The measurement of reactive oxygen species in human neat semen and in suspended spermatozoa: a comparison

BACKGROUND: It is generally accepted that oxidative stress is an important factor in male infertility because it may impair the physiological function of spermatozoa at the molecular level. Nevertheless, although several approaches have been reported, the imbalance between production of reactive oxygen species (ROS) and activity of the antioxidant defense system in semen is difficult to investigate and remains poorly understood. METHODS: This study compares measurement of ROS production in neat semen and in washed spermatozoa obtained from the same ejaculate, and suspended in phosphate buffered saline using exactly the same luminol-mediated chemiluminescence method. Ninety one samples were obtained from males of infertile couples and 34 from volunteers with proven fertility. RESULTS: As expected, ROS levels were markedly lower in neat semen than in washed spermatozoa suspensions where seminal plasma with its potent antioxidant capacity was removed. In the cases of both neat semen and washed spermatozoa, ROS production was lowest in samples from normozoospermic males and highest in samples containing more than half million peroxidase-positive leukocytes per milliliter. For all samples, there was a significant positive correlation between ROS production by neat semen and that by washed spermatozoa suspension. CONCLUSION: Measurement of ROS production in neat semen better reflects actual oxidative status because it detects only the overproduction of ROS which are not effectively scavenged by antioxidant capacity of seminal fluid. The results of our study show a good commutability of both measurements for identification of semen samples with high ROS production. The measurement in neat semen is even less time consuming and therefore easier to implement into laboratory routine

iTAP, a novel iRhom interactor, controls TNF secretion by policing the stability of iRhom/TACE

The apical inflammatory cytokine TNF regulates numerous important biological processes including inflammation and cell death, and drives inflammatory diseases. TNF secretion requires TACE (also called ADAM17), which cleaves TNF from its transmembrane tether. The trafficking of TACE to the cell surface, and stimulation of its proteolytic activity, depends on membrane proteins, called iRhoms. To delineate how the TNF/TACE/iRhom axis is regulated, we performed an immunoprecipitation/mass spectrometry screen to identify iRhom-binding proteins. This identified a novel protein, that we name iTAP (iRhom Tail-Associated Protein) that binds to iRhoms, enhancing the cell surface stability of iRhoms and TACE, preventing their degradation in lysosomes. Depleting iTAP in primary human macrophages profoundly impaired TNF production and tissues from iTAP KO mice exhibit a pronounced depletion in active TACE levels. Our work identifies iTAP as a physiological regulator of TNF signalling and a novel target for the control of inflammation.info:eu-repo/semantics/publishedVersio

Evaluation of day one embryo quality and IVF outcome – a comparison of two scoring systems

<p>Abstract</p> <p>Background</p> <p>The aim of our retrospective study was to compare the clinical usefulness of two non-invasive embryo scoring systems based either on a simplified pronuclear morphology of the zygote or on early cleavage rate, as well as their combination, for the selection of embryos with the best implantation potential in embryo transfer (ET).</p> <p>Methods</p> <p>Over a period of five years, the quality of 2708 embryos from 364 IVF cycles in women under the age of 39 years was assessed using these scoring systems in a university assisted reproduction centre. ET was always performed on day 3 of cultivation. The outcome of ETs of 702 embryos scored in the respective systems or their combination was retrospectively analyzed in terms of biochemical (bPR) and clinical pregnancy rates (cPR) and implantation rate (IR). Mann-Whitney U test and t-test for differences between relative values were used, p < 0.05 was considered statistically significant.</p> <p>Results</p> <p>There was no difference in outcome parameters in 109 cycles where only Pattern "0" zygotes, according to our simplified pronuclear morphology classification, were transferred and 140 cycles where only "other" pattern zygotes were transferred, regardless of their cleavage rate. On the contrary, significantly greater cPR and IR (p = 0.003 and p = 0.006, respectively) were achieved in 120 cycles where only early cleavage (EC) embryos were transferred compared with 152 cycles where only non early cleavage (NEC) embryos were transferred regardless of their pronuclear morphology. The best outcome in terms of cPR (56%) and IR (43%) was found in 50 cycles when Pattern "0" and EC embryos only were used for transfer.</p> <p>Conclusion</p> <p>The results indicate that early cleavage is a better independent marker of implantation potential than zygote morphology. The best outcome can be achieved if both embryo scoring systems are used jointly and the embryo is classified as EC and Pattern "0".</p

The measurement of reactive oxygen species in human neat semen and in suspended spermatozoa: a comparison

Abstract Background It is generally accepted that oxidative stress is an important factor in male infertility because it may impair the physiological function of spermatozoa at the molecular level. Nevertheless, although several approaches have been reported, the imbalance between production of reactive oxygen species (ROS) and activity of the antioxidant defense system in semen is difficult to investigate and remains poorly understood. Methods This study compares measurement of ROS production in neat semen and in washed spermatozoa obtained from the same ejaculate, and suspended in phosphate buffered saline using exactly the same luminol-mediated chemiluminescence method. Ninety one samples were obtained from males of infertile couples and 34 from volunteers with proven fertility. Results As expected, ROS levels were markedly lower in neat semen than in washed spermatozoa suspensions where seminal plasma with its potent antioxidant capacity was removed. In the cases of both neat semen and washed spermatozoa, ROS production was lowest in samples from normozoospermic males and highest in samples containing more than half million peroxidase-positive leukocytes per milliliter. For all samples, there was a significant positive correlation between ROS production by neat semen and that by washed spermatozoa suspension. Conclusion Measurement of ROS production in neat semen better reflects actual oxidative status because it detects only the overproduction of ROS which are not effectively scavenged by antioxidant capacity of seminal fluid. The results of our study show a good commutability of both measurements for identification of semen samples with high ROS production. The measurement in neat semen is even less time consuming and therefore easier to implement into laboratory routine.</p

iTAP, a novel iRhom interactor, controls TNF secretion by policing the stability of iRhom/TACE

The apical inflammatory cytokine TNF regulates numerous important biological processes including inflammation and cell death, and drives inflammatory diseases. TNF secretion requires TACE (also called ADAM17), which cleaves TNF from its transmembrane tether. The trafficking of TACE to the cell surface, and stimulation of its proteolytic activity, depends on membrane proteins, called iRhoms. To delineate how the TNF/TACE/iRhom axis is regulated, we performed an immunoprecipitation/mass spectrometry screen to identify iRhom-binding proteins. This identified a novel protein, that we name iTAP (iRhom Tail-Associated Protein) that binds to iRhoms, enhancing the cell surface stability of iRhoms and TACE, preventing their degradation in lysosomes. Depleting iTAP in primary human macrophages profoundly impaired TNF production and tissues from iTAP KO mice exhibit a pronounced depletion in active TACE levels. Our work identifies iTAP as a physiological regulator of TNF signalling and a novel target for the control of inflammation