Significance of Reactive Oxygen Species and Antioxidants in Defining the Efficacy of Sperm Preparation Techniques

doi:10.1002/J.1939-4640.1988.TB01067.X

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Significance of Reactive Oxygen Species and Antioxidants in Defining the Efficacy of Sperm Preparation Techniques

Journal Article•DOI•

Significance of Reactive Oxygen Species and Antioxidants in Defining the Efficacy of Sperm Preparation Techniques

12 Nov 1988-Journal of Andrology (Blackwell Publishing Ltd)-Vol. 9, Iss: 6, pp 367-376

TL;DR: Assessment of the ability of the antioxidants, butylated hydroxytoluene, and vitamin E, to curtail the peroxidative damage inflicted by cells in response to centrifugation revealed a significant improvement of sperm function in the presence of vitamin E.

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Abstract: The mechanisms responsible for mediating the influence of sperm preparation protocols on human sperm function have been investigated. Techniques that involved the separation of motile spermatozoa prior to centrifugation were found to yield sperm suspensions of highest quality. If the spermatozoa were centrifuged prior to isolation of the motile cells, sperm function was impaired. The detrimental effects of centrifugation were associated with a sudden burst of reactive oxygen species production by a discrete subpopulation of cells (characterized by significantly diminished motility and fertilizing capacity) that could be separated from normal functional spermatozoa on Percoll gradients. If unfractionated sperm suspensions were subjected to centrifugation, the reactive oxygen species generated by this subpopulation impaired the functional competence of normal spermatozoa in the same suspension. Assessment of the ability of the antioxidants, butylated hydroxytoluene, and vitamin E, to curtail the peroxidative damage inflicted by such cells in response to centrifugation revealed a significant improvement of sperm function in the presence of vitamin E.

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Citations

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WHO laboratory manual for the examination and processing of human semen

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T. Cooper, TG Cooper, J. Aitken, J Auger, G Noonan, J Backer - Show less +2 more

01 Jan 2010

2,199 citations

Journal Article•DOI•

Generation of reactive oxygen species, lipid peroxidation, and human sperm function.

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Robert John Aitken, J. S. Clarkson, S Fishel¹•Institutions (1)

University of Nottingham¹

01 Jul 1989-Biology of Reproduction

TL;DR: Results are consistent with a causative role for lipid peroxidation in the etiology of defective sperm function and also suggest a possible physiological role for the reactive oxygen species generated by human spermatozoa in mediating sperm-zona interaction.

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Abstract: Recent studies have demonstrated that human spermatozoa are capable of generating reactive oxygen species and that this activity is significantly accelerated in cases of defective sperm function. In view of the pivotal role played by lipid peroxidation in mediating free radical damage to cells, we have examined the relationships between reactive oxygen species production, lipid peroxidation, and the functional competence of human spermatozoa. Using malondialdehyde production in the presence of ferrous ion promoter as an index of lipid peroxidation, we have shown that lipid peroxidation is significantly accelerated in populations of defective spermatozoa exhibiting high levels of reactive oxygen species production or in normal cells stimulated to produce oxygen radicals by the ionophore, A23187. The functional consequences of lipid peroxidation included a dose-dependent reduction in the ability of human spermatozoa to exhibit sperm oocyte-fusion, which could be reversed by the inclusion of a chain-breaking antioxidant, alpha-tocopherol. Low levels of lipid peroxidation also had a slight enhancing effect on the generation of reactive oxygen species in response to ionophore, without influencing the steady-state activity. At higher levels of lipid peroxidation, both the basal level of reactive oxygen species production and the response to A23187 were significantly diminished. In contrast, lipid peroxidation had a highly significant, enhancing effect on the ability of human spermatozoa to bind to both homologous and heterologous zonae pellucidae via mechanisms that could again be reversed by alpha-tocopherol. These results are consistent with a causative role for lipid peroxidation in the etiology of defective sperm function and also suggest a possible physiological role for the reactive oxygen species generated by human spermatozoa in mediating sperm-zona interaction.

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1,171 citations

Journal Article•DOI•

Sperm chromatin structure assay: its clinical use for detecting sperm DNA fragmentation in male infertility and comparisons with other techniques.

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Donald P. Evenson¹, K.L. Larson¹, Lorna K. Jost¹•Institutions (1)

South Dakota State University¹

02 Jan 2002-Journal of Andrology

1,019 citations

Journal Article•DOI•

Role of reactive oxygen species in male infertility.

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Rakesh Sharma¹, Ashok Agarwal¹•Institutions (1)

Cleveland Clinic¹

01 Dec 1996-Urology

TL;DR: From the current data it appears that no single adjuvant will be able to enhance the fertilizing capacity of sperm in infertile men, and a combination of the possible strategies that are not toxic at the dosage used would be a feasible approach.

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908 citations

Journal Article•DOI•

Relative Impact of Oxidative Stress on the Functional Competence and Genomic Integrity of Human Spermatozoa

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Robert John Aitken, E L Gordon, Harkiss D, J.P. Twigg, P A Milne, Jennings Z, D.S. Irvine - Show less +3 more

01 Nov 1998-Biology of Reproduction

TL;DR: Human spermatozoa exposed to increasing levels of oxidative stress achieved through the stimulation of endogenous oxidant generation with NADPH or direct exposure to hydrogen peroxide exhibited significantly elevated levels of DNA damage and yet continued to express an enhanced capacity for sperm-oocyte fusion.

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Abstract: Reactive oxygen metabolites are known to disrupt sperm-oocyte fusion, sperm movement, and DNA integrity; however, the relative sensitivities of these elements to oxidative stress are unknown. In this study these factors were assessed in human spermatozoa exposed to increasing levels of oxidative stress achieved through the stimulation of endogenous oxidant generation with NADPH or direct exposure to hydrogen peroxide. At low levels of oxidative stress, DNA fragmentation was significantly reduced while the rates of sperm-oocyte fusion were significantly enhanced. As the level of oxidative stress increased, the spermatozoa exhibited significantly elevated levels of DNA damage (p < 0.001) and yet continued to express an enhanced capacity for sperm-oocyte fusion. At the highest levels of oxidative stress, extremely high rates of DNA fragmentation were observed but the spermatozoa exhibited a parallel loss in their capacities for movement and oocyte fusion. These studies emphasize how redox mechanisms can either enhance or disrupt the functional and genomic integrity of human spermatozoa depending on the intensity of the oxidative stimulus. Because these qualities are affected at different rates, spermatozoa exhibiting significant DNA damage are still capable of fertilizing the oocyte. These results may have long-term implications for the safety of assisted conception procedures in cases associated with oxidative stress.

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759 citations

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References

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Open Access

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The culture of mouse embryos in vitro.

[...]

J D Biggers, W K Whitten, D G Whittingham

01 Jan 1971

1,080 citations

Journal Article•DOI•

Cellular basis of defective sperm function and its association with the genesis of reactive oxygen species by human spermatozoa.

[...]

Robert John Aitken, J. S. Clarkson

01 Nov 1987-Reproduction

TL;DR: Studies with scavengers of reactive oxygen species revealed that, while reagents directed against singlet oxygen and the hydroxyl radical were without effect, cytochrome C reduced the response to A23187 by about 50%, suggesting that the superoxide anion radical is a major product of the activated human spermatozoon.

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Abstract: Addition of the divalent cation ionophore, A23187, to washed populations of human spermatozoa resulted in a sudden burst of production of reactive oxygen species which peaked within 3-5 min. This activity was dependent upon the presence of calcium in the external medium and was unaffected by the mitochondrial inhibitors, oligomycin, antimycin and rotenone. Studies with scavengers of reactive oxygen species revealed that, while reagents directed against singlet oxygen and the hydroxyl radical were without effect, cytochrome C reduced the response to A23187 by about 50%, suggesting that the superoxide anion radical is a major product of the activated human spermatozoon. The clinical implications of these studies stem from the considerable variation observed between individuals in the levels of reactive oxygen species produced by the spermatozoa. This variability was shown to be inversely related to the ability of the spermatozoa to exhibit sperm-oocyte fusion on exposure to A23187; defective samples exhibited a basal level of reactive oxygen species production which was 40 times that observed with normal functional cells.

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961 citations

"Significance of Reactive Oxygen Spe..." refers background or methods or result in this paper

...This inverse relationship be- tween the functional competence of human sperma- tozoa and their capacity for generating reactive oxygen species is in keeping with our previous studies concerning the biochemical basis of defective sperm function (Aitken and Clarkson, 1987a, 1987b )....
[...]
...…generated by the spermatozoa in response to centrifugation should be able to initiate lipid peroxidation and, as a conse- quence of the resulting loss of membrane fluidity and integrity, bring about the observed decline in sperm function (Jones et al, 1979; Aitken and Clarkson, 1987a, 1987b) ....
[...]
...In a recent analysis of the biochemical basis for defective sperm function (Aitken and Clarkson, 1987a), evidence was provided for an association between the excess generation of reactive oxygen species by human spermatozoa and the impaired capacity of these cells for fertilization (Aitken and…...
[...]
...The generation of reactive oxygen species by human spermatozoa was determined using the luminescent probe, luminol, as described by Aitken and Clarkson (1987a)....
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...The reactive oxygen species involved are not of mito- chondrial origin, but appear to be generated by a membrane-bound NADPH oxidase, the activity of which is regulated by intracellular calcium and protein phosphorylations dependent upon protein kinase C (Aitken and Clarkson, 1987a, 1987b)....
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Journal Article•DOI•

Spontaneous lipid peroxidation and production of hydrogen peroxide and superoxide in human spermatozoa. Superoxide dismutase as major enzyme protectant against oxygen toxicity.

[...]

Juan G. Alvarez¹, Joseph C. Touchstone¹, Luis Blasco¹, Bayard T. Storey¹•Institutions (1)

University of Pennsylvania¹

10 Sep 1987-Journal of Andrology

TL;DR: The results suggest that superoxide dismutase plays the major role in protecting human spermatozoa against lipid peroxidation, and the superoxide Dismutase activity of a fresh sperm sample appears to be a good predictor of the lifetime (up to the complete loss of motility) of that particular sample, and so may prove useful in semen analysis.

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Abstract: Spontaneous lipid peroxidation in washed human spermatozoa was induced by aerobic incubation at 32 C and measured by malonaldehyde production; loss of motility during the incubation was determined simultaneously. Malonaldehyde production at the point of complete loss of motility, defined as the lipoperoxidative lethal endpoint (LLE), was 0.10 +/- 0.03 nmol/10(8) cells (mean +/- SD, n = 40), and was independent of the time to complete loss of motility. Human spermatozoa produced both H2O2 and O2-. during aerobic incubation. Inhibition of superoxide dismutase in these cells with KCN showed that all the H2O2 production is due to action of the dismutase. The superoxide dismutase activity of individual human sperm samples varied between 1 and 10 U/10(8) cells, variations between samples from a single donor being nearly as great as those between different donors. The time to complete motility loss (tL) showed equal variation of 1 to 10 hours among samples. The rate of spontaneous lipid peroxidation, calculated as LLE/tL, for a given sperm sample and the superoxide dismutase activity of the same sample, determined prior to aerobic incubation, gave a good linear correlation (r = 0.97). Glutathione reductase, glutathione peroxidase, and glutathione were found to be present in human spermatozoa, but showed little variation among samples. These results suggest that superoxide dismutase plays the major role in protecting human spermatozoa against lipid peroxidation. In addition, the superoxide dismutase activity of a fresh sperm sample appears to be a good predictor of the lifetime (up to the complete loss of motility) of that particular sample, and so may prove useful in semen analysis.

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913 citations

"Significance of Reactive Oxygen Spe..." refers background in this paper

...Although superoxide dismutase has been shown to have an important protective role (Alvarez et al, 1987), the combination of superoxide anion and hy- drogen peroxide should, if catalytic amounts of iron are available, initiate the Haber-Weiss reaction to generate the extremely reactive hydroxyl…...
[...]
...The primary product of the sperm NADPH- oxidase system appears to be superoxide anion, a portion of which becomes converted to hydrogen peroxide through the action of superoxide dismutase (Mennella and Jones, 1980; Alvarez et al, 1987)....
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Journal Article•DOI•

The Use of Zona-Free Animal Ova as a Test-System for the Assessment of the Fertilizing Capacity of Human Spermatozoa

[...]

Ryuzo Yanagimachi¹, H. Yanagimachi¹, Rogers Bj¹•Institutions (1)

University of Hawaii¹

01 Nov 1976-Biology of Reproduction

TL;DR: The results suggest that zona-free hamster ova can be substituted for human ova in the preliminary assessment of the fertilizing capacity of human spermatozoa.

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Abstract: The zona-free hamster ovum was evaluated as a substitute for human o va in the assessment of the fertilizing capacity of human spermatozoa. Zona-intact ova completely resisted sperm penetration. Using nonpreincu bated spermatozoa sperm penetration of zona-free ova began 4-5 hours after insemination. However when spermatozoa were preincubated in a modified Krebs-Ringer solution for 4 hours sperm penetration began within 1 hour. There is some evidence that this is associated with the completion of sperm capacitation and the acrosome reaction. The results suggest that zona-free hamster ova can be substituted for human ova in the preliminary assessment of the fertilizing capacity of human spermatozoa.

...read moreread less

871 citations

Journal Article•DOI•

Peroxidative Breakdown of Phospholipids in Human Spermatozoa, Spermicidal Properties of Fatty Acid Peroxides, and Protective Action of Seminal Plasma

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Roy Jones¹, Thaddeus Mann¹, Richard J. Sherins¹•Institutions (1)

National Institutes of Health¹

01 May 1979-Fertility and Sterility

TL;DR: The antiperoxidant factor present in human seminal plasma effectively counteracts the toxic effect of exogenous peroxidized fatty acids upon human spermatozoa, but is unable to restore motility lost by lipid peroxide action.

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749 citations

"Significance of Reactive Oxygen Spe..." refers background in this paper

...The human spermatozoon is particularly susceptible to lipid peroxidation, since the sperm plasma mem- brane is rich in unsaturated fatty acids, particularly docosohexanoic acid (Jones et al, 1979)....
[...]
...…generated by the spermatozoa in response to centrifugation should be able to initiate lipid peroxidation and, as a conse- quence of the resulting loss of membrane fluidity and integrity, bring about the observed decline in sperm function (Jones et al, 1979; Aitken and Clarkson, 1987a, 1987b) ....
[...]
...…the damage inflicted by these radicals on functional cells in the same vicinity should theo- retically involve the peroxidation of unsaturated fatty acids in the sperm plasma membrane (Jones et al, 1979; Aitken and Clarkson, 1987a, 1987b ), the protective action of anti-oxidants was investigated....
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