Total antioxidant capacity of human seminal plasma

Although the importance of seminal plasma in the protection of spermatozoa against reactive oxygen species is well known, only a few studies have investigated its antioxidative properties and the possible relationship between infertility and plasmatic antioxidant defences. The aim of the present study was to assess the status of the total non-enzymatic antioxidant defences of human seminal plasma. Semen samples were obtained from 101 patients consulting for infertility and 15 fertile donors. A total reactive antioxidant potential (TRAP) in seminal plasma was determined by luminol-chemiluminescence. The relationship of seminal TRAP values to lipid peroxidation was also evaluated. Our results show that semen samples from fertile controls show total antioxidant capacity at higher frequency and value than equivalent samples from suspected subfertile men. This fact as well as the inverse relationship observed between antioxidant capacity and lipid peroxidation potential strongly suggest th

HUMAN SPERMATOZOA SELECTED BY PERCOLL GRADIENT OR SWIM-UP ARE EQUALLY CAPABLE OF BINDING TO THE HUMAN ZONA-PELLUCIDA AND UNDERGOING THE ACROSOME REACTION

Several techniques have been used for selecting motile spermatozoa including Percoll and albumin gradients, swim-up, and glass wool filtration. A high yield of motile spermatozoa as well as an enhancement of motility are the most desirable features of a practical method. An equally important consideration is whether or not these techniques select functionally normal spermatozoa. In this study we have compared two methods for separation of motile cells, swim-up and Percoll gradient. Normal semen samples from 12 different men were used in this study. Each sample was simultaneously processed by swim-up and Percoll gradient using modified Tyrode's medium. After the sperm concentration was adjusted to 1 x 10(7) spermatozoa/ml, the suspensions were incubated at 37-degrees-C, 5% CO2 in air. In each suspension the percentage of sperm recovery, percentage of motile spermatozoa, percentage of acrosome reacted spermatozoa (either spontaneously or stimulated with human follicular fluid), percentage of zona-free hamster oocytes penetrated, and number of spermatozoa bound to the human zona pellucida were determined. The results obtained indicated that the percentage of sperm recovery was higher with the Percoll gradient than with the swim-up procedure (P < 0.001). However, no significant differences were found between these two sperm populations in the percentage of motile cells, in the percentage of acrosome reacted spermatozoa, and in the percentage of zona-free hamster oocytes penetrated. In addition, the number of spermatozoa bound per zona pellucida was similar for spermatozoa selected by Percoll or swim-up. We conclude that there were no functional differences between the spermatozoa selected by either method

Preimplantation embryotoxicity after mouse embryo exposition to reactive oxygen species

Exposure of either gametes or embryos to conditions and/or factors that generate oxidative stress has been associated with impaired early embryogenesis. The effects of reactive oxygen species (ROS) on mouse preimplantation development, depending of the ROS-concentration and time of exposition, were studied. Two-cell embryos were incubated with 5, 10, 25 and 50 μM of hydrogen peroxide (H2O2) for 30 and 60 minutes of exposition and allowed to develop for 72 h to study the quality of development. The incubation with 50 μM H2O2 for 30 or 60 minutes, strongly inhibited the 2-cell embryo development as compared to the control (p&lt;0.001). Twenty-five μM H2O2 produced inhibition of blastocyst formation (p&lt;0.001) and 10 μM H2O2 significantly decreased the percentages of expanded and hatched blastocysts, which resulted morphologically altered (p&lt;0.05 and p&lt;0.01, respectively). The higher H2O2 concentrations were able to elicit necrotic morphology in the 2-cell arrested embryos, while 10 μM H2O 2 induced moderate damage with the arrested embryos partially fragmented. In conclusion, important causes for defective preimplantation development and for early embryo losses may be due to oxidative stress because early mouse embryos exposed to ROS for short times arrested at the first cellular cycle (2-cell) and/or impaired embryo differentiation and morphogenesis, being these effects ROS-concentration-dependent.Fil:Cebral, E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

The conversion of dehydroepiandrosterone into androst-5-ene-3β,17β-diol (androstenediol) is increased in endometria from untreated women with polycystic ovarian syndrome

The changes in endometrial homeostasis found in women with polycystic ovarian syndrome (PCOS) could be associated with alterations in the intracrine metabolism of steroid hormones. The uptake of dehydroepiandrosterone-sulphate (DHEA-S), precursor of the intracrine pathway, is achieved by transporters, such as organic anion transporter polypeptides (OATPs), and molecules with oestrogenic activity, such as androst-5-ene-3 ,17 -diol (androstenediol), can be generated. We aimed to determine androstenediol generation and the expression of OATPs in human endometria throughout the menstrual cycle and in endometria from PCOS women. Endometrial samples were obtained from control women in the proliferative phase (control endometria (CEp), n = 7), secretory phase (CEs, n = 7), and from PCOS patients (PCOSEp, n = 7). The mRNA levels of OATP-B, OATP-D and OATP-E were measured by reverse transcriptase polymerase chain reaction (RT-PCR) and protein levels of OATP-E by immunofluorescence; 3 -hydroxysteroid dehydrogenase (HSD) by immunohistochemistry/Western blot; the metabolism of DHEA to androstenediol was evaluated by thin layer chromatography–high-performance liquid chromatography (TLC–HPLC). Lower levels of OATP-E transcript were obtained in PCOSEp (p < 0.05) compared with CEp, while OATP-E protein levels (p < 0.05) and DHEA conversion to androstenediol (p < 0.01) were higher in PCOSEp. Lower 3 -(hydroxysteroid dehydrogenase) HSD protein levels were found in PCOSEp (p < 0.05) (Western blot, immunohistochemistry). These results reveal a higher capacity of the endometria from PCOS women to metabolise DHEA to androstenediol, which, coupled with the high oestrogen sensitivity previously found in these endometria, may account for the increase in cell proliferation in PCOSEp already reported.This study was supported by grant no. 1095127 from the Fondo Nacional de Desarrollo Científico y Tecnológico, Chile