• Journal of The Korean Society of Grassland and Forage Science
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• v.38 no.4
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• pp.320-324
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• 2018

In this study, we examined effect of sperm penetration of oocytes after in vitro fertilization (IVF) with cauda epididymal spermatozoa in Hanwoo bull after feeding of timothy hay. One testicle with epididymides was castrated from one Hanwoo bull (14 months of age) and spermatozoa recovered from cauda epididymis and cryopreserved. As control, frozen Hanwoo semen was used. Matured cumulus oocyte complexes were co-incubated with frozen-thawed cauda epididymal spermatozoa for 12 or 18 hours. After IVF, presumptive zygotes were cultured in modified synthetic oviductal fluid. In experiment 1, we examined sperm penetration rate at 12 hours of IVF with epididymal sperm. Total penetration rate among cauda epididymis and control was similar(mean${\pm}$standard error, cauda epididymis and control vs. $49.7{\pm}11.3$ and $54.4{\pm}12.8%$). In experiment 2, cleavage and blastocyst developmental rate were evaluated at day 2 and day 8 after IVF for 18 hours. Cleavage rate among cauda epididymis and control was similar(cauda epididymis and control vs. $81.2{\pm}3.4$ and $82.7{\pm}2.5%$). However, blastocyst developmental rate of cauda epididymis group was significantly higher than that of control group(cauda epididymis and control vs. $24.4{\pm}1.6$ and $12.2{\pm}2.8%$, p<0.05). In conclusion, cauda epididymal spermatozoa in Hanwoo bull has high embryo developmental competence and can be used as an alternative to ejaculated frozen sperm in vitro.

• Development and Reproduction
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• v.3 no.1
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• pp.87-93
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• 1999

The possible role of Na$^{+}$/H$^{+}$ antiporter in both the capacitation and the acrosome reaction (AR) was examined in mouse epididymal spermatozoa. Spontaneous acrosome reaction was inhibited by dimethyl amiloride (DMA), a specific inhibitor of Na$^{+}$/H$^{+}$ antiporter, with dose dependent manner. Follicular fluid- or A23l 87-induced acrosome reaction was not inhibited by DMA. It suggests that change in pH$_{i}$ by monovalent cation transport through the Na$^{+}$/H$^{+}$ antiporter is possibly engaged in the capacitation and that agonist- as well as A23l87-induced AR in capacitated sperm might be independent from the Na$^{+}$/H$^{+}$ antiporter. Conclusively, changes in pH$_{i}$ through the Na$^{+}$/H$^{+}$ antiporter might be important for sperm capacitation and it virtually occurs upstream of the $Ca^{2+}$ influx which precedes the acrosome reaction in mouse epididymal spermatozoa.pididymal spermatozoa.

• Applied Microscopy
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• v.24 no.2
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• pp.12-25
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• 1994

This research was undertaken to determine the effects of the anticancer and immunosuppressive drug cyclophosphamide (CP) on the epididymis of the male rat in terms of ultrastructural alteration and protein analysis by SDS-PAGE at different groups; control group, 1 week group, 3 weeks group, 5 weeks group were treated with saline (control group) or CP at doses of 20mg/Kg/week, 1 time a week, respectively. In the cytoplasm of the principal cells on the epididymis, the mitochondrial outer and inner membranes were significantly swollen or disrupted. The cisterns of rough endoplasmic reticulum (rER) were also swollen, and a number of Golgi vesicles were increased, respectively. It is suggested that treatment with CP alters the specific cell organelles in all segments of the epididymis. CP caused changes in protein concentrations in cauda of epididymis after CP treatment. Total proteins of 30 to 39 species such as lactate dehydrogenase, carnitine acetyltransferase and acid phosphatase were expressed in the cauda fluid. Then the more CP was increased, the more concentration of proteins caused to decrease, synthesize or increase in epididymal cauda. In contrast to the control group, in particular 29KD and the other 10 proteins in the cauda fluid were decreased or disappeared, respectively, whereas 89KD and the other 6 proteins in the cauda, were increased or synthesized, respectively. The other proteins are not showed distinctive difference. Therefore, it is possible that CP at a high dose accumulation alters epididymal function with dose-related increase or decrease in specific activity of marked proteins for all regions of the epididymis (particularly, specific segment of cauda). These alterations could be mediated by direct, toxic effects of the drug on the epithelium or be secondary to changes in the spermatozoa as a result of the CP treatment.

• The Korean Journal of Zoology
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• v.34 no.2
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• pp.159-172
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• 1991

It has been investigated what could be the selective marker distinguishing the immature from mature spermatozoa and whether fi -glucuronidase and fi -glucosidase are dependent on androgen in the luminal fluid of the epididymis or not. The contents of hexose, hexosamine and sialic acid in the epithelial cells, luminal fluid and spermatozoa of the epididymis were examined and the patterns of protein bands were compared in each group of the luminal fluid by SDS-PAGE. Lactate dehydrogenase, glucose-6-phosphatase, Na+ -K+ -ATPase and MgNa-ATPase showed higher activities in the cauda than the caput epididymal spermatozoa but only $Mg^2$+-ATPase activity appeared to be changed significantly. When the contents of hexose, hexosamine and sialic acid were analyzed and compared quantitatively, those of hexose were significantly different in the luminal fluid of caput and cauda epididymis, those of hexosamine in the epithelial cells and those of sialic acid in the epithelial cells and luminal fluid. When SDS-PAGE has been performed in each group, the band of MW 33-37 KD which was absent in the luminal fluid of caput epididymis appeared obviously in the luminal fluid of cauda epididymis and ako apeared in the cauda sperm crude membrane fraction. In addition, $\beta$ -glucuronidase and $\beta$ -glucosidase activities and their dependence on androgen were measured and the SDS-PAGE patiems of proteins and/or glycoproteins in the luminal fluid were examined. The activities of these two enzymes in the luminal fluid of the epididymis decreased significantly from the 5th day after castration. When testosterone was injected, the activity of $\beta$ -glucuronidase began to increase significantly from the 5th day following injection and that of $\beta$ -glucosidase from the loth day. On the other hand, the band of about MW 21 KD was newly observed in the lumen of caput epididymis when testosterone was administered.

• Korean Journal of Veterinary Research
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• v.14 no.2
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• pp.159-171
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• 1974

Histological and histochemical studies were made on the lining epithelia of the various ducts in epidymis of the Rooster and absorptive function of the canal epithelial cells in the Rooster epididymis were also investigated after administration of India ink. The results obtained were summerized as follows; 1. Epihtelium lining the rate testis was mainaly composed of single later of cuboidal cells, and was partially composed of flattened squamous or low columnar cells. Efferential ductules were characterized by having many villous projections orrfolds which extened into the lumen, and were lined by stereociliated pseudostratified epithelium which consisted of manily ciliated columnar cells, a few scattered clear cells and basal cells. Connecting ductules were lined by ciliated pseudostriatified colummnar epithelium in which ciliated columnar cells, clear cells and basal cells were noted. Epididymal ducts were lined by pseudostratified epidhelium in which columnar and basal cells were noted. 2. PAS-granules, saliva resistant were noted mainly in the epithelial cells of efferential and connecting ductules. 3. Sudan black B stained heavily the granules in the epithelial cells of sufferential and connecting ductules. 4. The granules reactive to acid phosphatase most abundant in the epithelial cells of efferential ductules and were lesser amount in the epithelial cells of connecting ductules where as very few or no granules were seen in the rest of the ducts. 5. Alkaline phosphatase activity was most prominent but discontinuous in the luminal surface of the epithelium of efferential ductules and less marked in the connecting ductless. No enzyme activity was noted in the canal epithelium of epididyml duct. 6. India ink granules were most numerous in the epithlial cells of efferential ductules and were a few in connecting ductules. Very few or no granules of India ink were noted in the other types of the ducts. India ink granules in the epithelium increased gradually as the time after the administration of India ink (one up to twenty-nine hours) has proceeded. From those results it is suggested that epithelial cells of efferential and connecting ductules have active absorptive function, whereas the rest of duct system in the epididymis of the Rooster may be the mere pathway of the seminal fluid without significant modification of its constituents.

• Development and Reproduction
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• v.18 no.4
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• pp.187-195
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• 2014

Polycyclic aromatic hydrocarbons (PAHs), which are ubiquitous in the air, are present as volatile and particulate pollutants that result from incomplete combustion. Most PAHs have toxic, mutagenic, and/or carcinogenic properties. Among PAHs, benzo[a]pyrene (B[a]P) and dimethylbenz[a]anthracene (DMBA) are suspected endocrine disruptors. The testis is an important target for PAHs, yet effects on steroidogenesis in Leydig cells are yet to be ascertained. Particularly, disruption of testosterone production by these chemicals can result in serious defects in male reproduction. Exposure to B[a]P reduced serum and intratesticular fluid testosterone levels in rats. Of note, the testosterone level reductions were accompanied by decreased steroidogenic acute regulatory protein (StAR) and $3{\beta}$-hydroxysteroid dehydrogenase isomerase ($3{\beta}$-HSD) expression in Leydig cells. B[a]P exposure can decrease epididymal sperm quality, possibly by disturbing the testosterone level. StAR may be a key steroidogenic protein that is targeted by B[a]P or other PAHs.

• Applied Microscopy
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• v.22 no.1
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• pp.89-102
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• 1992

This research was undertaken to determine the effects of the anticancer and immunosuppressive drug cyclophosphamide (CP) on the epididymal caput of the male rat in terms of ultrastructural alteration and protein analysis by SDS-PAGE at different groups; control group, 1 week group, 3 weeks group, 5 weeks group were treated with saline (control group) or CP at doses of 20 mg/kg/week, 1 time a week, respectively. In the cytoplasm of the principal cells on the epididymis, the mitochondrial outer and inner membranes were significantly swollen or disrupted. The lumens of rough endoplasmic reticulum (rER) were also swollen, and the number of Golgi vesicles were increased, respectively. It is suggested that treatment with CP alters the specific cell organelles in the epididymis. CP caused changes in protein concentrations in caput of epididymis after CP treatment. Total proteins of 32 to 37 species such as lactate dehydrogenase, carnitine acetyltransferase and succinate dehydrogenase were expressed in the caput fluid. Then the more CP was increased, the more concentration of proteins caused to decrease, synthesize or increase in epididymis. In contrast to the control group, in particular carnitine acetyltransferase and the other 9 proteins in the caput fluid were decreased or disappeared, respectively, whereas lactate dehydrogenase and the other 5 proteins in the caput fluid were increased or synthesized, respectively. The other proteins are not showed distinctive difference. These alterations could be direct mediated by toxic effects of the drug on the epithelium or be secondary to changes in the spermatozoa as a result of the CP treatment.

• Korean Journal of Animal Reproduction
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• v.14 no.1
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• pp.23-30
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• 1990

These studies were carried out to investigate the effects of fetal calf serum(FCS), estrous porcine serum(EPS), porcine follicular fluid(PFF), hormone and matured cumulus cell(MCC) on in vitro maturation and fertilization of porcine follicular oocytes. The ovaries and testes were obtianed from slaughtered Landrace sow and boars, respectively. The follicular oocytes surrounded with cumulus cells were recovered by aspirating follicular fluid from the visible follicles of diameter 3~5 mm and the semen were prepared from boar's epididymal cauda. The follicular oocytes were cultured in TCM-199 medium containing hormones, FCS, EPS, PFF and MCC for 48hrs. in a incubator with 5% CO2 in air at 36$^{\circ}C$ and then matured oocytes were again cultured for 18~20 hrs. with $1.5\times$106/ml motile capacitated sperm in the modified Tyroide solution containing 100$\mu\textrm{g}$/ml of heparin. The results obtained in these experiments were summarized as follows : 1. The maturation and fertilization rate of the follicular oocytes, cultured in the TCM-199 medium supplemented with 10% FCS and PMSG＋HCG were 55.6~64.5% and 33.3~37.1%, respectively. 2. The maturation and fertilization rate of the follicular oocytes cultured in the TCM-199 medium supplemented with 20% EPS and PMSG＋HCG were 50.0~55.0% and 30.3~33.3%, respectively. 3. The maturation rate(59.0~64.2%) and fertilization rate(34.8~39.3%) of follicular oocytes cultured in TCM-199 medium supplemented 20% FCS and 50% PFF were higher than those of follicular oocytes cultured in TCM-199 medium supplemented with 5%, 10% and 15% FCS and 10% and 50% PFF. 4. The maturation rate(60.0%) and fertilization rate(40.0%) of follicular oocytes cultured in TCM-199 medium supplemented with 20% FCS and granulosa cell (1$\times$106/ml) were significantly higher than those of fiollicular oocytes cultured in TCM-199 medium supplemented with 5%, 10% and 15% FCS and granulosa cell.

• Korean Journal of Animal Reproduction
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• v.14 no.1
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• pp.84-91
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• 1990

Experiments were disigned to define and optimize efficiency of a system whereby pig follicular oocytes could be matured and fertil ized in vitro. The pig oocytes removed from 1- 2 mm and 3-7 mm follicles were cultured in vitro in the mKRB(-BSA) solution containing estrous sow serum (ESS), FCS or dialyzed pig follicular fluid for 24 to 48 hr at 37$^{\circ}C$. The oocytes matured in vitro were evaluated after epididymal spermatozoa-oocyte incubation for 24 hr for pronucleus formation. 50-60% of the oocytes reached metaphase II during 36 to 48 hr of culture. There was no differernce in oocyte matura¬tion between two groups of follicular size but meiosis was slightly faster in the 3-7 mm follicular oocytes. The oocytes matured in mKRB (-BSA) plus 5% ESS, 15% FCS or dialyzed follicular fraction showed slightly higher maturation rates than the control mKRB. in vitro fertilization, pronucleus formation, tended to be increased when mKRBi-BSA) plus 5% ESS or 15% FCS was used for oocyte maturation and in vivo -capacitated spermatozoa were inseminated, respectively. It is concluded that ESS, FCS and dialyzed pig follicular fluid may be effective factors for in vitro maturation and fertilization of pig follicular oocytes.

• Korean Journal of Animal Reproduction
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• v.13 no.3
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• pp.171-178
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• 1989

These experiments were carried out to investigate the effect of a co-culture with granulosa cells on in vitro maturation, fertilization and development of bovine follicular oocytes. 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 diameter 2-6mm. Bovine oocytes were matured in vitro for 24-26 hr and then fertilized in vitro using epididymal spermatozoa capacitated by preincubation for 2-3hr in BO solution containing BSA(5mg/ml) and caffein(25mM). Eight hours after insemination, the oocytes were cultured in a co-culture system with granulosa cells. The rates of maturation of the follicular oocytes cultured in a co-culture system with granulosa cells were 83.1%, the rate of fertilization of the follicular oocytes culture in a co-culture in a co-culture system with granulosa cells were 76.9%, respectively. No significant difference are observed between control and treatment in maturation and fertilization rates. The rates of embryos developed to 2-, 4-, 8-, 16-cell and monula stages after co-cultured with granulosa cells were 65.8, 57.9, 39.5, 34.2 and 34.2%, respectively. The value for 16-and morula stages were significantly higher (P<0.05) than those of the embryos cultured in the basic medium.