• Clinical and Experimental Reproductive Medicine
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• 제23권3호
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• pp.351-355
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• 1996

Objective: To analyze the direct effect of nitre oxide, generated from sodium nitroprusside, on sperm motility and reactive oxygen species. Design: Human sperm samples were treated to allow swim-up and washing. And the samples were devided into four aliquots. Each aliquot was incubated with either concentration at 0, 100nM, $10{\mu}M$, 1mM of nitroprusside. Intervention: Samples were measured chemiluminosence for reactive oxygen species of each aliquot with concentrations at 0, 100nM, $10{\mu}M$, 1mM of nitroprusside at allowing swim-up and washing of sperm. Main Outcome Measures: Percent motion parameters and viability were asse-ssed at 0, 3, 6, 12, 24 hours incubation. Results: The percent viablity was lower slightly in control group (50.2%) than that in sperm treated with 100nM of nitroprusside(57.5%) at 24 hours after incubation, while was reduced significantly in sperm with concentra-tion of $10{\mu}M(42.1%)$ and 1mM(21.3%)of nitroprusside at 6 hours after incubation. And the sperm treated with 1mM of nitroprusside was immotile totally at 6 hours after incubation. The straight line$(35.3{\pm}5.6%)$, the rapid forward$(37.2{\pm}6.4%)$ and the weak curvilinear velocity$(9.6{\pm}2.4%)$were more favorable comparing with those ($32.4{\pm}4.2%$, $30.0{\pm}7.8%$ and $18.0{\pm}4.6%$ respectively) in control group at 3 hours after incubation, but reduced significantly in sperm treated with $10{\mu}M$ and 1mM of nitroprusside. The levels of reactive oxygen species in control(700 c.p.m.) is lower significantly than that in each experimental groups of sperm treated with nitroprusside. And the levels of reactive oxygen species were 2200 c.p.m. in 100nM, 6200c.p.m. in $1{\mu}M$ and 12800c.p.m. in 1mM respectively. Conclusion: These results suggested that the concentration of 100nM of nitroprusside on sperm is beneficial to the maintanance of viablity and motile velocity, but detriment in high concentration of $10{\mu}M$ or 1mM of nitroprusside.

• Reproductive and Developmental Biology
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• 제35권4호
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• pp.479-483
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• 2011

The objective of this study was to determine the effects of E. coli isolated from porcine semen on sperm viability, motility, and semen pH. Semen samples were prepared using commercial extender, $Seminark^{Pro}$ (Noahbio Tech, Korea) that did not contain antibiotics. And 4 different levels of E. coli were artificially innoculated to semen with following concentrations; 4,000 of sperms with 1 of E. coli (T1), 400 with 1 (T2), 40 with 1 (T3), and 4 with 1 (T4). Semen samples were preserved at $17^{\circ}C$ for 5 days in semen storage box until analyzed by flowcytometer. Aliquots were subjected to measure the sperm viability (Live/$Dead^{(R)}$ stain), motility (mitochondrial function), and semen acidity (pH) from day 0 (day of semen collection) to day 5. Sperm motility and viability were significantly decreased (p<0.05) on day 0 (4 hrs after preservation at $17^{\circ}C$) in T3 and T4 compared to control groups and were significantly decreased (p<0.05) in all groups from day 3. Sample pH was acidic in T3 (6.90~6.86) and T4 (6.86~6.65) from day 3 to day 5 (p<0.05). On the other hand, sample pH was maintained 7.0~7.1 in control, T1, and T2 during the experimental period. Sperm motility and viability were significantly decreased from day 0 to day 5 compared to control in samples contaminated with E. coli above a value of 40:1 ($20{\times}10^6$ sperm cells/ml : $5{\times}10^5$ cfu/ml). Even on day 1 in T4 and on day 3 in T3, semen pH was acidic probably due to the acidification of dead spermatozoa. These results suggest that E. coli contamination has a concentration-dependent detrimental effect on extended porcine semen quality.

• Advances in nano research
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• 제11권6호
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• pp.649-655
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• 2021

Nanotechnology is widely considered a major technology of the twenty-first century. Nanoparticles (NPs) has been shown to pass through reproductively significant biological barriers such as the blood-testicle and placental barriers. Thus, the purpose of this study was to determine the effect of silver Nanoparticles (Ag-NPs) on sperm-egg interaction and spermatozoa quality parameters in quail spermatozoa. Semen was suspended in Ringer solution containing Ag-NPs levels at 5.5 × 106 sperm/ml (0, 0.01, 0.1, 1 and 10 ppm). The results indicated that when sperm were counted at 0.1 ppm, the number of holes formed on the inner perivitelline layer was significantly increased compared to the control. The 10 ppm group had a significant reduction in sperm viability. At 0.1 and 1 ppm, the membrane integrity was significantly decreased (P < 0.05). All treatments (except 0.01 ppm Ag-NPs) had a significant (P < 0.05) effect on the percentage of spermatozoa with an intact acrosome when compared to the control group. At 0.1, 1, and 10 ppm Ag-NPs, morphological defects in the acrosome were observed. As a result, Ag-NPs is likely capable of destroying the acrosome membrane. This research indicates that Ag-NPs may be cytotoxic to spermatozoa by impairing sperm functionality and increasing sperm mortality.

• Advances in nano research
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• 제9권1호
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• pp.15-24
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• 2020

Iron has a crucial role in growth as part of metalo-proteins like haemoglobin or myoglobin, enzymes; they are also involved in energetic reactions. Iron plays a vital role in fertility. At high doses, Iron has a harmful consequence on the reproductive system, which can be strongly reflected the final stage of spermatogenesis. Nutritional products are claiming to use nanotechnology and it is important to recognize the potential toxicity of nano-sized nutrients. Recently iron nanoparticles were proposed as a food additive for poultry. The objective of this study was to investigate the effects of L-cystein coated iron oxide nanoparticles on reproductive performance in male quails. The results of Fourier Transform Infrared Spectrometer, Alternating Gradient Force Magnetometer and Scaning Electron Microscopy showed that iron oxide nanoparticles was produced and have been coated with L-cycstein (Fe₃O₄-Cys NPs). A total of 100 one-week-old quail chicks were randomly placed to five groups of five replicates. Four quails (two male and two females) were raised in an individual cage for each replicate. The five experimental treatment diets consisted; negative control diet, with no Iron supplementation; positive control diet supplemented with 60 mg/kg of Fe₃O₄; treatment diets supplemented with 0.6, 6 and 60 mg/kg of L-cystein coated iron oxide nanoparticles. The hemoglobin, Red blood cell, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, gonadal somatic index, daily sperm production, total testicular sperm and sperm viability of the male quails that were fed with diet supplemented by 0.6 mg/kg of Fe₃O₄-Cys NPs were improved as compare with negative control. This study showed that not only the use of the Fe₃O₄-Cys nanoparticles had no side effects but also it can be used as a feed additive to improve the reproductive performance in male quails.

• Asian-Australasian Journal of Animal Sciences
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• 제21권3호
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• pp.340-345
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• 2008

Oxidative stress is one of the major causes of failure of in vitro storage of boar semen. Reactive oxygen species (ROS) are one of the important mediators of oxidative stress during in vitro storage of boar semen. Our study examined the effects of taurine on sperm characteristic and on in vitro developmental embryos during in vitro storage of boar semen for 7 days. Semen was randomly aliquoted into 3 centrifuge tubes and treated with different concentrations of taurine (25-100 mM). The characteristics of boar sperm were analyzed for motility by light microscopy, viability by using a Makler counting chamber and membrane integrity by a hypoosmotic swelling test (HOST). The percentages of motile spermatozoa in taurine groups after 5 days were significantly higher compared to the control. Sperm viability in the control was lower than in taurine groups after 7 days irrespective of different taurine concentration. In the hyoosmotic swelling test (HOST), significantly higher results were obtained in taurine groups after 3 days. Also, the developmental rates of IVM/IVF porcine embryos from semen treated with pyruvate and taurine were significantly increased when compared with the control (p<0.05). These results indicate that supplementation of taurine as an antioxidant in boar semen extender can improve the semen quality.

• 한국임상수의학회지
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• 제34권2호
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• pp.156-160
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• 2017

We investigated the effect of ethylene glycol and antioxidants such as taurine, hypotaurine and trehalose with extenders during cryopreservation of Korean Jeju Black Bull spermatozoa. The cryopreservation of freshly collected spermatozoa was conducted with four different conditions. As a control, spermatozoa were cryopreserved with Tris egg-yolk extenders added 5% ethylene glycol (EG). Taurine (20 mM), hypotaurine (20 mM) and trehalose (20 mM) were individually added into tris egg-yolk extenders with 5% EG. After thawing of frozen spermatozoa with four different conditions, sperm viability, motility, acrosomal integrity, and membrane integrity were investigated. The significant (p < 0.05) improvement of sperm viability showed in all antioxidant treated thawed spermatozoa (taurine; $68.1%{\pm}4.4$, hypotaurine; $69.2%{\pm}6.7$ and trehalose; $68.0%{\pm}4.4$) when compared to control ($63.4%{\pm}5.6$). Neither positive nor detrimental effects of three antioxidants were shown sperm motility after thawing. The results of hypo-osmotic swelling test showed that the membrane integrity of taurine, hypotaurine or trehalose treated thawed spermatozoa ($64.1%{\pm}5.4$, $61.5%{\pm}3.7$ and $59.0%{\pm}4.0$, respectively) had significantly (p < 0.05) higher rate of the swollen sperm compared to control ($53.7%{\pm}9.7$). Hypotaurine treated frozen-thawed spermatozoa had siginificantly higher (p < 0.05) F pattern ratio than taurine, trehalose and control treated frozen-thawed spermatozoa. Trehalose added frozen-thawed spermatozoa had significantly higher (p < 0.05) acrosome reaction pattern ratio than taurine and hypotaurine added frozen-thawd spermatozoa. In this study, we found that antioxidants such taurine, hypotaurine and trehalose treatments during cryopreservation process could reduce damage of spermatozoa of Korean Jeju Black Bull and improved sperm capability of fertilization.

• Animal Bioscience
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• 제36권2호
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• pp.209-217
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• 2023

Objective: The conservation of Bali bulls, the Indonesian native breed of cattle, is crucial for cattle breeding in Indonesia. To guarantee the spread of Bali bulls through artificial insemination the quality of the frozen semen must be high. To this end, adding an extender material to semen that increases spermatozoa's survival during cryopreservation is important. Green tea extract (GTE) can be used as cryoprotectant because its high antioxidant activity can help avoid reactive oxygen species formation. Methods: Semen of five Bali bulls from the National Artificial Insemination Center at Singosari, Indonesia was collected routinely twice a week. First, fresh semen inspection was performed to determine the feasibility of using Bali bulls as animal samples. The extender used in this study was Tris-based egg yolk. The samples were divided into four treatments: T0, no GTE added to the extender; T1, 0.05 mg GTE plus 100 mL extender; T2, 0.10 mg GTE plus 100 mL extender; and T3, 0.15 mg GTE plus 100 mL extender. The semen freezing process was conducted according to standard procedures and sperm quality parameters, i.e., sperm motility, viability, abnormalities, and membrane integrity observed pre-freezing and post-thawing. Results: There were significant differences in total motility, progressive motility, viability, and integrity membrane of Bali bull sperm at both pre-freezing and post-thawing after adding GTE into the extender. In contrast, there were no differences in abnormalities among treatments. Conclusion: Adding GTE at a 0.15 mg into 100 mL Tris-based egg yolk extender can improve the quality of cryopreserved Bali bull sperm.

• 한국동물생명공학회지
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• 제39권2호
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• pp.88-94
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• 2024

Background: Using cryovial for freezing dog spermatozoa provides a practical method to increase extended sperm volume and shorten the time required for equilibration by using a simple freezing techniques. The purpose of this study was to determine the optimal thawing condition for dog sperm cryopreservation using cryovials. Methods: For sperm freezing, cryovials with 200 × 10⁶ sperm/mL were cooled after the addition of tris egg yolk extender (TEY) at 4℃ for 20 min, then TEY with 4% glycerol was added and equilibrated for another 20 min before being aligned over LN₂ vapor for another 20 min and plunged directly into LN₂. Spermatozoa were thawed in a water bath at 37℃ for varying times (25 sec, 60 sec, 90 sec, and 120 sec) in the first experiment. In the second experiment, spermatozoa were thawed in a water bath at various temperatures and times (37℃ for 1 min, 37℃ for 1 min with gentle stirring, 24℃ for 24 min, and 75℃ for 20 sec). In these experiments, the effect of thawing conditions on motility parameters, viability (SYBR-14/PI), and acrosome integrity (PSA/FITC) of spermatozoa were investigated. Results: The post-thaw sperm motility parameters, viability, and acrosome integrity were not significantly different across the experimental groups. Conclusions: In this study, the characteristics of spermatozoa frozen using cryovials were not significantly affected by various thawing conditions.

• Asian-Australasian Journal of Animal Sciences
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• 제29권9호
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• pp.1247-1255
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• 2016

Heparin binding proteins (HBPs) are produced by accessory glands. These are secreted into the seminal fluid, bind to the spermatozoa at the time of ejaculation, favour capacitation, acrosome reaction, and alter the immune system response toward the sperm. The present study was conducted with an objective to assess the effect of purified seminal plasma-HBPs (SP-HBPs) on cross bred cattle bull sperm attributes during two phases of cryopreservation: Pre freezing and freezing-thawing. SP-HBPs were purified from pooled seminal plasma by heparin affinity chromatography. Three doses of SP-HBPs i.e. 10, 20, $40{\mu}g/mLs$ semen were standardized to find out the optimum dose and $20{\mu}g/mLs$ was found to be an optimum dose. Semen as such and treated with SP-HBPs was diluted with sodium citrate-egg yolk diluter and cryopreserved as per the standard protocol. Sperm parameters i.e. motility, viability, Hypo-osmotic swelling test (HOST), acrosome damage, in vitro capacitation and lipid peroxidation were evaluated in SP-HBP treated and untreated (control) semen at both phases of cryopreservation. A considerable variation in percent sperm motility, viability, membrane integrity (HOST), acrosome damage, acrosome reaction and lipid peroxidation was observed at both phases among the bulls irrespective of the treatment. Incubation of neat semen with $20{\mu}g/mL$ SP-HBP before processing for cryopreservation enhanced the average motility, viability, membrane integrity by 7.2%, 1.5%, 7.9%, and 5.6%, 6.6%, 7.4% in pre-frozen and frozen-thawed semen in comparison to control. There was also an average increase of 4.1%/3.9% in in vitro capacitation and acrosome reaction in SP-HBPs-treated frozen-thawed semen as compared to control. However, binding of SP-HBPs to the sperm declined acrosome damage and lipid peroxidation by 1.3%/4.1% and 22.1/$32.7{\mu}M$/$10^9$ spermatozoa in SP-HBP treated pre-frozen/frozen-thawed semen as compared to control, respectively. Significant (p<0.05) effects were observed only in motility, HOST and in vitro acrosome reaction. It can be concluded that treatment of neat semen with SP-HBPs before cryopreservation minimized the cryoinjury by decreasing the generation of reactive oxygen species.

• Reproductive and Developmental Biology
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• 제30권3호
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• pp.219-224
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• 2006

본 연구는 내분비계 장애물질인 BPA와 DEHP가 돼지 정액 성상(운동성, 생존율, 원형질막의 정상성, 기형율)에 유해한 영향을 미치는지를 검토하였다. 일정 농도로 희석한 돼지 정액에 BPA와 DEHP를 각각 0, 1, 10, $100{\mu}M$의 농도로 처리하여 3, 6, 9시간 동안 체외배양을 실시하였다. BPA 처리에 따른 운동성과 생존율은 체외배양 시간과 첨가농도에 따라 감소하였으며, 체외배양시간이 경과함에 따라 대조구와 명백한 차이를 나타냈다. $100{\mu}M$의 농도로 처리한 경우의 운동성과 생존율은 체외배양시간에 관계없이 처리군이 대조구에 비해 유의적으로 감소하였다(p<0.05). DEHP처리에 따른 정자의 운동성과 생존율도 체외배양시간과 첨가농도에 따라 감소하여 BPA의 성적과 유사한 경향을 나타냈다. 정자 원형질막의 기능성은 배양시간에 따라 감소하였으며 BPA및 DEHP 첨가농도 간에 차이가 인정되었으며, 특히 $100{\mu}M$ 처리구에서 급격히 감소하는 성적을 나타났다. 정자의 기형율은 BPA및 DEHP 처리시간 및 농도에 크게 영향을 받지는 않았다. 본 연구의 결과로 볼 때, 고농도의 BPA와 DEHP($>10{\mu}M$에 정자의 장시간 노출은 유해한 영향을 줄 것으로 사료된다.