12 QUANTIFICATION OF REACTIVE OXYGEN SPECIES IN BOVINE SEMEN WITH DIFFERENT SEMINAL QUALITY AND WITH EXTENDER CONTAINING ANTIOXIDANT EVALUATED BY ARTIFICIAL INSEMINATION

2010 ◽  
Vol 22 (1) ◽  
pp. 164
Author(s):  
J. C. Borges ◽  
M. R. Silva ◽  
A. C. Tedesco ◽  
D. S. Costa ◽  
C. R. Esper ◽  
...  

The aim this study was to evaluate the effects on artificial insemination (AI) of minimizing oxidative stress by adding an antioxidant into the sperm microenvironment. For that purpose, semen of bulls with different seminal quality was evaluated in nature and after addition of extender with and without antioxidant, followed by freezing and thawing, with respect to thiobarbituric acid reactive substances (TBARS) concentration, total antioxidant capacity (TAC), and AI success. TBARS is an indicator of lipid peroxidation and TAC measures free radicals. Twelve ejaculates from 5 bulls were treated with Tris egg yolk extender (CE) and in half of each ejaculate the extender was supplemented with 200 μL of Trolox (AE). Three of these 5 bulls had high-quality semen (mean 11.9 ± 5.9 total defect, 7.7 ± 3.1 major defect, and 4.2 ± 3.9 minor defect) and 2 animals had poor-quality sperm (mean 31.9 ± 9.7 total defect, 26.3 ± 8.5 major defect, and 5.6 ± 5.0 minor defect) (P < 0.05). For AI, 300 Nelore heifers were divided in 10 groups and the females in each group were inseminated with semen of the same bull and extender treatment. To evaluate oxidative damage, TBARS concentration was measured to indirectly determine malondialdehyde (lipid-peroxidation metabolite) concentration, whereas TAC was measured using myoglobin, as a source of radicals, which interact with a chromogen 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonate) (ABTS), whose cation radical is spectroscopically detectable. The latency phase (lag) in the accumulation of ABTS cation is proportional to the antioxidant concentration. The pregnancy rates 60 days after AI were analyzed by chi-square test, and data of free radical production were evaluated by ANOVA, followed by the Tukey test with significance level of 5%. Reactive oxygen species (ROS) production in nature semen (0.351 nmol of TBARS in 50 × 108 spermatozoa) was lower (P < 0.05) when compared to diluted semen (0.367 nmol of TBARS in 50 × 108 spermatozoa). No difference (P > 0.05) between extenders was observed after thawing. The bulls with poor-quality semen presented total ROS production (0.967) superior (P < 0.05) to that of bulls with good-quality semen (0.753). Although the use of antioxidant in the extender did not decrease ROS and TAC production (P > 0.05), pregnancy rates after AI in bulls with poor quality semen with AE (60.0%) was superior (P < 0.06) to that in bulls with high-quality sperm with AE (56.6%). In conclusion, bulls with poor-quality semen present higher ROS and TAC production, and the use of antioxidant in the extender for cryopreservation with poor-quality bovine semen, in spite of not decreasing ROS and TAC production, improves pregnancy rates. Financial support: FAPESP.

2021 ◽  
Vol 33 (2) ◽  
pp. 121
Author(s):  
S. X. Yang ◽  
G. P. Adams ◽  
E. M. Zwiefelhofer ◽  
K. Rajapaksha ◽  
M. Anzar

Effective semen extenders are those with defined composition that promote sperm longevity. Generation of reactive oxygen species during semen cryopreservation results in sperm membrane lipid peroxidation and reduced longevity. Recent studies demonstrated that small cell permeating dimethyl tyrosine conjugated peptides, such as SS-31, translocate to the mitochondria and scavenge excess reactive oxygen species. The present study was conducted to test the hypothesis that the addition of SS-31 improves the post-thaw quality and fertility potential of bovine semen. The effect of SS-31 was tested in combination with two extenders: (1) conventional tris-egg yolk-glycerol (TEYG, control) and (2) cholesterol-cyclodextrin+tris-glycerol (CC+TG, a defined protein-free extender). Fifteen ejaculates were collected from 5 Black Angus bulls. Ejaculates were diluted to 400×106 spermmL−1 with tris-citric acid buffer, and treated with 0, 50, or 100 µmolmL−1 SS-31 for 15min at 32°C. Semen aliquots were diluted further with TEYG, 0.5mgmL−1 CC+TG, or 1mgmL−1 CC+TG extender to a final concentration of 50×106 spermmL−1. Semen was then cooled to 4°C and frozen in a programmable freezer. Post-thaw sperm motion parameters were evaluated at 0, 2, 4, 6 and 24h with CASA. Semen fertility was determined by fixed-time AI using Hereford-cross cows (n=100). Synchronized cows were inseminated once with semen extended in TEYG, 1mgmL−1 CC+TG, or 1mgmL−1 CC+TG + 100 µmolmL−1 SS-31. Pregnancy was diagnosed by ultrasonography at 27 days post-insemination. Post-thaw sperm motion parameters were compared by ANOVA for repeated-measures, and pregnancy rates were compared using binomial linear mixed-model ANOVA. No differences in sperm motion parameters were detected among SS-31 treatments within extenders. Semen extended in TEYG or 1mgmL−1 CC+TG had greater total and progressive motilities at 0 and 2h post-thaw than semen extended in 0.5mgmL−1 CC+TG (P&lt;0.05). Pregnancy rates after fixed-time insemination did not differ among semen extender groups [TEYG: 15/25 (60%), 1mgmL−1 CC+TG: 19/34 (56%), 100 µmolmL−1 SS-31+1mgmL−1 CC+TG: 21/37 (57%)]. Addition of the cell permeating antioxidant SS-31 did not improve post-thaw semen quality or fertility. The CC+TG extender was as robust as conventional egg yolk extender in protecting bovine sperm during cryopreservation. This research was supported by NSERC Canada, the Government of Saskatchewan, and Agriculture and Agri-Food Canada.


2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Elena Shekhova ◽  
Olaf Kniemeyer ◽  
Axel A. Brakhage

ABSTRACT Drug resistance in fungal pathogens is of incredible importance to global health, yet the mechanisms of drug action remain only loosely defined. Antifungal compounds have been shown to trigger the intracellular accumulation of reactive oxygen species (ROS) in human-pathogenic yeasts, but the source of those ROS remained unknown. In the present study, we examined the role of endogenous ROS for the antifungal activity of the three different antifungal substances itraconazole, terbinafine, and amphotericin B, which all target the fungal cell membrane. All three antifungals had an impact on fungal redox homeostasis by causing increased intracellular ROS production. Interestingly, the elevated ROS levels induced by antifungals were abolished by inhibition of the mitochondrial respiratory complex I with rotenone. Further, evaluation of lipid peroxidation using the thiobarbituric acid assay revealed that rotenone pretreatment decreased ROS-induced lipid peroxidation during incubation of Aspergillus fumigatus with itraconazole and terbinafine. By applying the mitochondrion-specific lipid peroxidation probe MitoPerOx, we also confirmed that ROS are induced in mitochondria and subsequently cause significant oxidation of mitochondrial membrane in the presence of terbinafine and amphotericin B. To summarize, our study suggests that the induction of ROS production contributes to the ability of antifungal compounds to inhibit fungal growth. Moreover, mitochondrial complex I is the main source of deleterious ROS production in A. fumigatus challenged with antifungal compounds.


1996 ◽  
Vol 15 (5) ◽  
pp. 422-427 ◽  
Author(s):  
Y. Shen ◽  
H-M. Shen ◽  
C-Y. Shi ◽  
C-N. Ong

Benzene is myelotoxic and leukemogenic in humans. The mechanisms leading to these effects, however have not been fully elucidated. One of the underlying mechanisms is believed to be the oxidative damage caused by its metabolites. A comparative study was undertaken to examine the relationships between reactive oxygen species (ROS) production, lipid peroxidation and subse quent cytotoxicity induced by five major benzene meta bolites. The generation of ROS by benzene metabolites was demonstrated by the significant and dose-dependent increase of intracellular ROS formation in HL60 human promyelocytic leukemia cells in vitro. 1,4-Benzoquinone (BQ) was found to be the most potent metabolite in induction of ROS formation, followed by 1,2,4-benzene triol (BT) and to a lesser extent, phenol (PH) and trans, trans-muconaldehyde (MD). No significant effect was observed when the cells were treated with trans, trans-muconic acid (MA). The enhancement of ROS production by BQ was effectively inhibited by the addition of catalase, deferoxamine (DFO) and dimethyl sulfoxide (DMSO), but unchanged by superoxide dismutase (SOD), suggest that hydrogen peroxide (H2O2) and hydroxyl radicals (OH.) are the two major forms of ROS involved. The results also demonstrate that the ability of benzene metabolites in enhancing ROS generation is closely correlated to their capacity in causing lipid peroxidation and subsequent cytotoxicity. These findings together with earlier parallel observations on DNA damage suggest that ROS play an important role in the mechanism of carcinogenesis induced by benzene metabolites.


2002 ◽  
Vol 49 (3) ◽  
pp. 775-780 ◽  
Author(s):  
Urszula Maciejewska ◽  
Lidia Polkowska-Kowalczyk ◽  
Ewa Swiezewska ◽  
Anna Szkopinska

The plant Solanum nigrum treated with the pathogen Phytophthora infestans-derived elicitor responded by elevated reactive oxygen species (ROS) production, lipid peroxidation and lipoxygenase (EC 1.13.11.12) activity in comparison with control plants indicating that oxidative stress took place. We demonstrate that these events are accompanied by a significant increase in plastoquinone (PQ) level. It is postulated that PQ may be associated with mechanisms maintaining a tightly controlled balance between the accumulation of ROS and antioxidant activity that determines the full expression of effective defence.


Plants ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1586
Author(s):  
Svetlana Veselova ◽  
Tatyana Nuzhnaya ◽  
Guzel Burkhanova ◽  
Sergey Rumyantsev ◽  
Igor Maksimov

Reactive oxygen species (ROS) play a central role in plant immune responses. The most important virulence factors of the Stagonospora nodorum Berk. are multiple fungal necrotrophic effectors (NEs) (SnTox) that affect the redox-status and cause necrosis and/or chlorosis in wheat lines possessing dominant susceptibility genes (Snn). However, the effect of NEs on ROS generation at the early stages of infection has not been studied. We studied the early stage of infection of various wheat genotypes with S nodorum isolates -Sn4VD, SnB, and Sn9MN, carrying a different set of NE genes. Our results indicate that all three NEs of SnToxA, SnTox1, SnTox3 significantly contributed to cause disease, and the virulence of the isolates depended on their differential expression in plants (Triticum aestivum L.). The Tsn1–SnToxA, Snn1–SnTox1and Snn3–SnTox3 interactions played an important role in inhibition ROS production at the initial stage of infection. The Snn3–SnTox3 inhibited ROS production in wheat by affecting NADPH-oxidases, peroxidases, superoxide dismutase and catalase. The Tsn1–SnToxA inhibited ROS production in wheat by affecting peroxidases and catalase. The Snn1–SnTox1 inhibited the production of ROS in wheat by mainly affecting a peroxidase. Collectively, these results show that the inverse gene-for gene interactions between effector of pathogen and product of host sensitivity gene suppress the host’s own PAMP-triggered immunity pathway, resulting in NE-triggered susceptibility (NETS). These results are fundamentally changing our understanding of the development of this economical important wheat disease.


2021 ◽  
Author(s):  
Biz R. Turnell ◽  
Luisa Kumpitsch ◽  
Klaus Reinhardt

AbstractSperm aging is accelerated by the buildup of reactive oxygen species (ROS), which cause oxidative damage to various cellular components. Aging can be slowed by limiting the production of mitochondrial ROS and by increasing the production of antioxidants, both of which can be generated in the sperm cell itself or in the surrounding somatic tissues of the male and female reproductive tracts. However, few studies have compared the separate contributions of ROS production and ROS scavenging to sperm aging, or to cellular aging in general. We measured reproductive fitness in two lines of Drosophila melanogaster genetically engineered to (1) produce fewer ROS via expression of alternative oxidase (AOX), an alternative respiratory pathway; or (2) scavenge fewer ROS due to a loss-of-function mutation in the antioxidant gene dj-1β. Wild-type females mated to AOX males had increased fecundity and longer fertility durations, consistent with slower aging in AOX sperm. Contrary to expectations, fitness was not reduced in wild-type females mated to dj-1β males. Fecundity and fertility duration were increased in AOX and decreased in dj-1β females, indicating that female ROS levels may affect aging rates in stored sperm and/or eggs. Finally, we found evidence that accelerated aging in dj-1β sperm may have selected for more frequent mating. Our results help to clarify the relative roles of ROS production and ROS scavenging in the male and female reproductive systems.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
V. Jaenen ◽  
S. Fraguas ◽  
K. Bijnens ◽  
M. Heleven ◽  
T. Artois ◽  
...  

AbstractDespite extensive research on molecular pathways controlling the process of regeneration in model organisms, little is known about the actual initiation signals necessary to induce regeneration. Recently, the activation of ERK signaling has been shown to be required to initiate regeneration in planarians. However, how ERK signaling is activated remains unknown. Reactive Oxygen Species (ROS) are well-known early signals necessary for regeneration in several models, including planarians. Still, the probable interplay between ROS and MAPK/ERK has not yet been described. Here, by interfering with major mediators (ROS, EGFR and MAPK/ERK), we were able to identify wound-induced ROS, and specifically H2O2, as upstream cues in the activation of regeneration. Our data demonstrate new relationships between regeneration-related ROS production and MAPK/ERK activation at the earliest regeneration stages, as well as the involvement of the EGFR-signaling pathway. Our results suggest that (1) ROS and/or H2O2 have the potential to rescue regeneration after MEK-inhibition, either by H2O2-treatment or light therapy, (2) ROS and/or H2O2 are required for the activation of MAPK/ERK signaling pathway, (3) the EGFR pathway can mediate ROS production and the activation of MAPK/ERK during planarian regeneration.


2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Biz R. Turnell ◽  
Luisa Kumpitsch ◽  
Anne-Cécile Ribou ◽  
Klaus Reinhardt

Abstract Objective Sperm ageing has major evolutionary implications but has received comparatively little attention. Ageing in sperm and other cells is driven largely by oxidative damage from reactive oxygen species (ROS) generated by the mitochondria. Rates of organismal ageing differ across species and are theorized to be linked to somatic ROS levels. However, it is unknown whether sperm ageing rates are correlated with organismal ageing rates. Here, we investigate this question by comparing sperm ROS production in four lines of Drosophila melanogaster that have previously been shown to differ in somatic mitochondrial ROS production, including two commonly used wild-type lines and two lines with genetic modifications standardly used in ageing research. Results Somatic ROS production was previously shown to be lower in wild-type Oregon-R than in wild-type Dahomey flies; decreased by the expression of alternative oxidase (AOX), a protein that shortens the electron transport chain; and increased by a loss-of-function mutation in dj-1β, a gene involved in ROS scavenging. Contrary to predictions, we found no differences among these four lines in the rate of sperm ROS production. We discuss the implications of our results, the limitations of our study, and possible directions for future research.


Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 233
Author(s):  
Tasuku Konno ◽  
Eduardo Pinho Melo ◽  
Joseph E. Chambers ◽  
Edward Avezov

Reactive oxygen species (ROS) are produced continuously throughout the cell as products of various redox reactions. Yet these products function as important signal messengers, acting through oxidation of specific target factors. Whilst excess ROS production has the potential to induce oxidative stress, physiological roles of ROS are supported by a spatiotemporal equilibrium between ROS producers and scavengers such as antioxidative enzymes. In the endoplasmic reticulum (ER), hydrogen peroxide (H2O2), a non-radical ROS, is produced through the process of oxidative folding. Utilisation and dysregulation of H2O2, in particular that generated in the ER, affects not only cellular homeostasis but also the longevity of organisms. ROS dysregulation has been implicated in various pathologies including dementia and other neurodegenerative diseases, sanctioning a field of research that strives to better understand cell-intrinsic ROS production. Here we review the organelle-specific ROS-generating and consuming pathways, providing evidence that the ER is a major contributing source of potentially pathologic ROS.


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