• Asian-Australasian Journal of Animal Sciences
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• 제31권11호
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• pp.1729-1737
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• 2018

Objective: The aim of the current study is to investigate the relationship between prohibitin (PHB), capping actin protein of muscle Z-line beta subunit (CAPZB), and tektin-2 (TEKT2) and sperm motility in Murrah buffalo. Methods: We collected the high-motility and low-motility semen samples, testis, ovary, muscle, kidney, liver, brain and pituitary from Murrah buffalo, and analysed the expression of PHB, CAPZB, and TEKT2 in mRNA (message RNA) and protein level. Results: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) result showed that the expression of PHB was higher and CAPZB, TEKT2 were specifically expressed in testis as compared to the other 6 tissues, and that in testis, the expression of TEKT2 was higher than that of CAPZB and PHB. Immunohistochemistry test revealed that all three genes were located on the convoluted seminiferous tubule and enriched in spermatogenic cells. Both qRT-PCR and Western Blot results showed that the expression levels of PHB, CAPZB, and TEKT2 were significantly lower in the low-motility semen group compared to the high-motility semen group (p<0.05). Conclusion: The expression levels of PHB, CAPZB, and TEKT2 in Murrah buffalo sperm have a high positive correlation with sperm motility. And the three genes may be potential molecular markers for the decline of buffalo sperm motility.

• 한국동물생명공학회지
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• 제36권2호
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• pp.91-98
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• 2021

The objective of this study was to establish a selection process for high quality sperm in bovine semen using sperm separation by magnetic activation (MACS). For this, semen from 21 Nellore bulls was collected using an artificial vagina. To guarantee the presence of pathologies in the ejaculate, animals previously declassified in four consecutive spermiogram were used. Semen was analyzed in five statuses: (1) fresh semen (fresh); (2) density gradient centrifugation (DGC), percoll column; (3) non-apoptotic fraction after separation by MACS (MAC); (4) apoptotic fraction from the separation (MACPOOR); and (5) MAC followed by DGC (MACDGC). Using a computerized analysis system (CASA), motility was measured. The sperm morphology was evaluated by phase contrast, and the supravital test was completed with eosin/nigrosin staining. For DGC, 20 × 10⁶ cells were used in a gradient of 90% and 45% percoll. MACS used 10 × 10⁶ cells with 20 μL of nanoparticles attached to annexin V, and filtered through the MiniMACS magnetic separation column. Membrane integrity was assessed with SYBR-14/IP and mitochondrial potential with JC-1 by flow cytometry. Processing sperm by MACDGC, was more effective in obtaining samples with high quality sperm, verified by the total of abnormalities in the samples: 35.04 ± 2.29%, 21.50 ± 1.47%, 17.30 ± 1.10%, 30.68 ± 1.94% and 10.50 ± 1.46%, respectively for fresh, DGC, MAC, MACPOOR, and MACDGC. The subpopulation of non-apoptotic sperm had a high number of live cells (82.65%), membrane integrity (56.60%) and mitochondrial potential (83.98%) (p < 0.05). These findings suggest that this nanotechnological method, that uses nanoparticles, is efficient in the production of high-quality semen samples for assisted reproduction procedures in cattle.

• 농업과학연구
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• 제49권1호
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• pp.103-112
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• 2022

Fipronil is a popular insecticide used in both agricultural and domestic fields. Factors that affect sperm and eggs have a direct influence on reproductive outcomes. This study was undertaken to assess the effect of varying concentrations (10 - 200 μM) of fipronil and incubation times (30 min and 2 hrs) on boar spermatozoa. Spermatozoa were evaluated for motility, motion kinematics, viability, chromatin stability, and for the generation of intracellular reactive oxygen species (ROS) and the results were compared to those from corresponding controls. The findings revealed a significant, dose-dependent reduction in sperm motility in all fipronil treatment groups at 30 min of incubation (p < 0.05). A similar dose-dependent reduction in sperm motility was observed subsequent to fipronil exposure for 2 hrs of incubation (p < 0.05). Groups treated with fipronil showed a gradual reduction in motion kinematics (p < 0.05). Moreover, a significantly higher percentage of dead sperm was observed at 200 μM fipronil, as compared to the highest live percentage obtained in controls (p < 0.05). Evaluating the sperm chromatin integrity revealed a significantly higher percentage of damaged chromatin in spermatozoa incubated with 200 μM of fipronil. Moreover, ROS production was significantly higher in fipronil-exposed sperm (p < 0.05). In conclusion, boar spermatozoa incubated with fipronil showed decreased levels of sperm motility and viability, weaker chromatin integrity, and increased levels of intracellular ROS generation, all of which indicate that exposure to fipronil potentially impairs the fertilization competence of boar spermatozoa.

• Animal Bioscience
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• 제37권2호
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• pp.203-209
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• 2024

Objective: This study aimed to assess the impact of the dilution ratio of Tris diluent, storage at 0℃, and long-distance transportation on the spermatozoa of Simmental cattle. It also validated the feasibility of the regional distribution of fresh semen. Methods: In experiment 1, semen was diluted at four dilution ratios (1:6, 1:9, 1:12, and 1:15) to determine the optimal dilution ratio of Tris diluent. In experiment 2, we assessed sperm viability, progressive motility (objectively assessed by computer-assisted sperm analyzer), and acrosome intactness in Tris dilutions kept at constant 0℃ for 1, 3, 6, 9, and 12 days. We compared them to Tianshan livestock dilutions (Commercial diluent). In experiment 3, semen was diluted using Tris diluent, and sperm quality was measured before and after long-distance transport. Artificial insemination of 177 Simmental heifers compared to 156 using Tianshan Livestock dilution. Results: The outcomes demonstrated that 1:9 was the ideal Tris diluent dilution ratio. The sperm viability, Progressive Motility, and acrosome integrity of both Tris and Tianshan dilutions preserved at 0℃ gradually decreased over time. sperm viability was above 50% for both dilutions on d 9, with a flat rate of decline. The decrease in acrosome integrity rate was faster for Tianshan livestock dilutions than for Tris dilutions when stored at 0℃ for 1 to 6 days. There was no significant difference (p>0.05) in sperm viability between semen preserved in Tris diluent after long-distance transportation and semen preserved in resting condition. The conception rates for Tris dilution and Tianshan livestock dilution were 49.15% and 46.15% respectively, with no significant difference (p>0.05). Conclusion: This shows that Tris diluent is a good long-term protectant. It has been observed that fresh semen can be successfully preserved for long-distance transport when stored under 0℃ conditions. Additionally, it is feasible to distribute semen regionally.

• Asian-Australasian Journal of Animal Sciences
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• 제19권11호
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• pp.1561-1565
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• 2006

The objective of this study was to investigate the anti-oxidative effects of taurine on sperm characteristics for in vitro storage of boar semen. Semen was randomly divided into 10 groups in conical tubes and treated with different concentrations of taurine (25-100 mM) with or without $250{\mu}M$ $H_2O_2$. The percentage of motile spermatozoa in taurine groups after 6 and 9 h were significantly higher at >94% and 87%, respectively, compared to the control group ($85.1{\pm}0.5$ and $72.4{\pm}0.3$, p<0.05). The sperm motility in taurine with $H_2O_2$ after 6 h incubation was slightly decreased compared to the taurine alone treatment, but after 9 and 12 h incubation % sperm motility dropped sharply in taurine with $H_2O_2$ ($75.3{\pm}0.3$ and $69.6{\pm}2.9$, p<0.05). For 3, 9 and 12 h incubation, sperm viability in the control was lower than in taurine groups, irrespective of taurine concentration. In eosin Y and nigrosin staining (ENS), the sperm survival rates (%) for 6 h incubation were significantly higher in 25 mM ($76.0{\pm}0.6$) and 50 mM taurine groups ($78.0{\pm}0.7$), respectively. Sperm survival rates for 9 and 12 h incubation were higher in taurine groups (${\geq}48%$ in 9 h and ${\geq}42%$ in 12 h) compared to controls ($43.0{\pm}2.1$ and $31.0{\pm}0.6$, respectively). In the hyoosmotic swelling test (HOST), sperm membrane integrity was similar to the results of sperm survival. These experiments indicate that supplementation of taurine to the semen extender can increase the sperm characteristics(motility, viability, survival and membrane integrity).

• Asian-Australasian Journal of Animal Sciences
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• 제27권10호
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• pp.1417-1425
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• 2014

The selection of morphologically normal spermatozoa is critical to obtain high breeding performances in boar breeding farms and artificial insemination (AI) centers. Parameters for the selection of semen mainly include total sperm motility, concentration, and morphology. However, these primary parameters are often not reliable for discriminating between normal and abnormal, non-fertilizable spermatozoa. The present study was designed to compare the motion characteristics, fertilization ability using in vitro fertilization (IVF), and acrosome formation of the semen from boars having low (boar number 2012) and normal (boar number 2004 and 2023) breeding performances. The ultimate goal was to identify additional simple and easy criteria for the selection of normal sperm. There was no significant difference between boar 2004 and boar 2023 sperm total motility in computer assisted sperm analysis. However, boar number 2012 semen presented a significantly reduced population of rapid moving spermatozoa and an increased population of slow moving spermatozoa compared to boar numbers 2004 and 2023. Analysis of detailed motion characteristics revealed that sperm from boar number 2012 had significantly reduced motility in progressiveness, average path velocity, straight-line velocity (VSL), curvilinear velocity (VCL), straightness, and linearity. The assessment of the fertilizing ability by IVF also showed that sperm from boar number 2012 showed a fertility rate of 3.4%, whereas sperm from boar number 2023 had a fertility rate of 75.45%. Interestingly, most of the sperm nuclei were found on the peripheral area of the oocytes, suggesting that the sperm from boar number 2012 lacked penetration ability into the oocyte zonapellucida. The acrosome formation analysis using Pisum sativum agglutinin staining demonstrated that the sperm from boar number 2012 had a defect in acrosome formation. Consequently, primary parameters for selecting semen before AI such as motility are not sufficient to select normal and fertilizable spermatozoa. In conclusion, the present study suggests that the acrosome staining and detailed motion characteristics such as progressiveness, VCL, and VSL should be included in determining semen quality together with primary parameters for successful AI and high breeding performance in the swine industry.

• Asian-Australasian Journal of Animal Sciences
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• 제5권3호
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• pp.449-454
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• 1992

Eight healthy Holstein bulls, 4-6 years old were used to study the effect of season of the year on the incidence of the morphologically abnormal spermatozoa. Semen was collected twice a week by A. V. over one-year period. The percentage of total abnormal spermatozoa was $14.1{\pm}0.5$. Ejaculates collected during hot summer season had significantly higher incidence of abnormal spermatozoa than those collected during winter time. Warm spring had moderate semen abnormality. In addition to its effect on the total number of morphologically abnormal spermatozoa, season affected significantly the primary as well as the secondary types of abnormalities. The differences between incidence of primary and secondary types of abnormalities were not significant for all seasons and seasons pooled together. The ratio between the total forms of abnormality in the head, mid-piece and tail of spermatozoa was as 1:1.5:1. Head and mid-piece had defected more during summer compared with both winter and spring. There was no variation in tail abnormalities due to season. The significant effect of season on head was observed by large, pyriform, free and detached heads, while that on mid-piece was by swollen, coiled mid-pieces and protoplasmic droplets.

• Animal Bioscience
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• 제35권11호
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• pp.1666-1674
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• 2022

Objective: Letrozole, a potent aromatase inhibitor, is known to have the potential to modify male reproductive function by altering sex hormone levels. This study aimed to evaluate the semen and testicular characteristics and hormonal profile of aged Mrakhoz bucks (Capra hircus) treated with letrozole. Methods: Twelve Markhoz male goats, aged between 4.5 to 5.5 years with an average body weight (BW) of 61.05±4.97 kg were used for the study. Animals were randomly divided into two equal groups and subcutaneously received either 0.25 mg/kg BW of letrozole or a control every week for 2 months. The semen collections were performed every 10 days, and blood samples and testicular biometric records were collected at 20 days intervals. Results: Letrozole causes increased testosterone and follicle-stimulating hormone levels, testosterone to estradiol ratio, semen index and reaction time during the period from 20th to 60th days (p<0.05). Furthermore, letrozole-treated bucks had higher semen volume, sperm concentration, and total sperm per ejaculate from 30th to 60th days (p<0.05). However, no differences occurred between the groups in scrotal circumference, relative testicular volume, semen pH, abnormality, acrosome integrity, and membrane integrity of sperm during the study (p>0.05). The serum luteinizing hormone levels, sperm viability, motility, and progressive motility increased, and estradiol levels decreased after 40th to 60th days of letrozole treatment (p<0.05). Conclusion: Letrozole application to aged Markhoz bucks provokes reproductive hormonal axis which, in turn, induces enhancement of semen production and quality.

• 한국발생생물학회지:발생과생식
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• 제19권4호
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• pp.253-258
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• 2015

Spermiation was stimulated in the mature grass puffer, Takifugu niphobles, with an injection of human chorionic gonadotropin (HCG) or carp pituitary extract (CPE). Spermatocrit and sperm density were reduced, but milt production was increased in both the HCG and CPE treatment groups relative to those in the control group (P < 0.05). These results should be useful for increasing the fertilization efficiency in grass puffer breeding programs.

• Asian-Australasian Journal of Animal Sciences
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• 제22권5호
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• pp.626-635
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• 2009

An experiment was conducted to investigate the effect of Butylated Hydroxy Anisole (BHA), Butylated Hydroxy Toluene (BHT), Pentoxifylline (PTX), Theophylline (TPY) and Theobromine (TBR) on cold protection ability of Murrah buffalo semen at room ($22-25^{\circ}C$) and refrigerated temperature ($4-7^{\circ}C$). Each semen sample was divided into six parts of equal volume and sperm concentration; the first was kept as a control and the remaining five were treated with BHA, BHT, PTX, TPY or TBR. Sperm motility, abnormal spermatozoa, live-dead count, hypo-osmotic swelling and acrosomal integrity were studied at room and refrigerated temperature for various incubation periods viz.; 0, 4, 8, 12 and 24 h at room and 0, 12, 24, 36, 48, 60 and 72 h at refrigerated temperature. Significant improvement in sperm motility, live-dead count, hypo-osmotic swelling and acrosomal integrity were observed in BHT, PTX and TPY fortified extender at room and refrigerated temperature for various incubation periods. From the present study it could be concluded that cold protection ability of buffalo semen can be improved through the addition of BHT followed by PTX and TPY.