320 SUSCEPTIBILITY OF GOAT SPERM TO DIFFERENT REACTIVE OXYGEN SPECIES

2010 ◽  
Vol 22 (1) ◽  
pp. 316
Author(s):  
R. O. C. Silva ◽  
E. G. A. Perez ◽  
R. P. Cabral ◽  
D. G. Silva ◽  
C. H. C. Viana ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Hydroxyl Radical ◽  
Semen Quality ◽  
Membrane Integrity ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Mitochondrial Potential ◽  
Acrosome Integrity

Semen quality is one of the main limiting factors for the success of artificial insemination in goats. It is well known that reactive oxygen species (ROS) lead to structural and functional damages to sperm, impairing or avoiding fecundation. The understanding of sperm oxidative mechanisms in goats may provide information on possible treatments to improve semen quality and fertility rates. The aim of the present study was to verify the resistance of goat spermatozoa to different reactive oxygen species. Sperm samples from 4 goats were collected using an artificial vagina. Sperm samples were then incubated (1 h, 37°C) with 4 ROS inducer mechanisms: xanthine/xanthine oxidase (produces superoxide anion), hydrogen peroxide (4 mM), ascorbate/ferrous sulfate (4 mM; produces hydroxyl radical), and malondialdehyde (MDA, lipid peroxidation product). Samples were analyzed for mitochondrial activity using the 3,3′ diaminobenzidine stain, for membrane integrity using the eosin/nigrosin staining, for acrosome integrity using the simple stain (fast green/Bengal rose), and for lipid peroxidation by dosing thiobarbituric acid reactive substances (TBARS). Results showed that goat sperm is more sensitive to hydrogen peroxide, when compared to superoxide anion, hydroxyl radical, and MDA, when considering acrosome integrity, membrane integrity, and mitochondrial potential (Table 1). On the other hand, TBARS production was increased in samples submitted to hydroxyl radical incubation. Strong negative correlations were found between sperm samples showing impaired mitochondrial potential and both membrane and acrosome integrity (r = -0.97, P < 0.0001 and r = -0.91, P < 0.0001, respectively). The concentration of TBARS correlated negatively with the percentage of sperm showing intact membranes (r = -0.53, P = 0.06), and the later correlated negatively with sperm showing no mitochondrial activity (r = -0.78, P = 0.0006). Results of the present experiment suggest that goat sperm are extremely susceptible to the attack of hydrogen peroxide, being resistant to other ROS. Therefore, an alternative to improve the use of goat semen in reproductive biotechnologies would be the treatment with catalase or glutathione peroxidase, important hydrogen peroxide scavengers. Table 1.Effect of different ROS on goat sperm The authors thank Nutricell for the media used in this experiment.

34 OVINE SPERM IS HIGHLY SUSCEPTIBLE TO ATTACK BY HYDROGEN PEROXIDE

2010 ◽  
Vol 22 (1) ◽  
pp. 175
Author(s):  
E. G. A. Perez ◽  
M. Nichi ◽  
F. A. Oliveira Neto ◽  
R. O. C. Silva ◽  
A. Dalmazzo ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Lipid Peroxidation ◽  
Hydroxyl Radical ◽  
Ferrous Sulfate ◽  
Present Experiment ◽  
Membrane Integrity ◽  
Oxygen Species ◽  
Mitochondrial Potential ◽  
Ram Sperm

Ram sperm membrane displays a particular lipid composition, especially regarding the high quantity of polyunsaturated cholesterol. This trait improves membrane fluidity; however, the spermatozoa become more susceptible to the attack of reactive oxygen species (ROS), which may lead to structural and functional damage, impairment or even impeded fecundity. The aim of the present experiment was to study the resistance of ovine spermatozoa to different ROS. Sperm samples from 4 rams were collected using an artificial vagina. Sperm samples were then incubated (1 h, 37°C) with four ROS inducer mechanisms: xanthine/xanthine oxidase (produces superoxide anion), hydrogen peroxide (4 mM), ascorbate/ferrous sulfate (4 mM; produces hydroxyl radical), and malondialdehyde (MDA, lipid peroxidation product). Samples were analysed using the 3-3′ diamino benzidine (DAB) stain as an index of mitochondrial activity, the eosin nigrosin stain as an index of membrane integrity; the simple stain (fast green/Bengal rose) as an index of acrosome integrity; and the measurement of thiobarbituric acid reactive substances (TBARS) as an index of lipid peroxidation. Results showed that acrosome and membrane integrity as well as mitochondrial potential were highly impaired by hydrogen peroxide, which was not the case for the other ROS (Table 1). Surprisingly, TBARS production was higher in samples incubated with ascorbate and ferrous sulfate (hydroxyl radical). Furthermore, sperm showing impaired mitochondrial potential were negatively correlated with membrane and acrosome integrities (r = -0.83, P < 0.0001 and r = -0.62, P = 0.01, respectively). Results of the present experiment suggest that semen of rams is extremely susceptible to attack by hydrogen peroxide. However, the mechanism by which this substance impairs sperm quality apparently does not involve oxidative stress, because no increase in TBARS was observed. Despite the necessity of further studies to investigate how hydrogen peroxide negatively influences sperm function, the use of catalase and glutathione peroxidase, important hydrogen peroxide scavengers, appears to be an alternative to improve the quality of ram sperm. Table 1.Effect of different reactive oxygen species in semen of rams The authors thank Nutricell for the media used in the experiment and CAPES for financial support.

75 INFLUENCE OF CRYOPRESERVATION ON THE SUSCEPTIBILITY OF GOAT SPERM AGAINST DIFFERENT REACTIVE OXYGEN SPECIES

2011 ◽  
Vol 23 (1) ◽  
pp. 143 ◽  
Author(s):  
P. A. A. Góes ◽  
M. Nichi ◽  
R. O. C. Silva ◽  
E. G. A. Perez ◽  
A. Dalmazzo ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Lipid Peroxidation ◽  
Hydroxyl Radical ◽  
Dna Fragmentation ◽  
Seminal Plasma ◽  
Semen Quality ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Hydroxyl Radical Scavengers

Semen quality after cryopreservation is one of the main limiting factors for the success of artificial insemination in goats. Previous studies indicate that cryo-injuries may be related to the oxidative stress which is caused by the reactive oxygen species (ROS) and leads to structural and functional damages to the sperm. The understanding of sperm oxidative mechanisms in goats may provide information on possible treatments to improve semen quality post cryopreservation. The aim of the present study was to verify the resistance of cryopreserved goat spermatozoa to different reactive oxygen species. Semen samples from 5 adult goats were collected and cryopreserved (Botubov®, Biotech Ltda.). After thawing, samples were washed twice with PBS and incubated (1 h, 37°C) with 4 ROS inducer mechanisms: xanthine/xanthine oxidase (produces superoxide anion), hydrogen peroxide (4 mM), ascorbate and ferrous sulfate (4 mM; produces hydroxyl radical) with and without the addition of seminal plasma. Samples were analysed for motility using computer-assisted sperm analysis (CASA); the 3–3′ diaminobenzidine stain, as an index of mitochondrial activity; the eosin nigrosin stain, as an index of membrane integrity; the simple stain (Fast green/Bengal rose), as an index of acrosome integrity; sperm chromatin structure assay as an index of DNA fragmentation; and the measurement of thiobarbituric acid reactive substances (TBARS), an index of lipid peroxidation. Statistical analysis was performed using the SAS System for Windows (SAS Institute Inc., Cary, NC, USA; least significant differences test and Spearman correlation; P < 0.05). Results showed that cryopreserved goat sperm after thawing is highly susceptible to the hydroxyl radical. No differences were found on CASA variables between the different ROS. On the other hand, lipid peroxidation and DNA fragmentation were higher for samples treated with hydroxyl radical when compared to samples treated with the other ROS. Furthermore, sperm showing low mitochondrial activity were lower also for samples treated with hydroxyl radical. Negative correlations were found between lipid peroxidation, and most of the variables evaluated by the CASA. A positive correlation was found between the percentage of sperm showing low mitochondrial potential and DNA fragmentation, indicating that impaired mitochondrial activity may be related to an increase on DNA fragmentation. Previous studies indicate that fresh goat semen is highly susceptible to the attack of hydrogen peroxide. We observed that after thawing there is a shift towards a higher susceptibility to the hydroxyl radical. This may indicate that seminal plasma in goats may be an important source of hydroxyl radical scavengers and that, due to the dilution of the seminal plasma with the extender, such antioxidant protection may be impaired. Therefore, an alternative to improve semen quality in cryopreserved goat semen would be the treatment with hydroxyl radical scavengers such as vitamins E and C, reduced glutathione, and other non-enzymatic antioxidants. Thanks to CAPES for financial support.

317 RESISTANCE AGAINST DIFFERENT REACTIVE OXYGEN SPECIES IN BOVINE EPIDIDYMAL SPERM UNDER DISTINCT MATURATION STATUS

2010 ◽  
Vol 22 (1) ◽  
pp. 314
Author(s):  
M. Nichi ◽  
E. G. A. Perez ◽  
C. H. C. Viana ◽  
A. C. Teodoro ◽  
P. A. A. Goes ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Lipid Peroxidation ◽  
Hydroxyl Radical ◽  
Reactive Oxygen ◽  
Lipid Peroxidation Product ◽  
Oxygen Species ◽  
Epididymal Sperm ◽  
Mitochondrial Potential

Oxidative stress is caused by reactive oxygen species (ROS) that may cause structural damage to biomolecules, DNA, lipids, carbohydrates and proteins, as well as other cellular components. Evidence indicates that oxidation products are also deleterious to biological systems. Spermatozoa are particularly susceptible the oxidative stress, mainly due to the reduced cytoplasm and the high content of polyunsaturated fatty acids in its membrane. The mechanisms by which sperm acquire antioxidant capacity are still not completely elucidated. The aim was to study the resistance of sperm derived from different epididymal compartments (caudae and head) to the different ROS and to the lipid peroxidation product malondialdehyde (MDA). Epididymal sperm samples from 4 testicles were collected from the head and caudae epididymides. Sperm samples were then incubated (1 h, 37°C) with 4 ROS inducer mechanisms: xanthine/xanthine oxidase (produces superoxide anion), hydrogen peroxide (4 mM), ascorbate and ferrous sulfate (4 mM; produces hydroxyl radical), and MDA. Samples were analyzed for 3-3′ diaminobenzidine stain, as an index of mitochondrial activity; the eosin nigrosin stain, as an index of membrane integrity; the simple stain (fast green/Bengal rose), as an index of acrosome integrity; and the measurement of thiobarbituric acid reactive substances (TBARS), an index of lipid peroxidation. Statistical analysis was performed using the SAS System for Windows (SAS Institute Inc., Cary, NC, USA; least significant differences test and Pearson correlation). Results showed that immature sperm (head epididymides) were significantly more susceptible to the MDA and to the hydroxyl radical in all studied variables, especially acrosomes, membranes, and mitochondrial potential. Semen derived from the caudae epididymides was more susceptible to the hydrogen peroxide and to the MDA, especially regarding mitochondrial potential. In semen from the epididymal head, a positive correlation was found between TBARS and sperm showing no mitochondrial potential (r = 0.66, P = 0.01). On the other hand, negative correlations were found between TBARS and sperm with damaged acrosome and membrane (r = -0.63, P = 0.01 and r = -0.58, P = 0.02, respectively) in samples collected from the caudae epididymides. The present results suggest that sperm susceptibility to the attack of ROS is different throughout maturation. Although immature sperm are more susceptible to the hydroxyl radical, mature sperm are more susceptible to the hydrogen peroxide. Furthermore, MDA, a product of lipid peroxidation, is also deleterious to the sperm, indicating that once oxidative stress starts, further damage may be caused by their products. The authors thankNutricell for the media used in the experiment andFAPESP for financial support (process #06/05736-1).

Antioxidant Activity of Hizikia fusiformis on Reactive Oxygen Species Scavenging and Lipid Peroxidation Inhibition

2003 ◽  
Vol 9 (5) ◽  
pp. 339-346 ◽  
Author(s):  
Nalin Siriwardhana ◽  
K.-W. Lee ◽  
Y.-J. Jeon ◽  
S.-H. Kim ◽  
J.-W. Haw
Keyword(s):  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Lipid Peroxidation ◽  
Hydroxyl Radical ◽  
Radical Scavenging ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Ros Scavenging ◽  
Hizikia Fusiformis ◽  
Lipid Peroxidation Inhibition

Water and organic extracts (diethyl ether, chloroform, ethyl acetate, acetone, ethanol and methanol) obtained from Hizikia fusiformis were screened on reactive oxygen species (ROS) scavenging assays (1,1-diphenyl-2-picrylhydrazyl (DPPH), superoxide anion, hydrogen peroxide and hydroxyl radical) and lipid peroxidation (inhibition of linoleic acid oxidation) inhibitory assays. Water, methanol and ethanol extracts showed significant ROS radical scavenging activities. Water extracts showed high scavenging activities on hydrogen peroxide (around 76%) and DPPH radicals (around 75%) while it presented a moderate scavenging activity on hydroxyl radicals (around 54%). Comparatively higher ROS scavenging activities were recorded in hydroxyl radical and DPPH scavenging assays. DPPH radical scavenging activities were well correlated with the polyphenolic content. ROS scavenging and lipid peroxidation inhibition activities indicated that H. fusiformis might be a valuable natural antioxidative source containing both water and fatsoluble antioxidative components.

242 PROTECTIVE EFFECT OF VITAMIN E TREATMENT IN HEAT-STRESSED BOS TAURUS BULLS

2013 ◽  
Vol 25 (1) ◽  
pp. 269 ◽  
Author(s):  
V. H. Barnabe ◽  
R. C. Barnabe ◽  
P. Goes ◽  
E. G. A. Perez ◽  
J. D. A. Losano ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Heat Stress ◽  
Vitamin E ◽  
Bos Taurus ◽  
Semen Quality ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Tropical Conditions

Bos taurus bulls, when raised under tropical conditions, are highly susceptible to heat stress, which leads to impaired semen quality, leading to significant economical losses because, in these regions, the reproductive mounting season occurs mainly during the summer. Previous studies have indicated that oxidative stress (i.e. attack by reactive oxygen species) may be the main mechanism of sperm damage in such conditions. Therefore, treatment with antioxidants may be an important alternative to improve semen quality in heat-stressed B. taurus bulls. The objective of the present study was to evaluate whether the treatment with vitamin E, an important antioxidant, could improve sperm quality in insulated bulls. Towards this aim, eight adult Holstein bulls were submitted for semen collection, and the sperm was submitted for motility evaluation by computer-assisted sperm analysis (Ivos, Hamilton Thorne Inc., Beverly, MA, USA), examination of membrane and acrosomal integrity (eosin/nigrosin and fast green/bengal rose stain, respectively), mitochondrial activity (diaminobenzidine stain; full mitochondrial activity or no mitochondrial activity), and sperm susceptibility to oxidative stress (thiobarbituric acid-reactive substances). Bulls were then insulated (testicles covered in a thermal bag for 3 days) and randomly assigned to two treatment groups: no vitamin E (placebo) and vitamin E (subcutaneous injection of 3000 IU of α-tocopherol each of 10 days). Subsequent semen analysis was performed 1 and 60 days after the insulation. Statistical analysis was performed with SAS (SAS Institute Inc., Cary, NC, USA) repeated-measures ANOVA, and significance of P < 0.05 was adopted. No differences were found on any of the variables before insulation. One day after insulation, animals treated with vitamin E showed a lower percentage of static sperm and a higher percentage of motile sperm when compared with animals treated with the placebo (28 and 63% v. 56 and 34%, respectively; P < 0.05). Also at this time, sperm susceptibility to oxidative stress was lower in animals treated with vitamin E (vitamin E: 410 ng/106 sperm; no vitamin E: 1760 ng/106 sperm; P < 0.05). Sixty days after insulation, sperm susceptibility to oxidative stress was still lower in animals treated with vitamin E when compared with the placebo group (1176 and 192 ng/106 sperm, respectively; P < 0.05). However, no differences were found on the other variables. Results indicate that vitamin E, an antioxidant whose main function is protection of the plasma membrane, may be an alternative to avoid the acute deleterious effects of the heat stress in B. taurus bulls raised under tropical conditions. In addition, even with no heat stress involved, vitamin E treatment may provide constant protection, increasing the resistance of the sperm against the reactive oxygen species.

Superoxide, hydroxyl radical, and hydrogen peroxide effects on single-diaphragm fiber contractile apparatus

2001 ◽  
Vol 90 (1) ◽  
pp. 45-54 ◽  
Author(s):  
L. A. Callahan ◽  
Z. W. She ◽  
T. M. Nosek
Keyword(s):  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Hydroxyl Radical ◽  
Reactive Oxygen ◽  
Calcium Sensitivity ◽  
Oxygen Species ◽  
Contractile Apparatus ◽  
Skeletal Muscle Function ◽  
Bathing Medium ◽  
Specific Species

Reactive oxygen species contribute to diaphragm dysfunction in certain pathophysiological conditions (i.e., sepsis and fatigue). However, the precise alterations induced by reactive oxygen species or the specific species that are responsible for the derangements in skeletal muscle function are incompletely understood. In this study, we evaluated the effect of the superoxide anion radical (O2 −·), hydroxyl radical (·OH), and hydrogen peroxide (H2O2) on maximum calcium-activated force (Fmax) and calcium sensitivity of the contractile apparatus in chemically skinned (Triton X-100) single rat diaphragm fibers. O2 −· was generated using the xanthine/xanthine oxidase system; ·OH was generated using 1 mM FeCl2, 1 mM ascorbate, and 1 mM H2O2; and H2O2 was added directly to the bathing medium. Exposure to O2 −· or ·OH significantly decreased Fmax by 14.5% ( P < 0.05) and 43.9% ( P < 0.005), respectively. ·OH had no effect on Ca2+ sensitivity. Neither 10 nor 1,000 μM H2O2 significantly altered Fmax or Ca2+ sensitivity. We conclude that the diaphragm is susceptible to alterations induced by a direct effect of ·OH and O2 −·, but not H2O2, on the contractile proteins, which could, in part, be responsible for prolonged depression in contractility associated with respiratory muscle dysfunction in certain pathophysiological conditions.

scholarly journals Fe-catalyzed sulfide oxidation in hydrothermal plumes is a source of reactive oxygen species to the ocean

2021 ◽  
Vol 118 (40) ◽  
pp. e2026654118
Author(s):  
Timothy J. Shaw ◽  
George W. Luther ◽  
Richard Rosas ◽  
Véronique E. Oldham ◽  
Nicole R. Coffey ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Hydroxyl Radical ◽  
Bottom Water ◽  
Sulfide Oxidation ◽  
Reactive Oxygen ◽  
Global Ocean ◽  
Oxygen Species ◽  
Fenton Chemistry ◽  
Hydrothermal Plumes

Historically, the production of reactive oxygen species (ROS) in the ocean has been attributed to photochemical and biochemical reactions. However, hydrothermal vents emit globally significant inventories of reduced Fe and S species that should react rapidly with oxygen in bottom water and serve as a heretofore unmeasured source of ROS. Here, we show that the Fe-catalyzed oxidation of reduced sulfur species in hydrothermal vent plumes in the deep oceans supported the abiotic formation of ROS at concentrations 20 to 100 times higher than the average for photoproduced ROS in surface waters. ROS (measured as hydrogen peroxide) were determined in hydrothermal plumes and seeps during a series of Alvin dives at the North East Pacific Rise. Hydrogen peroxide inventories in emerging plumes were maintained at levels proportional to the oxygen introduced by mixing with bottom water. Fenton chemistry predicts the production of hydroxyl radical under plume conditions through the reaction of hydrogen peroxide with the abundant reduced Fe in hydrothermal plumes. A model of the hydroxyl radical fate under plume conditions supports the role of plume ROS in the alteration of refractory organic molecules in seawater. The ocean’s volume circulates through hydrothermal plumes on timescales similar to the age of refractory dissolved organic carbon. Thus, plume-generated ROS can initiate reactions that may affect global ocean carbon inventories.

108 EFFECT OF DIMETHYLFORMAMIDE AND METHYLFORMAMIDE ON OXIDATIVE STATUS OF MANGALARGA STALLIONS CRYOPRESERVED SEMEN

2010 ◽  
Vol 22 (1) ◽  
pp. 213
Author(s):  
F. A. Oliveira Neto ◽  
M. Nichi ◽  
E. G. A. Perez ◽  
J. R. C. Gurgel ◽  
G. H. Ferreira ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Plasma Membrane ◽  
Membrane Integrity ◽  
The Other ◽  
Lipid Peroxidation Product ◽  
Oxygen Species ◽  
Mitochondrial Potential ◽  
Plasma Membrane Integrity ◽  
Peroxidation Product ◽  
Acrosome Integrity

Cryopreservation of equine semen has been widely studied by several research groups because of the large breed and individual variation in sperm freezability. A key factor in sperm cryopreservation is the high incidence of oxidative stress, an imbalance between reactive oxygen species (ROS) and antioxidant protection, which impairs sperm functionality by attacking plasma membrane, acrosome, mitochondria, and DNA. In order to study the resistance of equine spermatozoa to different reactive oxygen species (ROS), sperm samples from 4 Mangalarga stallions were collected using an artificial vagina. Samples were cryopreserved in extenders containing dimethylformamide (DMF) or methylformamide (MF). After thawing and washing, sperm samples were then incubated (1 h, 37°C) with 4 ROS inducer mechanisms: xanthine/xanthine oxidase (produces superoxide anion), hydrogen peroxide (4 mM), ascorbate/ferrous sulfate (4 mM; produced hidroxyl radical), and malondialdehyde (MDA, lipid peroxidation product). Samples were evaluated using the 3-3′ diamino benzidine (DAB) stain, as an indicator of mitochondrial activity; the eosin nigrosin staining, to evaluate plasma membrane integrity; the simple stain (fast green/Bengal rose), to assess acrosome integrity; and the measurement of thiobarbituric acid reactive substances (TBARS), a lipid peroxidation product. Statistical analysis was performed using the Student t-test and LSD test. Results showed that sperm mitochondrial potential of frozen-thawed samples in MF was highly susceptible to the attack of hydroxyl radical and hydrogen peroxide. No effect of ROS was observed on membrane and acrosome integrity. On the other hand, samples cryopreserved in DMF showed no differences in susceptibility to ROS. When evaluating the main effects of different extenders, results showed a higher protective effect of the MF extender on acrosome integrity and mitochondrial potential (MF: 12.1 ± 2.2 and 7.8 ± 2.3% v. DMF: 3.4 ± 0.7 and 1.1 ± 0.7%, respectively, P < 0.05). However, a negative effect of MF extender was observed regarding the percentage of sperm showing intact membrane and TBARS content (MF: 2.0 ± 0.8% and 517 ± 115 ng/106 sperm v. DMF: 20.6 ± 1.7% and 118 ± 44 ng/106 sperm, respectively, P < 0.05). A strong negative correlation was found between TBARS and plasma membrane integrity (r = -0.88; P = 0.004) for samples cryopreserved in DMF, whereas a positive correlation was found between TBARS and sperm with full mitochondrial potential (r = 0.73; P = 0.04). Results of the present study indicate that DMF may play a role in the protection of sperm against the attack of ROS. However, such action is apparently limited to the plasma membrane. On the other hand, the MF-supplemented extender exerts an intracellular protection. Therefore, the antioxidant therapy, especially hydrogen peroxide and hydroxyl radical scavengers, may be an alternative to improve the post-thaw quality of MF-supplemented cryopreserved semen in stallions, by increasing extracellular antioxidant capacity. The authors thank Nutricell for financial support and the media used in the present experiment.

232 INFLUENCE OF SEMINAL PLASMA ON THE SUSCEPTIBILITY OF DOG SPERM AGAINST DIFFERENT REACTIVE OXYGEN SPECIES

2011 ◽  
Vol 23 (1) ◽  
pp. 215
Author(s):  
A. Dalmazzo ◽  
P. A. A. Góes ◽  
M. Nichi ◽  
R. O. C. Silva ◽  
J. R. C. Gurgel ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Lipid Peroxidation ◽  
Hydroxyl Radical ◽  
Seminal Plasma ◽  
Superoxide Anion ◽  
The Other ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Acrosome Integrity

Due to the importance of dogs to humans, there is increasing interest in breeders in the use of reproductive biotechnologies. However, most of the biotechnologies would require the removal or dilution of the seminal plasma, which is known to exert both beneficial and deleterious effects on sperm quality. One of the beneficial effects of seminal plasma would be the antioxidant protection because sperm are particularly susceptible to oxidative stress, mainly due to the reduced cytoplasm and the high content of polyunsaturated fatty acids in their membrane. An alternative to overcome the injuries caused by oxidative stress is the antioxidant treatment, which requires the identification of those reactive oxygen species (ROS) that are the most deleterious. The aim of this study was to identify the most harmful ROS to dog semen. Semen samples from 6 adult dogs were collected and centrifuged. Seminal plasma (SP) was removed and samples were incubated (1 h, 37°C) with 4 ROS-inducing mechanisms: xanthine/xanthine oxidase (produces superoxide anion), hydrogen peroxide (4 mM), ascorbate and ferrous sulfate (4 mM; produces hydroxyl radical) alone or with additional SP. Samples were analysed for motility by computer assisted sperm analysis (CASA). The 3-3′ diaminobenzidine stain was used as an index of mitochondrial activity, the eosin nigrosin stain as an index of membrane integrity, the simple stain (fast green/Bengal rose) as an index of acrosome integrity, sperm chromatin structure assay (SCSA) as an index of DNA fragmentation, and measurement of thiobarbituric acid reactive substances (TBARS) as an index of lipid peroxidation. Statistical analysis was performed using the SAS System for Windows (SAS Institute Inc., Cary, NC, USA; least significant differences test and Spearman correlation; P < 0.05). Results showed that dog sperm is differentially modulated depending on the presence of SP. In addition, damage to the different sperm structures depended on the different ROS. Samples incubated with SP showed no differences concerning TBARS (1 233 in SP, 1 260 in Tris; P = 0.99). On the other hand, samples incubated without SP showed higher lipid peroxidation when treated with hydroxyl radical compared with the other ROS. Furthermore, although hydroxyl radical mostly altered mitochondrial activity in samples incubated with SP (DAB IV = 4.3%; P < 0.05 against all other ROS), the most significant ROS in samples incubated without SP was hydrogen peroxide (DAB IV = 4.7%; P < 0.05 against all other ROS). Superoxide anion was less harmful to acrosome integrity in samples incubated with SP and to motility in samples incubated without SP. The present results suggest that seminal plasma may play an important role in the susceptibility of dog sperm to oxidative stress. Moreover, the results indicate that different sperm compartments are susceptible to different ROS. It is concluded that the quality of frozen–thawed dog semen may be improved by treating with a combination of different antioxidants to destroy the chain reaction causing the oxidative stress. FAPESP is acknowledged for financial support.

Sign in / Sign up

Export Citation Format

Share Document