sperm motility, morphology, and concentration, confirming the close relationship between blood antioxidants and sperm quality. In conclusion, all these results suggested that the examination of blood and semen oxidative profiles might furnish useful information on sperm quality and function in infertile men.

Counteraction of oxidative damage in the rat liver by an ancient grain (Kamut brand khorasan wheat)

OBJECTIVE We previously demonstrated in rat plasma the antioxidant protective effect of whole-grain bread, particularly when made from Kamut brand khorasan wheat. In the present study, we investigated the effects of the same experimental breads in rat liver using two different bread-making procedures (bakers yeast and sourdough fermentation). METHODS Rats were examined in the basal condition and after the administration of doxorubicin, a pro-oxidative agent. The following parameters were measured in liver homogenates: glutathione peroxidase and thioredoxin reductase activities, as antioxidant enzymes containing selenium; glutathione, α-tocopherol and β-carotene, as major non-enzymatic cell antioxidants; malondialdehyde and advanced oxidation protein products, as markers of oxidative damage to lipids and proteins, respectively. A histologic evaluation of liver tissue was also conducted. RESULTS In agreement with our previous work, we observed a lower oxidative status and a different activity of glutathione peroxidase and thioredoxin reductase in rats fed the whole-grain Kamut khorasan bread than in rats fed the modern whole-grain durum wheat bread. Histologic evaluation of the hepatic tissue showed the onset of inflammation in response to doxorubicin only in rats fed the modern durum wheat bread. CONCLUSION Our data confirm that bread made from whole-grain Kamut khorasan protects rats from oxidative stress better than bread made from whole-grain durum wheat. This is consistent with their different antioxidant profiles. The type of wheat used for bread-making appeared to be the main determinant of the observed protective effect.

Influence of ovarian stimulation for IVF/ICSI on the antioxidant defence system and relationship to outcome.

Ovarian stimulation is used with IVF/intracytoplasmic sperm injection (ICSI) cycles to obtain multiple oocytes and improve pregnancy rates; however, it also induces perturbation in the oxidant-antioxidant balance leading to oxidation stress. The present study monitored the plasma antioxidant status in women undergoing a long agonist protocol of ovarian stimulation at three different time points: at baseline (T0), after pituitary suppression (T1) and on the day of oocyte retrieval (T2). The antioxidant composition of follicular fluid samples collected on T2 was also evaluated. Significant decreases (P < 0.05) of plasma vitamin C, vitamin E and carotenoids were found between T1 and T2 but not between T0 and T1. At T2, high plasma vitamin E was associated with high numbers of total and mature oocytes retrieved per patient, which, in turn, were favourable for achieving pregnancy. Accordingly, women who became pregnant presented higher vitamin E concentrations both in plasma and FF than those who did not. In conclusion, this study confirmed the occurrence of significant modifications of the plasma antioxidant profile during ovarian stimulation with gonadotrophins; at the same time, it was found that both systemic and follicular antioxidant status may be related to IVF/ICSI outcome.

Extracellular embryo genomic DNA and its potential for genotyping applications.

Background: Preimplantation genetic diagnosis (PGD) currently relies on biopsy of one or few embryo cells. Our aim was to evaluate the embryo extracellular matrices (spent medium and blastocoele fluid) as source of DNA for embryo genotyping. Results/methodology: We first evaluated the amplifiability and the amount of genomic DNA in spent embryo culture media from day 3 (n = 32) and day 5/6 (n = 54). Secondly, we evaluated the possibility to genotype the MTHFR polymorphism C677T from media at day 5/6 (n = 8) and blastocoele fluids (n = 9) by direct sequencing. The C677T polymorphism detection rate was 62.5 and 44.4% in medium and fluid, respectively. Conclusion: A noninvasive approach for embryo genotyping was possible, but still with limitations due to low detection rate and possible allele dropout.

Pre-implantation genetic diagnosis and screening: now and the future.

Abstract Since 1989, the year of the first pre-implantation genetic diagnosis (PGD), many developments occurred both in assisted reproduction techniques and in molecular tools. While PGD is a well-established and documented application, pre-implantation genetic screening (PGS) for the detection of aneuploid embryos is still debated due to the presence of mosaicism in the embryo, but especially to the knowledge of the limits that label an embryo as healthy or as appropriate to the life. The aim of this review is to present the state-of-the-art in the field of PGD and PGS, illustrating its benefits and limitations, along with biopsy techniques and the use of new high-throughput technologies.

Comparison of in vitro fertilization outcomes in ICSI cycles after human sperm preparation by density gradient centrifugation and direct micro swim-up without centrifugation

Objective The aim of this study was to evaluate the efficacy of a non-expensive, easy and fast technique (direct micro swim-up) for sperm preparation in intracytoplasmic sperm injection (ICSI) treatments without the use of centrifuge. Methods We carried out a multicentric study in which a total of 140 ICSI-cycles were included. Sibling oocytes were divided into two groups according to semen preparation procedures: group A, discontinuous gradients (DG) (oocytes n=668), and group B, direct micro swim-up (MSU) (oocytes n=660). We analyzed differences in some key performance indicators. Results Fertilization rates were not statistically different between the DG and MSU groups (76.0% vs. 81.8%, respectively, p=0.248); while significant differences were found in blastulation rates per fertilized oocytes (41.7% vs. 58.5%, p=0.009), blastulation rates per D3 embryos (46.1% vs. 63.7%, p=0.045), and pregnancy rates (25.8% vs. 41.9%, p=0.045). The abortion rate was reduced in the MSU group as compared to DG, but not in a significant manner (12.9% vs. 29.4%, p=0.161). Conclusion The MSU procedure has the advantage of reducing costs, time and mismatches, while ensuring comparable, and in some cases, better results than DG treatments. This technique can therefore be used as an alternative method to other conventional semen treatments.

A new micro swim-up procedure for sperm preparation in ICSI treatments: preliminary microbiological testing.

Objective This study aimed to assess the levels of microbial contamination in semen samples before and after the micro swim-up (MSU) procedure in intra-cytoplasmic sperm injection (ICSI). The new method is an upgrade to the classic wash swim-up procedure. Methods Semen analysis and microbiological tests were carried out before and after the MSU procedure. A total of twenty semen samples were analyzed. Results Pathogens were observed in semen samples only before MSU and never after ICSI. Microbiological tests revealed a large prevalence of gram-positive cocci [Staphylococcus spp. (n=16, 80%) and viridans streptococci (n=10, 50%)]. The results of this study indicate that direct MSU in ICSI improved the ICSI workflow. Conclusion The new workflow is faster and more affordable, and is likely to prevent infection problems that could arise from the normal microbial flora of the semen.