Role of amino acids as additives on sperm motility, plasma membrane integrity and lipid peroxidation levels at pre-freeze and post-thawed ram semen

2015 ◽  
Vol 161 ◽  
pp. 82-88 ◽  
Author(s):  
Sharon Sangeeta ◽  
A. Arangasamy ◽  
S. Kulkarni ◽  
S. Selvaraju
Keyword(s):  
Amino Acids ◽  
Lipid Peroxidation ◽  
Plasma Membrane ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Plasma Membrane Integrity

20 LIPID PEROXIDATION AND GENERATION OF HYDROGEN PEROXIDE FROM SUBFERTILE STALLION SPERMATOZOA DURING STORAGE AT REFRIGERATION TEMPERATURE

2013 ◽  
Vol 25 (1) ◽  
pp. 157 ◽  
Author(s):  
P. N. Guasti ◽  
C. P. Freitas-Dell'aqua ◽  
R. R. D. Maziero ◽  
G. A. Monteiro ◽  
F. P. Hartwig ◽  
...  
Keyword(s):  
Lipid Peroxidation ◽  
Plasma Membrane ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Membrane Lipid ◽  
Sperm Cells ◽  
Band Pass Filter ◽  
Membrane Lipid Peroxidation ◽  
Pass Filter ◽  
Plasma Membrane Integrity

The aim of this study was to evaluate the generation of hydrogen peroxide (H2O2) and membrane lipid peroxidation of subfertile spermatozoa stored at 5°C for 24 h. Semen samples, collected from 5 subfertile stallions (≤40% of conception rate), were diluted in a skim-based extender (BotuSemen; Botupharma) and stored in a passive transport container at 5°C over a period of 24 h. Sperm motility was determined by computer-assisted semen analysis (CASA; IVOS 12, Hamilton Thorne Inc., Beverly, MA, USA) for total motility (TM), progressive motility (PM), and rapid sperm (RAP). Plasma membrane integrity, generation of H2O2, and membrane lipid peroxidation were determined by flow cytometry (LSRFortessa cell analyzer, BD Biosciences, Franklin Lakes, NJ, USA). For evaluation of acrosome and plasma membrane integrity, samples were stained with Hoechst 33342 dye (H33342; Molecular Probes, Eugene, OR, USA), iodide propidium (IP; Sigma, St. Louis, MO, USA), and fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (FITC-PSA; Sigma). For the assessment of generation of H2O2, 2′,7′-dichlorofluorescein diacetate (DCFH-DA; Sigma), H33342, and IP were added to the sperm suspension. For the assessment of sperm lipid peroxidation, samples were stained with C11-BODIPY581/591 (Molecular Probes), H33342, and IP. Samples were evaluated after semen collection (0 h) and after 24 h of cooled storage. A total of 10 000 gated events were analyzed per sample by flow cytometry. The green fluorescence (FL1) was collected through a 580-nm band-pass filter and the red fluorescence (FL3) through a 635-nm band-pass filter. Statistical analysis was performed using GraphPad Prism version 4.03 (2005; GraphPad Software Inc., La Jolla, CA, USA), through paired t-test to identify the significant differences (P ≤ 0.05). In general, sperm motility parameters of TM and RAP, and acrosome and plasma membrane integrity significantly decreased after 24 h of refrigeration at 5°C (P ≤ 0.05). Interestingly, no differences were found in PM at 0 and 24 h (P ≥ 0.05). The percentage of sperm cells with high generation of H2O2 did not differ at 0 and 24 h (P ≥ 0.05), whereas the percentage of sperm cells with membrane peroxidation increased (0.56 ± 0.3 v. 2.24 ± 1.3; P ≤ 0.05) at these periods. The percentage of viable sperm cells with low generation of H2O2 had a significant decrease from 22.6 ± 11.2 at 0 h to 0.08 ± 0.1 after 24 h of storage (P ≤ 0.05), although no differences were found in the percentage of viable sperm cells without membrane peroxidation (P ≥ 0.05). In conclusion, the cooled storage of subfertile spermatozoa for 24 h drastically decreased the number of viable spermatozoa with low generation of H2O2 and increased the percentage of membrane lipid peroxidation, which is related to the decrease in sperm motility and increase in dead sperm. These results make it difficult to use refrigerated semen of subfertile stallions with poor semen quality in commercial breeding programs. São Paulo Research Foundation (FAPESP) is acknowledged for supporting this research.

scholarly journals Effects of Arsenite and Antioxidants on Sperm Motility, Plasma Membrane Integrity, Mitochondrial Activity, and Lipid Peroxidation in Pigs

2017 ◽  
Vol 27 (5) ◽  
pp. 517-523 ◽  
Author(s):  
Han-Su Kim ◽  
Yu-Sub Lee ◽  
Sang-Hee Lee ◽  
Hee-Tae Cheong ◽  
Choon-Keun Park ◽  
...  
Keyword(s):  
Lipid Peroxidation ◽  
Plasma Membrane ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Mitochondrial Activity ◽  
Plasma Membrane Integrity

87 OSMOTIC SENSITIVITY OF OKAPI SPERMATOZOA AND DEVELOPMENT OF CRYOPRESERVATION PROTOCOLS USING CRYOMICROSCOPY

2008 ◽  
Vol 20 (1) ◽  
pp. 124 ◽  
Author(s):  
L. M. Penfold
Keyword(s):  
Plasma Membrane ◽  
Sperm Motility ◽  
Repeated Measures ◽  
Membrane Integrity ◽  
Latin Square ◽  
Crystal Formation ◽  
Plasma Membrane Integrity ◽  
Power Of The Test ◽  
Before And After ◽  
Cooling Effects

Okapi would benefit from artificial insemination with frozen-thawed sperm in cases where aggression prevents mating or where individuals are geographically disparate. Effective sperm cryopreservation is a prerequisite to this goal. Ejaculates (n = 20) were collected from 7 anesthetized adult male okapi housed individually, or with a female for breeding, throughout the year by electroejaculation, and semen and sperm parameters were assessed. Semen aliquots were centrifuged; resuspended in 500 µL of PBS with the osmolarity adjusted to 35, 75, 150, 600, 1200, and 2400 mOsm; and incubated for 30 min before returning to isosmotic conditions. Semen was extended in TEST containing 1%, 2%, or 4% glycerol with or without 0.5% Equex (Minitube, Verona, WI, USA); 5-µL aliquots were cooled in a Latin square design on a Linkam BCS 196 cryomicrostage (Linkam Scientific, Tadworth, Surrey, UK) at 20�C min–1 to –6� –12�C at which point ice crystal formation was induced (seeded), and cooled further to –70�C before warming at 50�C min–1 to 35�C (okapi body temperature). To investigate cooling effects only, raw ejaculate was cooled to –6�C without seeding and warmed to 35�C. Percent sperm motility and plasma membrane integrity (PMI) were recorded before and after treatments. Differences were examined using one-way repeated measures analysis of variance. No differences in motility, total sperm numbers, or percent normal morphology were observed throughout the year (P > 0.05), although the power of the test was low so that negative findings should be interpreted cautiously. Mean semen volume was 1.3 � 0.19 mL, sperm motility was 29 � 3.2%, with a PMI of 39 � 6.8%; 48 � 2.8% were morphologically normal. High proportions of non-motile, plasma membrane-damaged cells were noted in every ejaculate, and whiplash motility, possibly indicating spontaneous capacitation, was observed in several ejaculates 1 h after collection. Motility was dramatically reduced on either side of isosmotic conditions and was more sensitive to osmotic pressure than was plasma membrane integrity. Cooling of raw ejaculate to sub-zero temperatures without freezing did not result in any loss of motility or PMI, indicating cold tolerence. Superior results were obtained when sperm were frozen-thawed in TEST containing 4% glycerol with 0.5% Equex. Findings indicate that okapi semen collected by electroejaculation routinely contain high numbers of non-motile and plasma membrane-damaged spermatozoa, apparently unrelated to season or the length of time since the male was housed with a breeding female. Okapi spermatozoa are remarkably intolerant of departures from isosmotic conditions, indicating a lack of ability to regulate or withstand volume excursions during osmotic stress events; however, cooling to sub-zero temperatures in the absence of cryoprotectant did not reduce percent sperm motility or PMI, indicating resistance to cold shock. Increasing and maintaining proportions of motile, membrane-intact spermatozoa prior to and during cryopreservation will be critical for development of freezing protocols for this species.

The effects of chronic administration of ghrelin on rat sperm quality and membrane integrity

2009 ◽  
Vol 59 (2) ◽  
pp. 159-168 ◽  
Author(s):  
Arash Kheradmand ◽  
Majid Taati ◽  
Homayoon Babaei
Keyword(s):  
Plasma Membrane ◽  
Treatment Group ◽  
Sperm Motility ◽  
Sperm Quality ◽  
Sperm Concentration ◽  
Membrane Integrity ◽  
Chronic Administration ◽  
Control Group ◽  
Plasma Membrane Integrity ◽  
Significant Difference

AbstractAlthough ghrelin acts as a modulator of feeding behavior and energy metabolism in the central nervous system, recent studies have implicated the peripheral actions of ghrelin in reproductive tissues. Here, we investigated the effects of chronic administration of ghrelin on the motility, plasma membrane integrity and concentration of rat spermatozoa. 45-d male Wistar rats were scheduled for the study and were divided into control and treatment groups. In the treatment group, 1 nmol of ghrelin was administered as sc injection for 10 consecutive days or vehicle (physiological saline) to the control rats. Sperm collection was achieved by killing of the rats on days 15, 25 and 50 after first injection. Total sperm motility and forward progressive movement did not exhibit significant difference during the experiment, although, there was a tendency for greater motion rate on d 15 and 25 in the treated rats compared to the control group. Plasma membrane integrity (HOS-reacted spermatozoa) was significantly higher in the treated animals, especially on day 15 as well as day 25, because of possible antioxidant properties of ghrelin. This value was statistically higher on day 15 than that of day 25 (P <0.05). Likewise, there was a significant correlation between the FPM (P <0.0001, r = 0.79) and TSM (P <0.01, r = 0.52) with the HOS test percentage in the treatment group. It was not observed statistically difference in the sperm concentration between groups during all of the experimental days. In conclusion, chronic administration of ghrelin (similar to induced by energy deficiency such as fasting) increased the integrity of sperm membrane, however, the sperm motility and concentration did not display any alterations.

Effect of platelet activating factor (PAF) supplementation in semen extender on viability and ATP content of cryopreserved canine spermatozoa

2010 ◽  
Vol 13 (4) ◽  
pp. 571-579 ◽  
Author(s):  
W. Kordan ◽  
M. Lecewicz ◽  
R. Strzeżek ◽  
A. Dziekońska ◽  
L. Fraser
Keyword(s):  
Plasma Membrane ◽  
Sperm Motility ◽  
Mitochondrial Function ◽  
Sperm Quality ◽  
Membrane Integrity ◽  
Platelet Activating Factor ◽  
Computer Assisted ◽  
Atp Content ◽  
Plasma Membrane Integrity ◽  
Semen Extender

Effect of platelet activating factor (PAF) supplementation in semen extender on viability and ATP content of cryopreserved canine spermatozoa The aim of this study was to investigate the effect of platelet activating factor (PAF) on the quality characteristics of cryopreserved canine spermatozoa. Cryopreserved semen of 5 mixed-breed dogs was treated with different concentrations of exogenous PAF (1 × 10-3M, 1 × 10-4M, 1 × 10-5M and 1 × 10-6M) and examined at different time intervals (0, 30, 60 and 120 min). Cryopreserved semen treated without PAF was used as the control. Sperm quality was evaluated for motility (computer-assisted semen analysis, CASA), mitochondrial function (JC-1/PI assay) and plasma membrane integrity (SYBR-14/PI assay and Hoechst 33258). Also, ATP content of spermatozoa was determined using a bioluminescence assay. Treatment of cryopreserved semen with 1 × 10-3 M PAF at 120 min of incubation resulted in significantly higher total sperm motility compared with the control. It was observed that PAF-improved total sperm motility was concurrent with enhanced sperm motility patterns after treatment of cryopreserved semen. Treatment of cryopreserved semen with PAF did not improve either sperm mitochondrial function or plasma membrane integrity, as monitored by different fluorescent membrane markers. Furthermore, ATP content of cryopreserved spermatozoa was significantly higher when PAF was used at a concentration of 1 × 10-3 M compared with the control and other PAF treatments, regardless of the incubation time. The findings of this study indicated that treatment with 1 × 10-3 M PAF at 120 min of incubation rendered better quality of cryopreserved canine semen, which was associated with improved sperm motility parameters and ATP content. It can be suggested that exogenous PAF addition is beneficial as a supplement for canine semen extender used for.

scholarly journals Effects of the addition of docosahexaenoic acid and ?-tocopherol on quality of equine spermatozoa stored at 5°C

2020 ◽  
Vol 41 (1) ◽  
pp. 167 ◽  
Author(s):  
Breno Fernandes Barreto Sampaio ◽  
Bruno Gomes Nogueira ◽  
Maria Inês Lenz Souza ◽  
Eliane Vianna da Costa-e-Silva ◽  
Carmem Estefânia Serra Neto Zúccari
Keyword(s):  
Lipid Peroxidation ◽  
Plasma Membrane ◽  
Docosahexaenoic Acid ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Control Group ◽  
Mitochondrial Activity ◽  
Membrane Composition ◽  
Artificial Vagina

Plasma membrane composition has impact on phase transition from liquid crystal to gel state of cooled sperm cell. The incorporation of polyunsaturated fatty acids increases its fluidity and can contribute to sperm motility. The aim of this study was to compare the effect of adding docosahexaenoic acid (DHA) and ?-tocopherol (?-Toh) to the cooling extender, singly or combined, to the equine sperm parameters, submitted to cooling, up to 72 hours. Two ejaculates of ten stallions collected with artificial vagina were used, and evaluated for motility, plasma membrane integrity, chromatin fragmentation, mitochondrial activity and lipid peroxidation, according to the following treatments: C; DHA; ?-Toh; DHA/?-Toh; EtOH 100: and EtOH 140 (corresponding to control; 10 ng mL-1 of DHA; 2 mM of ?-Toh; : 10 ng mL-1 of DHA + 2 mM of ?-Toh; 100 µL of ethanol and 140 µL of ethanol respectively). DHA treatment showed higher motility (68.2 ± 12.3; p < 0.05) when compared to control (62.1 ± 16.2), DHA/?-Toh (61.3 ± 12.7) and EtOH (58.1 ± 8.6) groups. In lipid peroxidation assay, the control group showed 2,506.2 ± 796.4 ng of MDA 108 spermatozoa-1, being significantly higher (p < 0.05) than the groups treated with DHA (2,036.0 ± 687.0), ?-Toh (1,890.8 ± 749.5) and DHA/?-Toh (1,821.1 ± 627.2). In conclusion, ?-Toh was effective in diminishing lipid peroxidation of equine sperm subjected to cooling, and DHA improved sperm motility and, in spite of being a polyunsaturated fatty acid with high susceptibility to peroxidation, reduced lipid peroxidation.

scholarly journals The Effect of Different Concentrations of Caffeine, Pentoxifylline and 2’-Deoxyadenosine on the Biological Properties of Frozen-Thawed Canine Semen

2019 ◽  
Vol 19 (3) ◽  
pp. 733-746
Author(s):  
Marek Lecewicz ◽  
Rafał Strzeżek ◽  
Anna M. Majewska ◽  
Piotr S. Purpurowicz ◽  
Władysław Kordan
Keyword(s):  
Plasma Membrane ◽  
Sperm Motility ◽  
Artificial Insemination ◽  
Membrane Integrity ◽  
Phosphodiesterase Inhibitors ◽  
Biological Properties ◽  
Kinematic Parameters ◽  
Plasma Membrane Integrity ◽  
Beneficial Effects

AbstractArtificial insemination (AI) and semen cryopreservation are the most accessible and commonly used techniques for breeding domestic animals. Among many parameters, such as plasma membrane integrity and acrosome structure, one of the key factors that determine the quality of frozen-thawed samples for artificial insemination is sperm motility. Sperm motility is one of the key parameters that determine the quality of frozen-thawed samples for AI. The total number of progressively motile spermatozoa in thawed canine semen is correlated with fertility. A variety of substances were used to compare sperm motility with the control. The aim of this study was to determine the effect of semen extender supplementation with motility stimulants, pentoxifylline (PTX), caffeine (CAF) and 2’-deoxyadenosine (DX), after different post-thaw incubation times (30, 60, 120 min) on the motility, selected kinematic parameters, plasma membrane integrity and mitochondrial membrane potential of cryopreserved canine spermatozoa. During attempts to improve the quality of cryopreserved semen, the applied substances exerted beneficial effects at a concentration of 10 mM. We demonstrated that both phosphodiesterase inhibitors, caffeine and pentoxifylline, as well as 2’-deoxyadenosine increased the motility and selected kinematic parameters of thawed canine spermatozoa.

scholarly journals Correlations Amongst Functional and Morphological Attributes and Oxidative Markers of Fresh and Cryopreserved Semen of Gir and Murrah Bulls

2020 ◽  
Vol 15 (03) ◽  
pp. 24-29
Author(s):  
A. J. Dhami ◽  
Tapasvi M Patel ◽  
DV Chaudhari
Keyword(s):  
Plasma Membrane ◽  
Sperm Motility ◽  
Semen Quality ◽  
Sperm Quality ◽  
Membrane Integrity ◽  
Winter Season ◽  
Correlation Coefficients ◽  
Plasma Membrane Integrity ◽  
Murrah Buffalo ◽  
Oxidative Markers

This study was undertaken during the winter season on healthy mature Gir cattle and Murrah buffalo bulls (n=3 each). The semen samples (6 ejaculates/bull, total 36 ejaculates) collected in the morning using artificial vagina were evaluated for routine seminal attributes, including acrosomal and plasma membrane integrity. The samples were then diluted @ 100 million sperm/ml with tris fructose yolk glycerol extender without and with sericin @ 0.1, 0.25, 0.5 and 1.0% (w/v), filled in French mini-straws, and frozen in LN2 using biofreezer as per standard freezing protocol. Straws were thawed in water bath at 37°C for 30 sec and evaluated for post-thaw quality, viz., motility, viability, morphology, acrosome integrity and plasma membrane integrity (HOST). Lipid peroxidation (malondialdehyde - MDA production) and activities of enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx) were assessed as oxidative markers in seminal plasma of freshly diluted and frozen-thawed semen samples. Sericn at 0.5% level significantly (p less than 0.01) improved the post-thaw sperm quality with reduced oxidative stress in both the species. The breed-wise correlation coefficients (r) among sperm quality attributes and oxidative markers were studied in fresh and frozen-thawed semen of each species, and also for fresh with frozen-thawed semen. The findings revealed significant interrelationships amongst most of the attributes of fresh as well as post-thawed semen and also of fresh semen attributes with those of cryopreserved semen including oxidative markers in both the species. Sperm motility estimation in fresh, pre-freeze and post-thawed semen was a legitimately good indicator of quality of spermatozoa at various steps of semen processing/freezing, and its fertilizing potential. Thus, the sperm motility, HOS test and either MDA or SOD/GPx activity alone may be used as valuable and practical tools for routine assessment of bovine semen quality considering significant correlations found between them.

scholarly journals Efficiency of Commercial Egg Yolk-Free and Egg Yolk-Supplemented Tris-Based Extenders for Dromedary Camel Semen Cryopreservation

Animals ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 999 ◽  
Author(s):  
Ayman Abdel-Aziz Swelum ◽  
Islam M. Saadeldin ◽  
Hani Ba-Awadh ◽  
Mohsen G. Al-Mutary ◽  
Abdullah F. Moumen ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Sperm Motility ◽  
Sperm Quality ◽  
Membrane Integrity ◽  
Egg Yolk ◽  
Dna Integrity ◽  
Computer Assisted ◽  
Dromedary Camel ◽  
Plasma Membrane Integrity ◽  
Semen Cryopreservation

This study compared the efficiency of commercial egg yolk-free (AndroMed, OPTIXcell) and egg yolk-supplemented (Triladyl, Steridyl) Tris-based extenders for semen cryopreservation in seven adult dromedary camels. The camel-specific extender SHOTOR was used as control. The collected semen samples were evaluated and diluted with SHOTOR, Triladyl, Steridyl, AndroMed, or OPTIXcell. The diluted semen was gradually cooled and equilibrated for two hours before liquid nitrogen freezing. Semen was evaluated prior to freezing and after freeze-thawing cycles for motility, kinetics, vitality, abnormality, plasma membrane integrity, and DNA fragmentation using computer-assisted sperm analysis. In pre-freezing evaluation, progressive sperm motility was higher in SHOTOR-diluted samples (21.54 ± 1.83) than in samples diluted with Steridyl, OPTIXcell, or AndroMed (15.76 ± 1.80, 17.43 ± 1.10, and 13.27 ± 1.07, respectively). Moreover, Triladyl and SHOTOR resulted in significantly (p < 0.05) better sperm vitality and DNA integrity than all other diluents, but Triladyl resulted in a significantly (p < 0.05) better plasma membrane integrity (87.77 ± 0.31) than SHOTOR (85.48 ± 0.58). In the post-thawing evaluation, Triladyl led to significantly (p < 0.05) higher sperm motility (38.63 ± 0.81%; p < 0.05) when compared to SHOTOR, Steridyl or AndroMed (35.09 ± 1.341%, 34.4 ± 0.84%, and 31.99 ± 1.48%, respectively), with OPTIXcell being the least efficient (28.39 ± 0.86%). Progressive sperm motility was the highest when using Triladyl. Post-thawing curvilinear, straight line and average path sperm velocities were highest with Triladyl and lowest with AndroMed. Triladyl led to the highest linearity coefficient and straightness sperm coefficient, while SHOTOR to the highest DNA and plasma membrane integrity. OPTIXcell and AndroMed resulted in poor post-thawing sperm vitality, while Steridyl was less efficient than Triladyl. The highest rate of sperm abnormalities was recorded with OPTIXcell and the lowest with SHOTOR or Triladyl. In conclusion, SHOTOR, Triladyl, Steridyl, AndroMed, and OPTIXcell can all be used for camel semen cryopreservation; however, SHOTOR and Triladyl provided the best post-thawing sperm quality. Based on our findings, Triladyl is the best commercially available extender for dromedary camel semen cryopreservation to date.

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