• Asian-Australasian Journal of Animal Sciences
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• v.29 no.5
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• pp.646-651
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• 2016

Cryopreservation of caudal epididymal spermatozoa is an effective technique to conserve genetic potentials of superior dogs when it is not possible to collect ejaculated spermatozoa. Although hen egg yolk is commonly supplemented into the semen extender, active substances within the egg yolk which protect sperm against cryoinjury remain to be discovered. Among its compositions, low-density lipoprotein (LDL) has been reported to have a cryoprotective property for sperm cryopreservation. However, the effects of LDL on dog epididymal spermatozoa during cryopreservation have not yet been investigated. This study aimed to investigate the effects of LDL on epididymal spermatozoa quality following cryopreservation and thawing. After routine castration of 12 dogs, caudal epididymides from individuals were separated from the testes and cut into a few pieces in a Tris-buffer. Spermatozoa recovered from each sample were examined at once for sperm quality and divided into six groups of extender: no LDL, 20% egg yolk, 4%, 8%, 16%, and 24% LDL, before cryopreservation. The sperm aliquots were then equilibrated and conventionally frozen. After thawing, sperm motility, morphology, plasma membrane integrity, and acrosome integrity were evaluated. The results revealed that 4% LDL and 20% egg yolk yielded significantly higher sperm motility (57.69% and 52.69%, respectively, p<0.05) than other LDLs. In addition, 4% LDL yielded the significantly highest plasma membrane integrity (70.54%, p<0.05). In conclusion, the supplementation of 4% LDL in Tris-glucose extender could be applied for cryopreservation of canine epididymal spermatozoa.

• Korean Journal of Animal Reproduction
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• v.11 no.3
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• pp.189-195
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• 1987

This study was conducted in order to obtain the information of the histological changes in each of six segments of the epididymal ducts in Korean native goats. Thirty-two Korean native male goats were examined, dividing into seven groups, at 4 weeks intervals from 8 to 32 weeks of age. The results obtained were as follows. 1. The epididymal ducts showed histologically an abrupt growth at the age of 16 weeks being followed by almost full maturation at the age of 24 weeks. Diameter of the cauda was steadily larger than that of the caput and corpus of the epididymal ducts. 2. Spermatozoa in the lumen of epididymal ducts were first observable at the age of 16 weeks, thereafter showing sparse in the lumen of caput, whereas most dense in the lumen of cauda in the density of spermatozoa. 3. Ducts in the caput and corpus were lined by ciliated columnar epithelium until the age of 12 weeks, and later by pseudostratified ciliated columnar epithelium which was composed of ciliated columnar cells, clear cells and basal cells. Ducts of cauda epididymis were lined by simple ciliated columnar epithelia until 12 weeks of age and later by simple or pseudostratified ciliated columnar epithelium, and two types of ducts (small ducts with high epithelium and large ducts with lower epithelium) were noted. Nucleus of the epithelial cells in the caput were located in the base of cells but in the corpus and cauda, those were located in the mid part of cells. cilia were most developed in the epithelia of the corpus.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.6
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• pp.791-796
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• 2014

Cryopreservation of epididymal sperm is an effective technique to preserve genetic materials of domestic cats and wild felids when they unexpectedly die. However, this technique inevitably causes detrimental changes of cryopreserved-thawed spermatozoa, for example, by physical damage and excessive oxidative stress. L-carnitine is an antioxidant that has been used to improve sperm motility in humans and domestic animals. This study aimed to investigate the effects of L-carnitine on cat epididymal sperm quality following cryopreservation and thawing. After routine castration, cauda epididymides were collected from 60 cat testes. The epididymal spermatozoa from 3 cauda epididymides were pooled as 1 replicate. Spermatozoa samples (16 replicates) were examined for spermatozoa quality and then randomly divided into 4 groups: 0 mM L-carnitine (control), 12.5 mM, 25 mM and 50 mM L-carnitine. The sperm aliquots were then equilibrated and conventionally frozen. After thawing, sperm motility, plasma membrane integrity, DNA integrity and acrosome integrity were evaluated. The 25 mM L-carnitine significantly improved sperm motility compared with a control group (p<0.05), although this was not significantly different among other concentrations. In conclusion, supplementation of 25 mM L-carnitine in freezing extender improves cauda epididymal spermatozoa motility. The effects of L-carnitine on the levels of oxidative stress during freezing and thawing remains to be examined.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.3
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• pp.447-456
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• 1999

Objective: The purpose of this study was to evaluate outcome of intracytoplasmic sperm injection (ICSI) using epididymal and testicular sperm in patients with azoospermia. Methods: From March, 1993 to May, 1999, a retrospective clinical analysis was done of a total of 140 cycles in 112 patients who underwent ICSI. Subjects were divided into three groups: ejaculated-ICSI group included 42 cycles in 34 patients with ejaculated sperm who underwent ICSI due to severe oligospermia and past history of failed or poor fertilization in the previous in vitro fertilization and embryo tranfer (IVF-ET) cycles, microsurgical epididymal sperm aspiration and intracytoplasmic sperm injection (MESA-ICSI) group included 50 cycles in 42 patients with congenital absence of the vas deferens (CAVD) or unreconstructable obstructive azoospermia and testicular sperm extraction and intracytoplasmic sperm injection (TESE-ICSI) group included 48 cycles in 36 patients with no spermatozoa which can be retrieved from epididymis or non-obstructive azoospermia. Results: Normal two-pronuclear fertilization rates were similar in three groups: 64.4% for ejaculated-ICSI group, 59.4% for MESA-ICSI group and 60.4% for TESE-ICSI group. The pregnancy rates were 26.2%, 26.0% and 25.0% respectively. There were no significant differences in the fertilization, cleavage, and clinical pregnancy rates among ICSI cycles using ejaculated, epididymal and testicular sperm. Conclusion: Epididymal and testicular sperm obtained in azoospermic patients can fertilize oocyte successfully and may lead to be similar fertilization rates and clinical pregnancy rates to ejaculated sperm.

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.2
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• pp.130-135
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• 2022

Epididymal sperm cryopreservation provides a potential method for preserving genetic material from males of endangered species. This pilot study was conducted to develop a freezing method for tiger epididymal sperm. We evaluated post-thaw sperm condition using testes with intact epididymides obtained from a Siberian tiger (Panthera tigris altaica) after castration. The epididymis was chopped in Tyrode's albumin-lactate-pyruvate 1x and incubated at 5% CO₂, 95% air for 10 min. The Percoll separation density gradient method was used for selective recovery of motile spermatozoa after sperm collection using a cell strainer. The spermatozoa were diluted with modified Norwegian extender supplemented with 20 mM trehalose (extender 1) and subsequent extender 2 (extender 1 with 10% glycerol) and frozen using LN₂ vapor. After thawing at 37℃ for 25 s, Isolate^® solution was used for more effective recovery of live sperm. Sperm motility (computerized assisted sperm analysis, CASA), viability (SYBR-14 and Propidium Iodide) and acrosome integrity (Pisum sativum agglutinin with FITC) were evaluated. The motility of tiger epididymal spermatozoa was 40.1 ± 2.0%, and progressively motile sperm comprised 32.7 ± 2.3%. Viability was 56.3 ± 1.6% and acrosome integrity was 62.3 ± 4.4%. Cryopreservation of tiger epididymal sperm using a modified Norwegian extender and density gradient method could be effective to obtain functional spermatozoa for future assisted reproductive practices in endangered species.

• Journal of Embryo Transfer
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• v.17 no.1
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• pp.55-59
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• 2002

The objective of this study is to determine the developmental competence of in vitro matured bovine oocytes after intracytoplasmic sperm injection(ICSI) with frozen-thawed epididymal spermatozoa. The ovaries were obtained from slaughtered Korean native cows. Oocytes matured in vitro for 24 hrs were fertilized by ICSI with frozen-thawed epididymal spermatozoa. After ICSI, a group of oocytes was activated with 7% ethanol fur 5 min, and the other group was not activated. The oocytes were cultured in TCM-199 medium containing hormones and 10% FCS for 24~30 hrs in a incubator with 5% $CO_2$ in air at 38.5$^{\circ}C$. The percentage of oocytes reaching M II after 24 hrs and 30 hrs of incubation were significantly higher(p<0.05) after culture with TCM-199 media(80.0% and 88.3%) than M I(8.3% and 6.7%). The rate of cleavaged embryos to blastocyst obtained by ICSI treated activation oocytes was significantly higher(p<0.05) than that of nonactivation oocytes(22/46, 47.8% vs 10/39, 25.6%). The rates of embryos development to blastocyst obtained by ICSI treated sperm of flesh, epididymal and frozen-thawed epididymal were 24/45(53.3%), 15/40(37.5%), 11/43(25.6%), respectively and these values of fresh sperm injection were higher than frozen-thawed epididymal sperm. We also concluded that embryos can be produced with ICSI of in vitro matured oocytes by ICSI using frozen-thawed epididymal semen.

• Korean Journal of Animal Reproduction
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• v.21 no.3
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• pp.247-253
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• 1997

The objective of this study was to examine the fertilization and embryonic development rates of porcine oocytes matured in vitro according to the morphological normality of epididymal sperm in porcine IVF. The results obtained in this experiment were summarized as follows : 1. When the ratio of morphological normality of epididymal sperm was divided into the three groups with $\leq$ 10%, 10-30% and $\geq$ 50%, the fertilization and embryonic devel-opment rates of 능50% group (64%. 26%) w were significantly higher than those of other two groups ($\leq$10%: 27%, 6% and 10-30%: 36%. 5%) (p$\leq$50% group of morphologically normal epididymal sperm was adjusted to 100% (5 x 10$^5$ cells/ml), the fertilization a and development rates (63%, 27%) of epidymal sperm were similar to those of frozen-thawed ejaculated sperm (56%, 35%). 3. Also, when the fertilization and development rates of epididymal sperm were analyzed according to the oocyte : sperm ratio (1:6000, 1: 6650, 1: 7700, 1: 10000), the fertilization and development rates indicated high, in 1:6000 (68%, 32%) and 1:6650 (89%, 31%), the ratio of oocyte: sperm. Therefore, when the percentage of morphological normality of epididymal sperm is more than 50, the embryonic development a can be obtained similar to that of frozen thawed ejaculated sperm. Also, these result suggested that the morphological evaluation of normal sperm in porcine IVF using epididymal sperm sho비d be prerequisite for the more effective embryonic developm ment.

• Korean Journal of Animal Reproduction
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• v.11 no.3
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• pp.196-205
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• 1987

This study was conducted in order to obtain the informatin of the histochemical changes in each of 6 segments of the epididymal ducts in 32 Korean native male goats. The male goats were examined, dividing into 7 groups, at 4 잔 intervlas from 8 to 32 wks of age. The reuslts obtained were as follows: 1. PAS reaction showed positive on the basal and upper part beyond the nucleus of the peithelium of effernt ductules throughout all the classes of age. It was also positive on the free border and basal and upper part beyond the nucleus of the caput, on the free border andbasal parts of the corpus, and on the basal part of the cauda of the epididymal epithelium. 2. Acid phosphatase reaction was negative on the every part of epididymal epithelium at the age of 8 weeks, however, with the aging it became strangly positive on the areas between the free border and the nucleus, and moderately positive on the basal part of epithelium of the caput and corpus. In the free border adn basal part of the cauda, it was slightly positive. alkaline phosphatase reactin was negative on the every part of epididymal epithelium until 12 weeks of age. From 16 weeks, free border of epididymal epithelium becaqme slightly positive, and from 20 weeks, the reaction became stronger on the basal part but weekend on the free border with the aging. 3. In the sudan black B staining, many blue black granules between the free border and the nucleus, some granules on the basal part, and a few granules on the cytoplasm around the nucleus of the epididymal epithelium were observed from 8 weeks of age as early, and the granules were increased in number with the aging. 4. In Azan staining, reddish violet granules below the nucleus and blue granules on the upper part beyond the nucleus in some cells of epithelia of efferent ductules were noted at 12th and 16th week, and after 24th week, the granules were decreased with the aging. Golgi apparatus were clearly observable on the upper part beyond the nucleus of all parts of epididymal epithelium from 8th week, and also number of intracytoplasmic vacuoles(smaller ones on the upper part and larger ones on the basal part beyond the nucleus) and fine granules were increased with the aging. 5. In the toluidin blue staining, reddish purple granules on the basal part of the epithelium in all the parts of epididymal ducts, particularly brilliant in the cauda, were observed from 8th week as early. 6. In the Cowdry staining, numerous mitochondria, according to aging, were observed between the free border of epithelium and the upper part beyond the nucleus particularly in the catus and corpus of the epididymal ducts.

• Korean Journal of Animal Reproduction
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• v.26 no.2
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• pp.105-110
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• 2002

The objective of this study was to determine the developmental competence of in vitro matured oocytes after intracytoplasmic sperm injection(ICSI) with epididymal spermatozoa. The ovaries were obtained from slaughtered small species dogs. Oocytes matured in vitro for 24 hrs were fertilized by ICSI with epididymal spermatozoa. After ICSI, one group of oocytes was activated with 2.0 mM dimethylaminopurine or 7% ethanol for 5 min. and second group was not activated. The follicular oocytes were cultured in synthetic oviductal fluid(SOF) and TCM-199 medium containing hormones and 10% FCS for 24~48 hrs in a incubator with 5% $CO_2$ in air at 38.5$^{\circ}C$. 1. Results of IVM showed that the percentage of oocytes reaching MII after 24 h and 48 hrs of incubation were significantly higher(p<0.05) after culture with 48 hrs(9/30, 30.0%) than that after culture with 24hrs(a/30, 26.7%). 2. Results of IVM showed that the percentage of oocytes reaching MII after 48 hrs of incubation were significantly higher(p<0.05) after culture with SOF media(10/30, 30.3%) than TCM-199 media (7/30, 23.3%). 3. The rate of cleavaged embryos to blastocyst obtained by ICSI treated activation oocytes was significantly higher(p<0.05) than that of nonactivation oocytes(5/16, 25.0% vs 1/13, 5.0%). 4. The rates of development of cleavaged embryos to blastocyst obtained by ICSI treated sperm of fresh, epididymal and frozen-thawed epididymal were 8/18(44.43%), 5/16(31.3%), 2/14(14.3%), respectively. and these values of frozen-thawed epididymal sperm injection were lower than fresh sperm injection.

• The Journal of Internal Korean Medicine
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• v.33 no.4
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• pp.429-437
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• 2012

Objectives : Obesity is an important cause of diabetes, and lipotoxicity causes insulin resistance. In this study, we investigated the effects of Salvia miltiorrhiza on high fat diet-induced obese type 2 diabetic mouse models. Methods : Diabetes was induced in ICR male mouse (23~25 g) with Surwit's high fat, high sucrose diet. Mice were divided into 4 groups (n=10) of normal, control, Salvia miltiorrhiza, and metformin. After 8 weeks, body weight, OGTT, fructosamine, lipid profile, serum level of adiponectin and leptin, epididymal fat pad, liver weight and epididymal adipocyte size were measured. Results : Salvia miltiorrhiza significantly reduced oral glucose tolerance levels, fructosamine serum level, epididymal fat weight, and epididymal adipocyte size. Salvia miltiorrhiza also increased HDL-cholesterol, adiponectin and leptin serum levels. Conclusions : These results show that Salvia miltiorrhiza improves insulin resistance. Therefore we suggest that Salvia miltiorrhiza would be an effective treatment for obese type 2 diabetic patients.