Effects of negative pressure applied before storage (liquid storage, 17°C) on boar semen quality and fertilization ability

Author(s):  
LI. Jingchun ◽  
LI. Qi ◽  
LI. Yanbug ◽  
WEI Guosheng ◽  
SUN Dongbo

The present study was aimed to investigate the effects of negative pressure applied before storage on the quality and fertilization ability of boar semen. Boar semen samples were collected and pooled, and diluted with Modena solution containing 0.4% (w/v) of bovine serum albumin. Negative pressure was applied for 2–5 min using a vacuum pump with a barometer. The pressure applied were 0 (Control), -0.02 MPa (P200), -0.04 MPa (P400), and -0.08 MPa (P800). The sperm motility, acrosome integrity and sperm fertilizing ability were evaluated. Application of –0.04 MPa improved the sperm motility, acrosome integrity and fertilizing ability, compared with the other groups. The sperm motility and acrosome integrity decreased with increasing storage time in vitro. After 5 days, the sperm motility and acrosome integrity of the P400 group were all higher than those of the other groups (P less than 0.05). The cleavage rate (64.5% ± 2.4%) and blastocyst development rate (33.9% ± 2.8%) for semen stored for 7 days were similar to those of fresh semen. In conclusion, application of –0.04 MPa before liquid storage at 17°C can improve the quality and fertilization ability of boar semen.

2019 ◽  
Vol 71 (2) ◽  
pp. 489-499
Author(s):  
B.A. Pereira ◽  
L.G.P. Rocha ◽  
M.C. Teles ◽  
W.E. Silva ◽  
J.A. Barbosa ◽  
...  

ABSTRACT A study was conducted to evaluate the effect of chlorogenic acid (ChA) added pre-cooling and its combination with caffeine added during warming on cooled-stored boar semen parameters. Ten ejaculates were diluted in commercial extender with or without 4.5mg/ml ChA and stored at 15°C. After 0, 24 and 72 hours of storage, aliquots of these doses were taken and incubated at 37°C in the presence or absence of 8.0mM caffeine. Semen quality was evaluated after 10 and 120 minutes of incubation. The ChA increased (P <0.01) the sperm motility, viability, acrosomal integrity and the percentage of spermatozoa with high mitochondrial activity (PMHA), however, decreased (P <0.01) the malondialdehyde (MDA) concentration. Caffeine increased (P<0.05) the sperm motility, viability, PMHA and the MDA concentration and reduced (P <0.05) the acrosome integrity. When associated (ChA+caffeine), there was an increase (P <0.05) in sperm motility and viability, PMHA and acrosome integrity. The addition of ChA to the dilution medium improves the quality of the swine inseminating doses. The addition of caffeine during re-warming is only recommended when the semen is stored for prolonged periods (72h), and the inseminating dose should be used immediately after its addition.


Author(s):  
Jingchun Li ◽  
Qi Li ◽  
Guosheng Wei ◽  
Jiabao Zhang ◽  
Yanbing Li

This study aimed to investigate the influence of negative pressure on boar semen quality during liquid preservation at 17°C. Semen samples from ten large white boars were collected and pooled, divided into four equal parts, and diluted with Modena containing 0.4% (w/v) of bovine serum albumin. The semen samples were placed in a closed container with valve, and a negative pressure was applied for 2–5 minutes using a vacuum pump with a barometer. The control group had no treatment, the P200 group was treated at 200 mbar, the P400 group at 400 mbar, and the P800 group at 800 mbar. During liquid preservation, sperm motility, total antioxidant capacity, and semen H2O2 content were analyzed every 24 h. The effective survival time of boar semen during preservation was evaluated. The results indicated that a suitable negative pressure decreased the effects on reactive oxygen species on boar sperm quality during liquid preservation compared with the control group. Among all the groups, the 400 mbar negative pressure group had the highest sperm motility, total antioxidant capacity, and the percentage of spermatozoa with high mitochondrial membrane potential. The P400 group also had semen H2O2 content than the other groups. A suitable negative pressure improves sperm quality by reducing oxidative stress and the respiratory metabolism of sperm, and the optimum negative pressure is 400 mbar.


2014 ◽  
Vol 83 (1) ◽  
pp. 13-18 ◽  
Author(s):  
Janko Mrkun ◽  
Tamara Dolenšek ◽  
Tanja Knific ◽  
Anja Pišlar ◽  
Marjan Kosec ◽  
...  

One of the features of apoptosis is the externalization of phosphatidylserine which could be used to remove apoptotic cells from semen preparations. Magnetic-activated cell sorting using annexin V-conjugated microbeads which bind to phosphatidylserine could be used to enhance semen quality. Twelve boar semen samples after 3 days of liquid storage at 16­­–17 °C were subjected to magnetic-activated cell sorting. Bound and unbound fractions and control samples were subjected to flow cytometry following the staining of spermatozoa with Annexin V conjugated with Alexa Fluor 488 and propidium iodide. Four subpopulations were obtained: live, early apoptotic live, late apoptotic, early necrotic dead and late necrotic dead. The frequency of early apoptotic and late necrotic spermatozoa was significantly higher (P< 0.05) in bound (14.1 ± 10.6% and 24.1 ± 10.2%, respectively) than in unbound fractions (3.4 ± 2.1% and 12.7 ± 3.1%) and control (3.5 ± 1.6% and 12.0 ± 5.0%). The lowest concentration of live spermatozoa was found in the bound fraction (10.6 ± 8.0 %), which differed significantly (P< 0.05) from the control. In unbound fractions there was a significantly higher concentration (P< 0.05) of morphologically normal spermatozoa (31.8 ± 12.6%) compared to bound ones (5.9 ± 7.3%). A significantly (P< 0.05) lower proportion of morphologically normal spermatozoa was observed in both fractions compared to control (67.2 ± 17.0%). Boar spermatozoa were separated by the above method for the first time, however, the results showed this method to be inappropriate for boar semen separation under the tested conditions.


Biomolecules ◽  
2018 ◽  
Vol 8 (3) ◽  
pp. 65 ◽  
Author(s):  
Saleem Banihani

Since 1963, various research studies and reports have demonstrated the role of uric acid (2,6,8-trihydroxypurine), an end product of adenosine and guanosine catabolism, on semen quality and sperm function. However, this effect has not yet been collectively discussed, even though uric acid has been a well-recognized constituent in semen. Here, we systematically and comprehensively discuss and summarize the role/effect of uric acid in semen quality by searching the main databases for English language articles considering this topic. Additionally, certain significant and relevant papers were considered to support discussions and perceptions. In conclusion, uric acid contributes to maintaining and enhancing sperm motility, viability, and morphology; therefore, protecting sperm function and fertilizing ability. This contribution is performed mainly by neutralizing the damaging effect of oxidizing (e.g., endogenous free radicals and exogenous toxins) and nitrating agents and enhancing certain bioactive enzymes in spermatozoa. In contrast, high levels of uric acid may induce adverse effects to sperm function, at least in part, by reducing the activity of vital enzymes in spermatozoa. However, further research, mainly clinical, is still required to fully explore the role/effect of uric acid in semen.


2020 ◽  
Vol 89 (1) ◽  
pp. 47-53
Author(s):  
Aiste Iljenkaite ◽  
Sigita Kerziene ◽  
Agila Dauksiene ◽  
Zoja Mikniene ◽  
Henrikas Žilinskas ◽  
...  

The current study was carried out to investigate the protective effects of royal jelly supplementation on sperm motility, viability and pH value during the liquid storage of boar semen at 16 °C and 4 °C, at various periods of time (0, 24, 48, 72 and 96 h). Semen samples were collected from 11 boars, diluted with a long-term extender and supplemented with different concentration of royal jelly (0%, 0.5%, 1% and 2%) at a final concentration of 50 × 106 sperm/ml. In the laboratory, the semen was assessed for sperm morphology, viability (eosin-nigrosin staining), subjective motility and objective sperm motility by sperm class analyzer. In total, 396 tests for sperm viability and motility were performed. The longer storage time and the lower incubation temperature showed lower sperm motility and viability results. The results showed that royal jelly supplementation at 1% concentrations protected the functionality of the sperm plasma membrane during the liquid storage time of 96 h at 16 °C. Sperm subjective and objective motility results in samples stored at 4 °C decreased with higher royal jelly concentrations and longer storage time, and differ significantly from the results in samples stored at 16 °C (P < 0.05). Our data showed that royal jelly supplementation at lower concentrations can improve boar semen motility and viability parameters during liquid storage at 16 °C for 96 h.


2017 ◽  
Vol 62 (No. 5) ◽  
pp. 245-252 ◽  
Author(s):  
S. Gungor ◽  
C. Ozturk ◽  
AD Omur

The aim of this study was to determine the effects of trehalose and cysteine on sperm motility, viability, mitochondrial activity and acrosome integrity during liquid storage of Merino ram semen. Ejaculates were collected using artificial vaginas from five Merino rams, microscopically evaluated and pooled at 37 °C. The pooled semen samples were diluted in a Tris-based extender, including cysteine (2 mM and 4 mM), trehalose (10 mM and 25 mM) and no antioxidant (control). Diluted semen samples were kept in tubes and cooled from 37 to 5 °C in a cold cabinet, and maintained at 5 °C. Cooled samples were evaluated for sperm motility, viability, mitochondrial activity and acrosome integrity at 0, 24, 48, 72 and 96 h. Extender supplemented with trehalose (10 and 25 mM) and cysteine (2 and 4 mM) led to higher motility in comparison to the control at 24, 48, 72 and 96 h of liquid storage (P &lt; 0.05). Trehalose at the doses of 10 mM, 25 mM and 2 mM cysteine led to higher viability between 24–48–72 h and at 96 h of liquid storage (P &lt; 0.05). Further, 4 mM of cysteine improved sperm viability rates at 24 and 48 h of storage compared to the control group (P &lt; 0.05), and resulted in improved acrosome integrity rates compared to the control group at 72 and 96 h of storage (P &lt; 0.05). Extender supplemented with 10 and 25 mM trehalose at 24 and 72 h and 4 mM cysteine at 24 and 96 h of storage led to higher sperm mitochondrial activity rates when compared to the control group (P &lt; 0.05). In conclusion, the findings of this study show that trehalose and cysteine provided significant protection to ram sperm parameters during liquid storage.


2016 ◽  
Vol 28 (2) ◽  
pp. 221
Author(s):  
D. Le Bourhis ◽  
S. Camugli ◽  
P. Salvetti ◽  
L. Schibler ◽  
E. Schmitt

SensiTemp, a new in vitro maturation (IMV) bull straw concept, presents the advantage of colour changing while the straw is thawed. The colour of frozen straws is blue and straws start to become white when the temperature reaches 33°C, with a complete change of colour at 37°C. The objective of this study is to assess sperm quality after thawing of semen frozen in SensiTemp from 2 bulls, by analysing, in experiment 1, sperm motility and membrane integrity using computer-assisted semen analysis (CASA) and flow cytometry (FC), and, in experiment 2, the in vitro embryo production (IVP) using IVP technologies [IVM, IVF, and in vitro culture (IVC)]. The ejaculates of 2 bulls, selected during preliminary experiments on high in vitro fertility, were harvested at CIA L’Aigle, France, and split ejaculates were frozen in experimental (SensiTemp) and conventional (control) straws. In experiment 1 after thawing semen from the 2 types of straws (5 pooled straws each; 2 replicates), motility was assessed using the IVOS CASA system (Hamilton Thorne Inc., Beverly, MA, USA) and membrane integrity was evaluated through FC with Cytosoft software (Millipore-Guava Technologies Inc., Hayward, CA, USA). In experiment 2, IVF was used to evaluate the non-toxicity of SensiTemp and control straws. Cumulus-oocyte complexes (COC; n = 1178; 4 replicates) collected from slaughterhouse ovaries were matured in IVM medium (TCM-199 with bicarbonate, Sigma-Aldrich, Saint Quentin Fallavier, France; 10 µg mL–1 FSH-LH, Reprobiol, Liège, Belgium; and 10% FCS, Thermo Fisher, Illkirch, France) for 22 h. After fertilization, presumptive zygotes of each group (SensiTemp and control for each bull) were cultured in synthetic oviduct fluid medium (SOF, Minitube, Tiefenbach, Germany) with 1% estrous cow serum (ECS) and 0.6% BSA (Sigma-Aldrich, France) up to 8 days. All cultures were conducted at 38.5C in 5% CO2, and 5% O2. The cleavage and blastocysts rates were evaluated on Days 3 and 7, respectively, for each group. Embryo quality was recorded on Day 7 according to the IETS evaluation. Data from each bull were analysed separately using the chi-squared test (P < 0.05). In experiment 1, neither sperm motility from bull 1 (61.2 and 60.5%) and bull 2 (66.2 and 66.5%) nor membrane integrity from bull 1 (58.6 and 52.2%) and bull 2 (61.0 and 61.9%) were different between SensiTemp and control, respectively. Results from experiment 2 showed no difference (P > 0.05) in cleavage rate between SensiTemp and control for the 2 bulls: 92.1 and 91.7% for bull 1 and 94.2 and 94.6% for bull 2 respectively. The blastocysts rate on Day 7 did not differ (P > 0.05) among groups (47.5, 47.1 and 51.3, 50.4% for SensiTemp and control bull 1 and bull 2, respectively) nor the quality of embryos retrieved in the different groups: 25.4, 23.3, and 30.8, 29.6% in grade 1 embryo for SensiTemp and control bull 1 and bull 2, respectively. Those results demonstrate, in vitro, that the new SensiTemp straws were non-toxic and did not affect the semen quality after thawing nor did the SensiTemp straws affect the ability of sperm cells to fertilize oocytes and produce 8-day-old embryos.


2020 ◽  
Vol 47 (5) ◽  
pp. 48-57
Author(s):  
O. A. Sokunbi ◽  
O. Alaba ◽  
B. Ogunwumiju ◽  
S. Eboh ◽  
T. Iruo

Fertilizing potential of spermatozoa is pivotal for successful artificial insemination. Preservation of boar semen is associated with the production of reactive oxygen species which leads to decline of spermatozoa quality and decrease in fertilizing ability. The study aimed at investigating the effects of Ethanolic Allium sativum Extracts (EASE) as a cytoprotective agent on the quality and fertilizing potential of spermatozoa in extended boar semen. Semen was collected and aliquot portions were divided into 60 sample bottles comprising of five treatments with replicates. T1 served as the control (Beltsville Thawing Solution (BTS) + semen) in ratio 1:3 semen-extender, while other treatments T2 to T5 contained semen-extender at the same ratio, but supplemented with EASE at varying concentrations of 50, 75, 100 and 125 μg/L respectively stored at 17 °C and were evaluated at 0, 24, 48 and 72 hours for pH, normal spermatozoa (NS) (%), liveability (%), acrosome integrity (AI) (%) in a completely randomized design. Data obtained were subjected to descriptive statistics and analysed using one way ANOVA and means were separated using Duncan Multiple Range Test. Treated semen samples (75, 100 and 125 μg/L) gave superior (P<0.05) liveability than control, up to 72 hours. Similar (P>0.05) mean values were observed for NS from all treatments except 50 μg/L at 24 hours. At 72 hours, treated samples (75, 100 and 125 μg/L) gave superior (P<0.05) NS. Significant differences (P<0.05) in mean values for pH were observed among treatments through the extension period. However, all values were within accepted range for quality semen. Acrosome integrity were similar (P>0.05) among treatments from 0 to 72 hours, but reduced (P<0.05). Results suggest that the supplementation of BTS with EASE 125 μg/L, gave better cytoprotection to the spermatozoa compared to the control.     L'ablitédes spermatozoïdes de fertiliserest essentiel pour une insémination artificielle réussie. La conservation de la semence de l'our est associée à la production d'espèces réactives de l'oxygène, ce qui entraîne un déclin de la qualité des spermatozoïdes et une diminution de la capacité de fertilisation. L'étude visait à étudier les effets des extraits éthanoliques d'Allium sativum (EEAS) en tant qu'agent cytoprotecteur sur la qualité et le potentiel fertilisant des spermatozoïdes dans le sperme de sanglier étendu. Le sperme a été collecté et les portions aliquotes ont été divisées en 60 bouteilles d'échantillons comprenant cinq traitements avec des répliques. T1 a servi de contrôle (BeltsvilleThawing Solution (BTS) + sperme) dans un rapport 1: 3 sperme-extenseur, tandis que d'autres traitements T2 à T5 contenaient sperme-extenseur au même rapport, mais complété avec EEAS à des concentrations variables de 50, 75 , 100 et 125 μg / L respectivement conservés à 17 ° C et ont été évalués à 0, 24, 48 et 72 heures pour le pH, les spermatozoïdes normaux (SN) (%), l'habitabilité (%), l'intégrité de l'acrosome (IA) (%) dans une conception complètement aléatoire. Les données obtenues ont été soumises à des statistiques descriptives et analyses en utilisant une méthode ANOVA et les moyennes ont été séparées en utilisant le test Duncan Multiple Range. Les échantillons de sperme traités (75, 100 et 125 μg / L) ont donné une habitabilité supérieure (P <0.05) à celle des témoins, jusqu'à 72 heures. Des valeurs moyennes similaires (P> 0.05) ont été observées pour le SN pour tous les traitements, sauf 50 μg / L à 24 heures. A 72 heures, les échantillons traités (75, 100 et 125 ug / L) ont donné une SN supérieure (P <0.05). Des différences significatives (P <0.05) dans les valeurs moyennes du pH ont été observées entre les traitements pendant la période d'extension. Cependant, toutes les valeurs se situaient dans la plage acceptée pour le sperme de qualité. L'intégrité des acrosomes était similaire (P> 0.05) parmi les traitements de 0 à 72 heures, mais réduite (P <0.05). Les résultats suggèrent que la supplémentation en BTS avec EEAS 125 μg / L, a donné une meilleure cytoprotection des spermatozoïdes par rapport au témoin.


2011 ◽  
Vol 23 (1) ◽  
pp. 115 ◽  
Author(s):  
Z. Namula ◽  
R. Kodama ◽  
Y. Kaedei ◽  
F. Tanihara ◽  
V. L. Vien ◽  
...  

Liquid preservation of semen can be an alternative to frozen–thawed semen for artificial insemination. The success of a selection of boar semen extenders has been studied over storage periods of 5 to 7 days. The objective of this study was to evaluate the effects of skim milk on the viability and in vitro fertility of boar spermatozoa preserved in Modena-based extenders at 5°C and 15°C for 2 weeks. A total of 7 ejaculates were collected from one boar. The sperm-rich fraction of each ejaculate was centrifuged and diluted in Modena extenders supplemented with 0 (control), 7.5, and 15 mg mL–1 of dry skim milk. The final sperm concentration was adjusted to 1 × 108 cells mL–1, and then the semen was stored at 5°C and 15°C for 2 weeks. In the first experiment, the motility, viability (live/dead fluorescence viability assay), plasma membrane integrity (hypoosmotic swelling test; HOST), and acrosome integrity (FITC-labelled peanut agglutinin staining) of semen stored for 2 weeks were assessed. In the second experiment, the fertilization of stored semen after 20 h of co-incubation with in vitro matured oocytes and their development were examined. Data were analysed using ANOVA. When the semen was stored at 5°C for 2 weeks, the mean total sperm motility of semen stored with 7.5 and 15 mg mL–1 of dry skim milk was significantly higher than that of semen in the control group (41.4% and 41.5% v. 17.4%; P < 0.05). However, the beneficial effects of skim milk on the sperm motility were not observed in the semen stored at 15°C. Moreover, there were no significant differences in the other parameters of semen quality among the groups in each storage temperature. Significantly higher penetration rates of semen stored with 7.5 and 15 mg mL–1 of dry skim milk were observed in the storage at 5°C (41.1% and 34.8% v. 19.8%; P < 0.05) but not at 15°C (38.9% and 26.0% v. 30.0%; P > 0.05) when compared with the control group. When the semen was stored at 5°C, the development rate to the blastocyst stage of oocytes fertilized with semen stored with 7.5 mg mL–1 of dry skim milk was significantly higher than that with control and 15 mg mL–1 of dry skim milk (15.4% v. 1.1% and 7.8%; P < 0.01). However, there were no significant differences in the development rates of oocytes fertilized with semen stored at 15°C among the groups (9.6–11.9%). In conclusion, our results indicate that the effect of skim milk on the viability and in vitro fertility of liquid-stored boar spermatozoa is dependent on the storage temperature. The addition of 7.5 mg mL–1 of dry skim milk may be effective for the improvement of viability and fertility of semen stored at 5°C but not at 15°C.


2019 ◽  
Vol 64 (No. 5) ◽  
pp. 209-216 ◽  
Author(s):  
R Ratchamak ◽  
T Vongpralub ◽  
W Boonkum ◽  
V Chankitisakul

The purpose of this study was to examine sperm quality after cryopreservation of ejaculates collected as a bulk sample, which is routinely part of semen collection, and to compare this quality with the sperm-rich fraction in boars. Ejaculates were collected as sperm-rich fractions (SRF) and bulk samples (BE) using a gloved-hand technique. Fresh semen quality in terms of semen volume, sperm concentration, total sperm motility and pH were conventionally evaluated. Then, semen was cryopreserved using the liquid nitrogen vapour method. The post-thaw sperm quality was evaluated by assessing sperm motility, live sperm with normal apical ridge and high mitochondrial energy status, lipid peroxidation was evaluated using CASA and fluorescent multiple staining and MDA levels were determined using a spectrophotometer, respectively. In terms of fresh semen quality, sperm motility in fresh semen did not differ significantly between the two groups. The treatment with the greater mean volume (BE; P &lt; 0.05) had a lower mean sperm concentration (P &lt; 0.05); meanwhile, the mean ejaculate pH collected as BE was more basic compared with SRF (P &lt; 0.05). However, there were no significant post-thaw quality changes between sperm-rich fractions and bulk samples of semen. In conclusion, ejaculates can be collected as bulk samples without the need to classify fractions for boar semen cryopreservation.


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