• Journal of Ginseng Research
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• v.34 no.4
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• pp.327-335
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• 2010

We reported previously that the administration of Korean red ginseng water extract (KRG-WE) protected the guinea pig testis against damage induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (a potent endocrine disruptor). We also found that crude saponin from ginseng was the active ingredient responsible for this protection. Here, we examined the biological role of KRG-WE in an animal model of age-induced dysfunction of spermatogenesis. Twenty-four male Sprague-Dawley (six 2-month-old and eighteen 12-month-old) rats were used. The young and old control groups received only vehicle. The ginseng saponin (GS)- and KRG-WE-treated groups received GS (40 mg/kg body weight/day) and KRG-WE (200 mg/kg body weight/day), respectively, for 4 months. The number of cells, Sertoli cell index, Johnsen's score, and sex hormone levels decreased significantly with age. However, the administration of KRG-WE and GS markedly improved the number of germ cells, seminiferous tubular size, and Johnsen's score in the old rats. Ginseng produced a distinct testicular histological improvement in old rats. KRG-WE and GS elevated testosterone levels, while attenuating the aberrant increase in follicle stimulating hormone and luteinizing hormone levels. Sperm kinematics evaluated by a computer-assisted sperm analyzer demonstrated improvement in the percentage of motile sperm, progressive sperm motility, and curvilinear velocity associated with sperm quality, supporting the beneficial role of red ginseng in senile spermatogenesis. Overall, the total water extract had a more potent effect than the corresponding saponin fraction. In conclusion, Korean red ginseng rejuvenated age-induced testicular dysfunction. Additionally, the total water extract was more potent than the corresponding saponin fraction.

• Journal of Animal Reproduction and Biotechnology
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• v.38 no.2
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• pp.70-76
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• 2023

Background: Despite considerable technological advancements, polyspermy remains a significant challenge in in vitro fertilization (IVF) procedures in pigs, disrupting normal embryonic development. Here, we aimed to determine whether optimal fertilization conditions reduce the polyspermy incidence in pigs. Methods: In vitro-matured oocytes were co-incubated with sperm according to a modified two-step culture system. Results: In the first experiment, oocytes were briefly co-incubated with sperm, washed in IVF medium, and then moved to fresh IVF medium for 5 or 6 h. Although the 6 h sperm-free cultured group had a higher penetration rate than the 5 h cultured group, the polyspermy rate significantly increased in the 6 h sperm-free cultured group. The gamete co-incubation period was either 20 or 40 min. The 40 min cultured group had a higher rate of blastocyst formation and number of total cells in blastocysts than the 20 min cultured group. In experiment 2, oocytes were inseminated with sperm separated by Pecroll treatment. Percoll treatment increased the rate of oocyte penetration and blastocyst formation compared to the control. In experiment 3, fertilized oocytes were cultured in 25 µL microdroplets (10 gametes/drop) or 500 µL (100 gametes/well) of culture medium in 4-well plates. The large volume of medium significantly reduced the number of dead oocytes and increased the rate of blastocyst formation compared to the small volume. Conclusions: Collectively, these results demonstrate that various fertilization conditions, including modified co-culture period, active sperm separation, and culture medium volume, enhance fertilization efficiency and subsequent embryonic development by decreasing polyspermy occurrence.

• Fisheries and Aquatic Sciences
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• v.4 no.1
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• pp.39-46
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• 2001

To examine the effect of copy number-dependent transgenic genotype on the expression of foreign gene, stable hemizygous and homozygous transgenic breeding line was established using artificial parthenogenesis. For this purpose, induced diploid gynogenetic transgenesis was optimized in mud loach (Misgurnus mizolepis) using UV-irradiated cyprinid loach (M. anguillicaudatus) sperm and thermal shocks. Optimum UV range for inactivation of cyprinid loach sperm was between 3,150 to $4,050\;ergs/mm^2$ The UV-irradiated sperm were inseminated into eggs from recessive color strain (yellow) or heterozygous transgenic mud loach containing CAT gene. Cold shock at $2^{\circ}C$ for 60 min, 5 min post fertilization successfully restored the diploidy of eggs inseminated with UV-irradiated sperm. Restoration to diploidy was confirmed by flow cytometry and gynogenetic status was verified by examining maternal exclusive inheritance of multi-locus DNA fingerprints, body color and transgenic marker. Putative isogenic transgenic fish clearly showed homozygous status at trans gene locus based on Southern blot hybridization and progeny testing. Further, such homozygous gynogenetic diploids revealed the increased levels of transgene expression, when compared to those of heterozygous (hemizygous) transgenic fish.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.12
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• pp.1678-1682
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• 2001

The total proteins were estimated in both deoxycholate (DOC)-extract of sperm membrane and seminal plasma of chilled as well as frozen semen obtained from five Murrah buffalo bulls. Proteins were further characterized by polyacrylamide gel electrophoresis (PAGE) in three bulls. The protein content of sperm membrane extract (SME) and that of seminal plasma (SP) decreased gradually with increase in freezing period from 6 to 24 mo when compared with the values observed in freshly chilled semen in all bulls. The total decrease in protein content of SME and SP varied from 30-40% and 28-59% respectively during 6-24 mo of freezing. The number of glycoproteins/proteins (GP/P) in SME varied from 4-8 in freshly-chilled semen of all bulls and reduced to 2-4 after 24 mo of freezing. In SP, the number of proteins varied from 6-10 in freshly chilled semen of all bulls and reduced to 3-8 after 24 mo of freezing. Some of the proteins in SME and SP disappeared, others got altered and appeared with change in molecular weight after different freezing times. These studies reveal that alterations in the sperm membrane proteins may be responsible for damage to their membrane during freezing and thus lowering their fertilizability.

• Natural Product Sciences
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• v.18 no.3
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• pp.204-210
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• 2012

Sulfur has been used as a general physical strengthening agent from ancient times in Asia. On the basis of this point, we measured spermatogenic effect of sulfur and methyl sulfonyl methane (MSM) which is used a functional food for arthritis in rats. MSM, a kind of organic sulfur, is used as an alternative of sulfur by some people. Sulfur was administered as a dietary supplement and MSM was administered orally as a solution to 7 week old rat for 6 weeks. All sulfur administered groups showed a significant dose dependent increase in the number of sperm in the testes compared with the control group. Moreover, a histological examination showed an apparent increase in the number of seminiferous tubular cell layers in the testes of the sulfur treated rats. However, there were no observed any increase of sperm in MSM dosing group. In sulfur treated rats, the weights of body, liver, spleen, kidney, testes and epididymides didn't show significantly differences compared with the control. Histopathological examination was not revealed any morphological change in the liver, spleen and kidney. Thus, sulfur may be effectively used to treat sperm deficiency of men, but not MSM.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.31 no.3
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• pp.353-358
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• 1998

Experiments were performed to find out the physico-chemical properties of milt, and morphological changes of cryopreserved spermatozoa in tiger puffer, Takifugu rubripes. The average number of sperm and spermatocrit in milt stripped were $9.81{\pm}0.34{\times}10^{10}/m{\ell}$ and $97.8{\pm}0.8$, respectively. While total lipid concentration from seminal fluid was higher than that from sperm, total protein concentration from sperm was higher than that from seminal fluid, Na and K concentrations in sperm and those in seminal fluid were similar each other, However, glucose from sperm and seminal fluid were not detectable. Spermatozoon of tiger puffer was consisted of head, middle Piece and tail. Size of head showing horseshoe shape was $0.65{\pm}0.10{\mu}m$ in diameter and $1.35{\pm}0.30{\mu}m$ in length. The head fully containing chromatin did not have acrosome. Mitochondrion in middle piece was $0.2{\mu}m$ in average diameter and flagellum showed 9+2 structure. A few of cryopreserved spermatozoa showed morphologically loose or swollen plasma membranes.

• Animal cells and systems
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• v.9 no.2
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• pp.65-73
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• 2005

In this study, we found that sperm ball of Urechis unicinctus consisted of a somatic cell and spermatogenic cells. After separation from the sperm ball, individual spermatid floated freely in the coelomic fluid and differentiated into a mature sperm. Because of many nuclear vacuoles, spermatid nucleus was observed to be heterogeneous. Later, the spermatid nucleus condensed into the homogeneous round nucleus of the mature sperm. Perinuclear microtubules could be seen but did not seem to be organized into manchette microtubules. To understand the nature of nuclear condensation during spermiogenesis, the sperm and spermatids (spermiogenic cells) were treated with FITC-phalloidin, or anti-actin-FITC, or labeled with antiactin immunogold particles (AAIP; 10 nm) followed by transmission electron microscopy or confocal laser scanning microscopy. The anti-actin-FITC and FITC-phalloidin reactions occurred distinctly in the nuclei of both spermiogenic cells. FITC-phalloidin reacted more intensely with acrosomes. The AAIP were incorporated mainly into nuclei of both cells sometimes showing local distribution in the nucleus. Nuclear vacuoles of spermatids disappeared progressively with condensation of the nucleus, as the number of incorporated $AAIP/{\mu}m^2$ increased. These results suggest that nuclear actin microfilaments might be closely related to nuclear condensation.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.3
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• pp.335-340
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• 2006

A large number of toxicological substances and pharmacological and physical agents can cause reproductive intervention at the cellular and molecular level. The present study was designed to assess the effect of mercury ($HgCl_2$) at 50 to $550{\mu}M$ concentration ranges, in vitro, on the sperm membrane and DNA integrity, viability, and acrosomal status of normal bull spermatozoa. The samples were processed for sperm analyses using semen-diluting fluid (PBS, pH 7.2). We recorded a sharp increase in the lipid peroxidation/LPO rate; the highest was at $550{\mu}M$ mercury concentration, indicating a deleterious effect of mercury on the sperm membrane intactness. There was also a strong negative correlation between LPO rate and % viable spermatozoa (R = 0.987, p<0.001). Data obtained from a comet assay technique revealed that mercury is capable of inducing DNA breaks in the sperm nuclei. Interestingly, 92% of DNA breaks were double-stranded. The correlation between LPO rate and % DNA breaks was 0.984. Performing the gelatin test indicates that mercury is able to alter the integrity of acrosomal membranes showing an abnormal acrosome reaction. In this regard, a strong link was found between LPO rate and % halos (R = 0.990, p<0.001). Collectively, mercury proved to be a potent oxidant in the category of environmental factors affecting bull spermatozoa. Hence, considering the wide spread use of mercury and its compounds, these metals should be regarded with more concern.

• Journal of Embryo Transfer
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• v.26 no.2
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• pp.117-122
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• 2011

In this study, we examined the effectiveness of in vitro fertilization of porcine immature oocytes on the embryo development of blastocysts or hatched blastocysts and the number of cells according to the in vitro fertilization conditions. In the in vitro fertilization of in vitro matured porcine oocytes, there were no significant differences between treatment groups regarding fertilization rate, blastocyst rate, and embryo development of hatched blastocysts according to the storage periods of liquid sperm of 24, 48, and 72 hours. The embryo development rate of hatched blastocysts after the fertilization according to different spermatozoa concentrations ($0.4{\times}10^5$, $1.2{\times}10^5$, and $3.6{\times}10^5$ cells/ml) showed the highest rate in the group with a spermatozoa concentration of $1.2{\times}10^5$ cells/ml; in particular, this rate was significantly higher than that in the $0.4{\times}10^5$ cells/ml group (p<0.05). The total number of blastocysts cells as well as trophectoderms (TE) that developed in each treatment group were also significantly higher in the $1.2{\times}10^5$ cells/ml group than in any other groups (p<0.05). In contrast, the embryo development rate of blastocysts according to different co-incubation periods of sperm and oocyte (1, 3, and 6 hr) was high in the 6-hour group; in particular, the rate was significantly higher than that of the I-hour group (p<0.05). Furthermore, the total number of oocytes cells and TEs that developed was significantly higher in the 6-hour group than any other group (p<0.05). In this study, the most effective treatment conditions for porcine embryo development and high cell number were found to be as follows: a sperm storage period of less than 72 hours, a spermatozoa concentration of $1.2{\times}10^5$ cells/ml, and a 6-hour co-incubation period for sperm and ooocyte.

• 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.