• Clinical and Experimental Reproductive Medicine
• /
• v.36 no.1
• /
• pp.35-43
• /
• 2009

Objectives: Seminal concentration of tumor necrosis factor-alpha (TNF-${\alpha}$) relevant to sperm nuclear DNA integrity has not been studied. The present study aimed to evaluate seminal concentration of TNF-${\alpha}$ in correlation with sperm parameters and nuclear DNA integrity in asymptomatic healthy donors. Methods: Semen samples were obtained by masturbation from forty-five healthy donors. Results: Sperm quality was assessed by computer-assisted semen analysis and nuclear DNA integrity measured by the TUNEL assay in raw semen. TNF-${\alpha}$ concentrations were measured by ELISA in frozen-thawed seminal plasmas. Sperm DNA fragmentation rates were ranged between 1.9% and 53.0% (mean${\pm}$SD, 12.4${\pm}$9.6%). Univariate analysis revealed that DNA fragmentation rate was not associated with sperm concentration or motility but had a correlation with linearity negatively (r=-0.325, p=0.03) and age positively (r=0.484, p=0.001). The mean seminal concentration of TNF-${\alpha}$ was 4.9 pg/mL with a range from 1.1 to 22.6 pg/mL. The TNF-${\alpha}$ concentration had no correlation with clinically relevant parameters of sperm quality or nuclear DNA fragmentation rate. Conclusion: Our results indicate that sperm nuclear DNA fragmentation may be not associated with seminal TNF-${\alpha}$ level or sperm quality in asymptomatic healthy donors.

• Korean Journal of Animal Reproduction
• /
• v.10 no.2
• /
• pp.188-191
• /
• 1986

This experiment was carried out to examine the correlation among the semen characteristics of Holstein bulls. Seventy-two samples of semen were collected from 6 Holstein bulls, 2 to 10 years old. The characteristics of the semen were estimated by ordinary methods. The results obtained in this experiment were summarized as follows: 1. Semen characteristics on an average were 6.0 ml of volume, 16${\times}$108/ml of concentration, 92.4${\times}$108 of total sperm number(TSN), 72.1% of motility and 4.6% of abnormality. 2. Semen volume was positively high correlated (P<0.01) with total sperm number, but negative correlation was observed with the sperm motility (P<0.05) and semen concentration. 3. In relation to their age, older bulls semen volume and total sperm number were increased (P<0.01), but semen concentration and motility were decreased (P<0.01).

• Asian-Australasian Journal of Animal Sciences
• /
• v.8 no.2
• /
• pp.123-127
• /
• 1995

In vitro embryo production (IVP) is affected by various factors during in vitro maturation, fertilization, and development. In this experiment, the effect of ovary type, quality of follicular oocyte, medium used for fertilization, presence of hormone in medium, sperm concentration on in vitro maturation and fertilization were examined for effective IVP. In vitro maturation was carried out using TCM-199 supplemented with 15% FCS and hormones in 5% $CO_2$ incubator for 24h. In vitro fertilization was performed with frozen-thawed sperm in modified mTALP medium containing 0.3% BSA, $10{\mu}g/ml$ heparin, and 5mM/ml caffeine for 24h. The fertilized embryos were co-cultured on monolayer of cumulus cells in TCM-199. When oocytes were collected from functionally active and inactive ovaries, maturation rate was 76.9 and 7.7%, respectively. When oocytes were classified morphologically to good and poor grades, maturation rate was 75 and 58.8%, respectively. FSH + LH + $E_2$ (86.4%) showed higher maturation rate than control (53.0%) and FSH (73%). The fertilization rate was 28.2, 100 and 91.7% in $1.6{\times}10^5$, $5.0{\times}10^5$ and $10.0{\times}10^5$ sperm concentration per ml. When oocytes were fertilized in mTALP and BO media, fertilization and cleavage rates of oocytes in mTALP were higher (84.3 and 56.9%) than those (67.4 and 23.3%) in BO medium. In this experiment, in vitro maturation, fertilization and development of oocytes were affected by type of ovary, grade of oocyte, hormones, sperm concentration and fertilization medium.

• Animal Bioscience
• /
• v.34 no.2
• /
• pp.192-197
• /
• 2021

Objective: The present study evaluated the preservation of ram semen at 0℃ using soybean lecithin with a Tris-fructose extender. Methods: Semen was collected by artificial vagina ejaculation from six rams with proven fertility. High quality ejaculates were diluted by soybean lecithin (0.25%, 0.5%, 0.75%, 1.0%, 1.25%) using Tris-fructose extender and control (Tris-fructose egg yolk extender), respectively. The ejaculates were diluted to a concentration of 5×108 sperm/mL, followed by cooling to 0℃ in 90 min and maintaining the temperature for 12 days. The diluted semen samples were examined and recorded for sperm progressive motility, acrosome integrity at 0, 24, 72, 144, 216, 288 h, respectively. Two hundred and twenty-three ewes were inseminated for 216 h with optimal soybean lecithin concentrated semen or control via trans-cervical insemination. Results: The results showed that there were no differences in sperm progressive motility at 0, 24, 72, and 144 h (p>0.05). After 216 h, the sperm progressive motility in the control group and 0.5% concentration groups was significantly higher when compared to 0.25% concentration (p<0.05). The 0.5% concentration group demonstrated the highest survival rate and had no difference with the control group (p>0.05). At 216 h, the sperm progressive motility of all groups was still above 50%. The acrosome integrity of all groups was decreased with prolongation of storage time, but there was no difference at each time point (p>0.05). There was no significant difference in the lambing rate and pregnancy rate between the 0.5% concentration group and the control group (p>0.05). Conclusion: These results suggest that ram sperm is capable of fertilization after preservation at 0℃ with 0.5% of soybean lecithin in Tris-based extender substituted for egg yolk and produce normal offspring after insemination.

• Proceedings of the KSAR Conference
• /
• 2001.10a
• /
• pp.57-57
• /
• 2001

This study was carried out to investigate the general characteristics such as volume, sperm concentration, sperm motility, sperm abnormality on whole semen, RSP-S and RSP-T semen and fractional semen of small size dogs, and the effect of temperature and preservation time and cryoproservation on motility of whole and RSP-S and RSP- T semen. Multiple ejaculates were collected from small dogs by the digital manipulation of penis. 1. The volume per ejaculate semen, sperm of concentration and motility and abnormal sperm rate of 1st fractional semen were 0.65±0.09㎖, 4.52±0.35×10/sup 6/ cells/㎖, 15.64±3.85% and 5.50±0.62%. Also, 2nd fractional semen were 1.25±0.20㎖, 3.35±0.48×10/sup 6/cells/㎖, 96.25±4.65% and 4.24±0.46%. And 3rd fractional semen were 1.45±0.21㎖, 3.85±0.52×10/sup 6/cell/㎖, 92.82±4.24% and 4.66±0.58%, respectively. 2. The sperm of concentration and motility and abnormal sperm rates of whole, RSP-S and RSP-T semen were 5.45±0.82×10/sup 6/ cells/㎖, 95.55±4.65%, 4.58±0.45% and 4.82±0.36×10/sup 6/cells/㎖, 90.10±3.42%, 6.48±0.68% and 4.55±0.45× 10/sup 6/cells/㎖, 93.25±3.85%, 4.82±0.58%, respectively. 3. The motility of whole, RSP-S and RSP-T semen were higher at 4℃ than at 38℃. When preservation temperature was at 4℃, survival rates of RSP-S and RSP-T sperm were 97.54%-6.25% at 1-72 hrs, 97.40%-5.62% at 1-100 hrs, respectively. 4. The survival rates of slow and rapid frozen 2nd fraction, RSP-S and RSP-T semen were 67.3±4.45%, 88.8±4.46% and 46.4±3.84%, 74.4±4.20%, respectively. Survival rates was significantly higher in frozen RSP-S and RSP-T semen than that in control group(8.5±2.12%).

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.32 no.2
• /
• pp.238-241
• /
• 1999

Experiments were performed to find out the physico-chemical properties of milt and the sperm motilities in various conditions using the grey mullet, Mugil cephalus. The average concentration of sperm in the milt was $1,11 \pm0.36\times10^{10}/ml$. Spermatocrit was 96.7$\pm$2.6. pH and osmolality of seminal fluid were 7.8$\pm$0.1, 370$\pm$6 mOsm/kg, respectively, Total protein concentration of sperm was higher than that of seminal fluid, but total lipid concentration of seminal fluid was higher than that of sperm. The sperm motility was high in the diluent of milt : artificial seawater (1:10, by volume) and in 822 mOsm/kg and 983 mOsm/kg similar to seawater osmolality, but it decreased after 20 minutes. But activity of sperm was highly maintained in 482 mOsm/kg which was a little higher than osmolality of seminal fluid, and was high in pH 7$\~$9.

• Korean Journal of Animal Reproduction
• /
• v.14 no.4
• /
• pp.245-252
• /
• 1990

This study was carried out to investigate the factors affecting fertilization in vitro of follicular oocytes with frozen-thawed spermatozoa in Korean Native Cattle. The bovine ovaries were obtained at a slaughter house and the follicular oocytes were recovered by aspirating the follicular fluid from the visible follicles of 3~6mm. The bovine oocytes were matured in vitro for 20~24 hours in TCM-199 containing FCS and hormones. The matured oocytes were fertilized in vitro using Percoll-separated frozen-thawed spermatozoa in BO solution. The effects of dilution and fertilization media, capacitating method, concentration of inseminated sperm and time after insemination of fertilization, were observed. The results obtained are summarized as follows : 1. The fertilization rate of frozen-thawed sperm inseminated in BO solution with caffeine and heparin together(56.4%) was higher than that of sperm inseminated in BO solution with either caffeine(10.5%) or heparin(8.9%) and without both caffeine and heparin(0%)(P<0.05). 2. The fertilization rate(56.3%) of frozen-thawed sperm inseminated in BO solution with both caffeine and heparin without preincubation was higher than that of sperm preincubated(2.9%)(P<0.05). 3. The fertilization with high concentration of frozen-thawed sperm(1.4~1.8$\times$107cells/ml) in BO solution containing caffeine and heparin resulted in higher fertilization rate, 76.7%, than the low concentration of sperm(0.8~1.0$\times$107cells/ml), 32.7%(P<0.01). 4. When the oocytes were inseminated with frozen-thawed sperm in BO solution containing caffeine and heparin without preincubation, fertilization rate increased by time and the rates were 5.9, 46.0 and 59.4% at 8, 16 and 24 hours, respectively.

• Proceedings of the KSAR Conference
• /
• 2003.06a
• /
• pp.63-63
• /
• 2003

This study was carried out to investigate the effects of liquid boar sperm stored at 4$^{\circ}C$ on sperm motility, normal acrosome, and in-vitro fertilization and culture of pig oocytes matured in-vitro. The sperm-rich fraction (30~60 ml) of ejaculate was collected into an insulated vacuum bottle. Semen was slowly cooled to room temperature (20~23$^{\circ}C$) by 2 h after collection. Semen was transferred into 15 ml tubes, centrifuged at room temperature for 10 min at 800$\times$g, and the supernatant solution was poured off. The concentrated sperm was resuspended with 5 ml of lactose, egg yolk and N-acetyl-D-glucosamine (LEN) diluent to provide 1.0$\times$10$^{9}$ sperm/ml at room temperature. The resuspended semen was cooled in a refrigerator to 4$^{\circ}C$ and preserved for 5 days to examine sperm motility and normal acrosome. The medium used for oocyte maturation was modified tissue culture medium (TCM) 199. After about 22 h of culture, oocytes were cultured without cysteamine and hormones for 22 h at 38.5$^{\circ}C$, 5% $CO_2$ in air. Oocytes were inseminated with liquid boar sperm stored at 4$^{\circ}C$ for 2 days after collection. Oocytes were coincubated for 6 h in 500 ${mu}ell$ mTBM fertilization media with 0.2, 1, 5 and 10$\times$10$^{6}$ /ml sperm concentration, respectively. At 6 h after IVF, oocytes were transferred into 500 ${mu}ell$ Hepes-buffered NCSU-23 culture medium for further culture of 6, 48 and 144 h. There were significant differences in sperm motility and normal acrosome among preservation days and incubation times, respectively. The rates of sperm penetration and polyspermy were higher in 5 and 10$\times$10$^{6}$ sperm/ml than in 0.2 and 1$\times$10$^{6}$ sperm/ml. Male pronuclear formation was lower in 0.2$\times$10$^{6}$ sperm/ml than in 1, 5 and 10$\times$10$^{6}$ sperm/ml. Mean numbers of sperm in penetrated oocyte were highest in 10$\times$10$^{6}$ sperm/ml compared with other sperm concentrations. The rate of blastocysts from the cleaved oocytes (2~4 cell stage) was highest in 1$\times$10$^{6}$ sperm/ml compared with other sperm concentrations. In conclusion, we found out that liquid boar sperm stored at 4$^{\circ}C$ could be used for in-vitro fertilization of pig oocytes matured in-vitro. Also, we recommend 1$\times$10$^{6}$ ml sperm concentration for in-vitro fertilization of pig oocytes.

• Clinical and Experimental Reproductive Medicine
• /
• v.15 no.1
• /
• pp.10-16
• /
• 1988

Artificial insemination with donor sperm(AID) or husband sperm(AIH) has been a major form of treatment for the infertile couples with severe male factors. The conception rate in oligoasthenoteratozoospermia is very low. Therefore the aim of this study was to examine those factors associated with improved success rate. The results are obtained as the follows; 1. The husband semenalysis(n=639) revealed normal findings in 32.2%. The abnormal findings included the oligospermia(18.5%), oligoasthenospermia(20.7%), oligoasthenotetatozoospermia(8.5%) and azoospermia(20.1 %). 2. The causes of abnormal semenalysis are idiopathic(most common), varicocele, congenital anomalies, vaso-vasostomy, etc.. 3. The semen washing to improve sperm motility and concentration was effective in case of over 20 ${\times}lO^6$/ml sperm count, but there was no significant improvement in case of be low 10-5${\times}lO^6$/ml sperm count. 4. The improvement of motility after cryopreservation depended on the initial sperm concentration. 5. The pregnancy rates following AIH are higher in normospermia than oligospermia.

• Korean Journal of Veterinary Research
• /
• v.32 no.3
• /
• pp.435-450
• /
• 1992

To evaluate the fertilizing capacity of domestic animal spermatozoa by hamster test, semen were collected from 15 boars(Duroc, Landrace, and Yorkshire) and 2 mixed dogs which had been proved to be fertile in the past then, the semen were preserved in BWW medium at $4^{\circ}C$ or $18^{\circ}C$ for about 20 hours and coincubated with zona-free hamster ova for 5 hours. The ova were stained by lacmoid and examined under phase contrast microscope to investigate the rates of sperm binding to the ova, penetration and formation of a male pronucleus, and the numbers of both bound and penetrated sperm per ovum. Both the semen preserved at $18^{\circ}C$ for about 20 hours and that treated by swim up procedure showed considerably higher rates of sperm binding and penetration as well as higher number of penetrated sperm than that preserved at $4^{\circ}C$ for about 20 hours, respectively(p<0.01). Motility of boar sperm at insemination was from 40 to 90% and no difference in hamster test was obtained according to different degree of sperm motility. Abnormality in morphology of boar sperm at insemination was from 6 to 45% and no difference in hamster test was obtained according to different degree of sperm abnormality. The sperm concentrations of $7{\times}10^7$ and $7{\times}10^6$ showed considerably higher rates of sperm binding and penetration as well as higher number of bound sperm than that of $7{\times}10^4$ (p<0.01) along with the same higher results than that of $7{\times}10^5$(0<0.05), respectively. Boar sperm showed considerably higher rates of sperm binding and penetration as well as higher numbers of both bound and penetrated sperm than dog sperm, when both semen were treated by BWW＋heparin medium and swim up procedure, respectively. These results indicated that fertile boar sperm showed considerably lower rates in the results of hamster test, when preserved at $4^{\circ}C$ for about 20 hours and in lower concentration of sperm than when preserved at $18^{\circ}C$ for about 20 hours and in higher concentration of sperm, respectively, and at the same time considerably higher results than fertile dog sperm, consequently to prove that hamster test would be of great value in assaying the fertilizing capacity of boar sperm.