• Journal of Preventive Medicine and Public Health
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• v.43 no.2
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• pp.131-137
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• 2010

Objectives: Bisphenol A diglycidyl ether (BADGE) is a liquid compound obtained by condensation of two molecules of epichlorohydrin with one molecule of bisphenol A. General and reproductive toxicity with BADGE has been reported higher than 1000 mg/kg/day. This study was performed to show the effects of acute exposure to BADGE below 1000 mg/kg/day on the testis in adult male rats. Methods: BADGE was administered by gastric lavage in a single dose of 500, 750, 1000, and 2000 mg/kg/day in 8-week old male SPF Sprague-Dawley rats. The right testis was processed for light microscopic analysis. The left testis was homogenized and spermatids were counted to determine the daily sperm production and daily abnormal sperm production. The sperm count, sperm motility, and incidence of abnormal sperm were estimated in the epididymis. In testicular sections, the seminiferous tubules were observed for qualitative changes. The progression of spermatogenesis was arbitrarily classified as full-matured, maturing, and immature. The specimen slide was observed at 3 points and 10 seminiferous tubules were evaluated at each point. Results: The male rats exposed to single oral dose of BADGE at 750, 1000, and 2000 mg/kg/day were significantly increased the number of immature and maturing sperm on the testis. There were no significant differences with respect to sperm head count, sperm motility, and sperm abnormality in the BADGE treatment groups. Conclusions: These results suggest that single oral exposure of BADGE 750 mg/kg/day can affect adult male testis development.

• Journal of Embryo Transfer
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• v.31 no.4
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• pp.355-365
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• 2016

The cryopreservation of sperm has become the subject of research for successful artificial insemination technologies. Antifreeze proteins (AFPs), one of the factors necessary for effective cryopreservation, are derived from certain Antarctic organisms. These proteins decrease the freezing point of water within these organisms to below the temperature of the surrounding seawater to protect the organism from cold shock. Accordingly, a recent study found that AFPs can increase the motility and viability of spermatozoa during cryopreservation. To evaluate this relationship, we performed cryopreservation of boar sperm with AFPs produced in the Arctic yeast Leucosporidium sp. AFP expression system at four concentrations (0, 0.01, 0.1, and $1{\mu}g/ml$) and evaluated motility using computer assisted sperm analysis. DNA damage to boar spermatozoa was measured by the comet assay, and sperm membrane integrity and acrosome integrity were evaluated by flow cytometry. The results showed that motility was positively affected by the addition of AFP at each concentration except $1{\mu}g/ml$ (p<0.001). Although cryopreservation with AFP decreased the viability of the boar sperm using, the tail DNA analyses showed that there was no significant difference between the control and the addition of 0.1 or $0.01{\mu}g/ml$ AFP. In addition, the percentage of live sperm with intact acrosomes showed the least significant difference between the control and $0.1{\mu}g/ml$ AFP (p<0.05), but increased with $1{\mu}g/ml$ AFP (p<0.001). Our results indicate that the addition of AFP during boar sperm cryopreservation can improve viability and acrosome integrity after thawing.

• Reproductive and Developmental Biology
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• v.40 no.4
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• pp.33-37
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• 2016

This study was conducted to evaluate effect of ${\alpha}$-linolenic acid (ALA) and bovine serum albumin (BSA) on viability, acrosome reaction and mitochondrial intact in frozen-thawed boar sperm. The boar semen was collected by gloved-hand method and cryopreserved using freezing extender containing 3 ng/mL ALA and/or $20\;{\mu}g/mL$ BSA. Cryo-preserved boar sperms were thawed in $37^{\circ}C$ water-bath for 45 sec to analysis. Viability, acrosome reaction, and mitochondrial intact were analyzed using flow cytometry. In results, viability of frozen-thawed boar sperm was significantly higher in only ALA+BSA supplement group than control group (p<0.05), whereas there was no difference either in ALA or BSA supplement. However, acrosome reacted sperm in both of live and all sperm population were significantly decreased in all treatment groups than control (p<0.05). Interestingly, mitochondrial intact of boar sperm was enhanced in ALA and ALA+BSA groups compared with control (p<0.05). In this study, we showed that supplementation of ALA and BSA in freezing extender enhanced the sperm viability, mitochondrial intact and decrease acrosomal membrane damage. In conclusion, our findings suggest that quality of frozen-thawed sperm in mammalians could improve by using of ALA and BSA.

• Clinical and Experimental Reproductive Medicine
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• v.27 no.2
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• pp.151-157
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• 2000

Objective: The purpose of this study was to evaluate the effect of polycystic ovarian follicular fluid on sperm motility in human in vitro fertilization (IVF). Methods: From May, 1998 to July, 1999, 55 patients who complained of infertility were involved in this study. We obtained ovarian follicular fluids from the patients by ultrasono-guided aspiration. Subjects were divided into two groups. 20 patients who had polycystic ovarian disease were belong to study group, and 25 patients who had normal ovarian follicular fluid were belong to control group. The follicular fluid dilution was done with Ham's fluid as 10%, 20%, 50%, 100%. The sperm motility was analyzed by CASA at 6hr and 12hr after incubation in follicular fluids. Results: The levels of average path velocity (VAP) in all concentration fluid didn't show significant difference between study and control group. The other parameters including curvilinear velocity (VCL), amplitude of lateral head displacement (ALH), and linerity (LIN) were didn't show any significant difference between both groups. Conclusion: PCOD fluid had seemed to have an adverse effect on the sperm biological function. But, this study showed that PCOD fluid had no different effect on sperm motility with normal follicular fluid.

• Clinical and Experimental Reproductive Medicine
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• v.42 no.1
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• pp.22-29
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• 2015

Objective: Sperm must be properly prepared in in vitro fertilization (IVF)-embryo transfer (ET) programs in order to control the fertilization rate and ensure that embryos are of high quality and have appropriate developmental abilities. The objective of this study was to determine the most optimal sperm preparation method for IVF. Methods: Patients less than 40 years of age who participated in a fresh IVF-ET cycle from November 2012 to March 2013 were included in this study. Poor responders with less than three mature oocytes were excluded. Ham's F-10 medium or sperm-washing medium (SWM) was used in combination with the density-gradient centrifugation/swim-up (DGC-SUP) or SUP methods for sperm preparation. A total of 429 fresh IVF-ET cycles were grouped according to the media and methods used for sperm preparation and retrospectively analyzed (DGC-SUP/Ham's F-10, n=82; DGC-SUP/SWM, n=43; SUP/Ham's F-10, n=181; SUP/SWM, n=123). Results: There were no significant differences among these four groups with respect to the mean age of the female partners, duration of infertility, number of previous IVF cycles, and retrieved oocytes. We determined that both the DGC-SUP and SUP methods for sperm preparation from whole semen, using either Ham's F-10 or SWM media, result in comparable clinical outcomes, including fertilization and pregnancy rates. Conclusion: We suggest that both media and both methods for sperm preparation can be used for selecting high-quality sperm for assistive reproductive technology programs.

• Journal of Embryo Transfer
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• v.32 no.3
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• pp.249-255
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• 2017

In this experiment, we determined the effect of curcumin supplementation in freezing buffer for miniature pig sperm cryopreservation. Each ejaculate was diluted with modified Modena B extender and mixed with lactose-egg yolk (LEY extender, 80% v/v lactose solution [310 mM], 20% v/v egg yolk, and $100{\mu}g/mL$ kanamycin sulfate) and LEY-glycerol Orvus ES Paste (LEYGO, 89.5% v/v LEY, 5% v/v glycerol, 1.5% v/v Orvus ES Paste), 100 mM trehalose supplemented with 0, 10, 50, 100, and $500{\mu}M$ of curcumin from turmeric, respectively. Following equilibration, the 0.5 mL French straws were frozen and plunged into $LN_2$ tank for 7 days at least. Sperm parameter and oxidative byproducts were determined by the computer assisted sperm motility analysis (CASA) and fluorescence-activated cell sorting (FACS) as compared with each groups. Supplementation of curcumin had no effect on sperm motility, progressive motility and curvilinear velocity. However, average-path velocity and straight-line velocity were significantly higher in $10{\mu}M$ curcumin group ($100.9{\pm}8.8{\mu}m/s$, $61.7{\pm}2.9{\mu}m/s$, respectively) than control group ($77.8{\pm}3.9{\mu}m/s$, $46.4{\pm}3.0{\mu}m/s$, respectively) (p < 0.05). In addition, the level of the O2 radical and H2O2 were comparatively decreased in curcumin groups by evaluation of ethidium and DCF fluorescence. According to the results, curcumin can improve sperm kinetic variables and alleviate ROS induced cryoinjury to pig sperm.

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.3
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• pp.202-208
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• 2022

This study investigated the influence of sodium bicarbonate (NaHCO₃) and progesterone on acrosome reaction and proportion of polyunsaturated fatty acid (PUFA) composition boar sperm. The sperm were diluted with semen extender and incubated with NaHCO₃ and progesterone at 38℃, 5% CO₂ for 6 h. Plasma membrane integrity and acrosome reaction were analyzed using SYBR14/propidium iodide (PI) and FITC-PNA/PI doubling staining method, and proportion of PUFA was analyzed using gas chromatography. In results, Plasma membrane integrity was significantly decreased in 50 mM NaHCO₃ group and acrosome reaction was significantly increased by over the 100 mM NaHCO₃ group compared to control group (p < 0.05). In addition, progesterone significantly increased decreased plasma membrane integrity at 100 mM progesterone and acrosome reaction at over the 5.0 µM progesterone (p < 0.05), but there was no difference among the 5.0 to 100 µM groups. PUFAs were significantly decreased in 100 mM NaHCO₃ and 50 µM progesterone treatments compared to control group. In summary NaHCO₃ and progesterone induce acrosome reaction and reduce PUFA composition in boar sperm, therefore, the results maybe help to understand basically knowledge for the acrosome reaction and PUFA composition in boar sperm.

• Reproductive and Developmental Biology
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• v.30 no.1
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• pp.27-34
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• 2006

This study was conducted to compare the reproduction ability of the wild type boar and recombinant human erythropoietin (hEPO) transgenic boar semen. Ejaculated boar semen was analyzed by flow cytometry, Elisa and IVF methods. In experiment 1, flow cytometric analysis showed that the live sperm ratio of transgenic boar sperm significantly lower (P<0.05) than that of wild type boar after incubation at 20, 22, 24 and 26 hr. In experiment 2, the presence and levels of various cytokines (IL-6, IL-10 and $TNF-{\alpha}$) to related animal reproduction in the seminal and blood plasma were examined using specific enzyme immunoassay. There was no significant difference between both groups. In experiment 3, the fertilizing capacity and developmental ability of both boar sperm were compared. The transgenic boar sperm had a significantly low capacity of penetration, sperm-zona binding, embryo development, and blastocyst formation compared to wild type sperm (P<0.05). These results suggest that transgenic boar sperm harboring hEPO gene has low sperm viability than wild type boar, and it is a reason to decrease of fertility and litter size.

• Journal of Embryo Transfer
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• v.30 no.3
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• pp.121-128
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• 2015

The objective of this study was to evaluate the efficiency of sperm cryosurvival in boar sperm separated by Percoll containing antioxidant enzymes. The boar semen was collected into a pre-warmed ($37^{\circ}C$) thermos bottle by gloved-hand method and was separated by 65% Percoll with superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) before freezing. The frozen sperm was thawed at $38.5^{\circ}C$ for 45 sec in water-bath for sperm characteristic analysis. The sperm were estimated with SYBR14/PI double staining for viability, FITC-PNA/PI double staining for acrosome reaction, Rhodamine123/PI double staining for mitochondrial integrity and were analyzed using flow cytometry. In results, sperm viability, acrosome reaction and mitochondrial integrity were improved in separated sperm groups compared with unseparated sperm by Percoll (UP) group. Especially, viability was significantly higher in sperm separated by Percoll containing 400 IU CAT group compared with other groups (P<0.05). And acrosome reaction was decreased in sperm separated by Percoll with 300 IU SOD, 400 IU CAT and 0.5 mM GSH groups compared with other groups, however, there were no significantly difference mitochondrial integrity among sperm separated by Percoll with antioxidant enzymes. In conclusion, we suggest that use of Percoll containing antioxidant enzymes for sperm separation will be beneficial for sperm cryopreservation in pigs.

• Applied Microscopy
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• v.44 no.1
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• pp.1-7
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• 2014

The spermatogenesis of Siamese fighting fish, Betta splendens, belongs to Osphronemidae was investigated by light and electron microscopic observations. In primary spermatocyte stage, the nucleus was comparatively large ellipsoidal, and mitochondria showed a marked development in cytoplasm. In secondary spermatocyte stage, the germ cells were smaller than that of primary spermatocytes. The nucleus was a spherical shape and intercellular space was formed between germ cells. In spermatid stage, the early spermatids were not much different from a secondary spermatocyte. But, the chromatin condensation was occurred from the outside to the inside. The nucleus was more condensed. Intracellular space was larger than early spermatid. The mitochondria were rearranged in a middle piece, and occupied about half of the head part in early sperm. In sperm stage, the head of mature sperm was a spherical shape and had no acrosome. The flagellum was showed the typical 9+2 array of microtubules. Also, the tail of sperm had no lateral fins and outer coarse fibers. These ultrastructural characteristics can be used in classification of species.