• Reproductive and Developmental Biology
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• 제36권3호
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• pp.225-230
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• 2012

Interferon induced transmembrane protein-1 (Ifitm-1) has been reported to have an important role in primordial germ cell formation, and it has expressed in female reproductive organ. In the present study, Ifitm-1 gene expression was identified in testes and all part of epididymis using western immunoblot and immunohistochemistry. Interestingly, Ifitm-1 expression was observed on the head of spermatozoa. To investigate the role of Ifitm-1 gene expression in behavior of spermatozoa after acrosome reaction, fresh sperm was incubated with calcium ionophore to induce acrosome reaction, whereas the expression of Ifitm-1 was not altered after the acrosome reaction. Then to identify the effect of Ifitm-1 in sperm motility and other seminal parameters, different concentration of Ifitm-1 antibody was incubated with spermatozoa, and seminal parameters were assessed using computer-assisted semen analysis (CASA). Interestingly, motility, progressive, and VAP were increased in the sperm with Ifitm-1 antibody treated compared to rabbit serum, however other parameters such as straightness were not changed. In order to identify the functional significance of Ifitm-1 in fertilization, capacitated spermatozoa were pre-incubated with anti-Ifitm-1 antibody and subsequently examined the ability to adhere to mouse oocytes. However, any defection or alteration in sperm-egg fusion was not found, Ifitm-1 antibody treated or non-treated spermatozoa showed a normal penetration. Although the precise role of Ifitm-1 in sperm motility and following fertilization need to be elucidated, this study suggests that the activation of Ifitm-1 on the sperm may enhance the motility of spermatozoa in mice.

• 한국환경보건학회:학술대회논문집
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• 한국환경보건학회 2005년도 국제학술대회
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• pp.319-322
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• 2005

Porous building materials are not only sources of indoor air pollutants such as volatile organic compounds (VOCs) but they are also strong sinks of these pollutants. Volatile organic compounds have been implicated in impaired spermatogenesis, increase in the incidence of malformed sperm and decrease in the percentage of moving sperm. The aim of this study was to determine and compare the direct effects of various volatile organic compounds (phenol, formaldehyde; HCHO, ethanol, toluene, styrene) on motility and survival rate of human sperm in vitro. Semen samples from 3 health subjects were prepared using swim-up method and 1-10mM volatile organic compounds were added to the test medium. HCHO and phenol produced significant decreases in the motility and survival rate with a different potency. The most potent inhibition of motility and survival rate was observed after exposure to HCHO. Less than 1mM HCHO significantly inhibited sperm motility. When ethanol is added directly to sperm, at concentrations equivalent to that in serum after heavy drinking, these damaging effects were lowest compared with other volatile organic compounds. Present study shows that each compound has differential toxic potency to human sperm and we need special caution for the use of HCHO and phenol.

• Reproductive and Developmental Biology
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• 제32권2호
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• pp.123-126
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• 2008

The objective of this study was to determine the potential hazardous effects of sorting process by flowcytometry on the quality of boar spermatozoa by flowcytometer. Freshly collected boar semen was diluted and divided into two groups; control none sorted and sorted. Sperms in sorted group were processed with flowcytometer for cell sorting with $100\;{\mu}M$ nozzle under the 20 psi pressure. Measurements on each parameter were made at two time points, 0hr (right after sorting) and 24 hr post sorting. Although there was a tendency of lower viability in sorted group than none sorted control group, the percentage of live cells in control ($75.83{\pm}6.92\;&\;59.53{\pm}10.34$) was not significantly different from sorted ($59.70{\pm}7.37\;&\;43.97{\pm}3.76$) at both 0 and 24 hr post sorting. However, sorted sperm showed significantly lower mitochondrial function compared to the control at both 0 h ($79.37{\pm}3.22\;vs.\;63.50{\pm}10.05$) and 24 hr ($67.27{\pm}3.22$ vs. $46.97{\pm}5.37$) time points (p<0.007). Sperm DNA fragmentation rate was significantly lower in control ($22.0{\pm}7.04$) than that of sorted ($32.27{\pm}7.49$) at 24 hr time point (p<0.0002). Taken together, these data suggested thatsorting process by flowcytometer may have influenced sperm motility rather than viability. Also high speed sperm sorting by flowcytometer has significant effects on DNA fragmentation on elapsed time after sorting.

• 대한본초학회지
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• 제30권3호
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• pp.19-24
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• 2015

Objectives : HPL-1, a novel herbal medicine which is composed of five herbs such as Kalopanacis Cortex, Chaenomelis Fructus, Raphani Semen, Atractylodis Rhizoma and Pulvis Aconiti Tuberis Purificatum, was developed for treatment of osteoarthritis. This study is aimed to develop analytical method for consistent quality control of HPL-1 and validate chromatographic method. Methods : Chromatographic analysis was performed using ultra-high performance liquid chromatography - diode array detector (UHPLC-DAD) equipped with RP-amide column, column oven, and auto sampler. Marker compounds [protocatechuic acid, chlorogenic acid, liriodendrin, 3,5-dicaffeoylquinic acid, ${\beta}$-D-(3-O-sinapoyl)-fructofuranosyl-$\alpha$-D-(6-O-sinapoyl)glucopyranoside and benzoylmesaconine] were separated by step gradient elution of acetonitrile and 0.1% phosphoric acid/water. The method validation was evaluated by quantitative validation parameters of linearity, accuracy, precision, limit of detection (LOD) and limit of quantification (LOQ) according to KFDA guideline.Results : An optimized method for six marker compounds in HPL-1 was established by UHPLC-DAD. The correlation coefficient (R²) with each calibration curve was greater than 0.99. The LOD and LOQ were within the range of 0.008-0.090 and $0.023-0.274{\mu}g/mL$, respectively. The relative standard deviation (RSD) of intra- and inter-day variability were less than 4.0%. The result of recovery test was range from 93.3-106.3% with RSD < 4.0%.Conclusions : These results suggest that the quantitative UHPLC method is precise, accurate, effective for quality evaluation of HPL-1. The method may also contribute to improve quality of crude drug preparations used for treatment of various diseases.

• 한국수정란이식학회지
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• 제13권1호
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• pp.77-86
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• 1998

A combined technology of transvaginal ovum pick-up(OPU) system with in vitro-oocyte manipulation technique can be used for improving reproductive efficiency in the cattle. The objective of this study was to establish a newly-conceived breeding program using OPU in the pregnant cows. The OPU trial was performed in pregnant cows every 10 days from 40 through 90 days of artificial insemination (Al), and number of follicles in ovary, number of retrieved oocytes and embryo development following in vitro-fertilization, were evaluated. Reduced number of follicles in the ovaries of pregnant cows was firstly detected from 70 days after A' and a significant (P<0.05) decrease in the follicle number (5.4 follicles /donor) was found at 90 days than at 40, 50, 60 and 80 days after Al (8.0~9.2). A similar pattern was also observed in the number of oocytes retrieved by OPU apparatus during experimental period. When retrieved oocytes were matured and inseminated in vitro with frozen bull semen, development of the oocytes to the blastocyst stage was not significantly affected by the retrieval time. Four embryos (morula or blastocyst stage) derived from oocytes retrieved from pregnant cows were nonsurgically transferred to four recipient cows on day 7 of estrus cycle. For the first time in Korea, three of four transferred embryos developed to live calves with normal physiological parameters. In conclusion, an effective breeding program employing pregnant cow can be developed by use of OPU trial and in vitro culture techniques of oocytes ; OPU system could be repeated in pregnant cows with no risk of abortion and viable offsprings were borne after transfer to the recipients.

• Clinical and Experimental Reproductive Medicine
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• 제21권1호
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• pp.31-41
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• 1994

Morphological estimation of human spermatozoa is complicated by the fact that there is great natural variation in shape. This natural variation in shapes makes it difficult to say which forms are associated with infertility and which are normal variations. Possibly post coital test or in vitro cervical mucus penetration tests will help to clarify this question by showing which sperm are capable of penetration. The purpose of this investigation was performed to assess distribution of various morphological abnormalities according to the ability of sperm to penetrate cervical mucus. The sperm-mucus penetration using hen's egg white as substituting mucus for human cervical mucus was done in 45 fertile men with normal semen analysis and 122 infertile men with abnormal seminal parameters more than one. The female partners of 122 infertile couples showed normal results in the female fundamental test for fertility. Conventional semen analysis was evaluated according to the WHO standard normal(l980). The detailed classification of the abnormal sperm was made according to David et al(l975). The vitality of the sperm samples determined by eosin yellow-nigrosin stainig according to the method of Eliasson(l977). Results were as follw; 1. The patients had significantly lower total sperm count, motility (%), normal morphology (%), viability and total functional sperm fractions(TFSF) than fertile donors. 2. The mean value of sperm penetration distance of the patients(28.69${\pm}$11.02mm) showed significantly lower than fertile donors(37.33${\pm}$5.49mm). And 43/45 fertile donors(95.5%) as well as 57/122 patients(46.7%) had over 30mm in sperm penetration distance respectively. While 2/45 fertile donors(4.5 %) and 65/122 patient(53.3%) had under 30mm in sperm penetration distance respectively. 3. The morphological abnormalities in fertile donors were significantly lower 23.04${\pm}$5.83% (head = 12.89${\pm}$4.98, neck=6.11${\pm}$3.83%, and tail=3.43${\pm}$2.65%), compared to 36.03${\pm}$14. 40% in patients(head = 15.98 8.60%, neck 11.20${\pm}$6.56% and tail=8.70${\pm}$6.55%). Also, 3 types of sperm abnormalities including head, neck and tail were significantly lower in patient than fertile donors, respectively. Both the patients and fertile donors showed higher distribution of sperm with abnormal head than abnormal neck and tail. 4. The mean morphological abnormalities(SP>30mm) of the patients(30.68 11.64%; head = 15.95${\pm}$9.35%, neck=8.14${\pm}$4.21 %, tail=6.56${\pm}$5.64%) were significantly lower compared to patients(40.72${\pm}$15.01 %; head=16.02${\pm}$7.69%, neck 13.89${\pm}$7.82%, tail=1O.58${\pm}$6.75%) under 30mm in sperm penetration distance. Also, both groups over 30mm and under 30mm in sperm penetration showed distance higher distribution of sperm with abnormal head than abnormal neck and tail. The morphological abnormalities of head did not show significant difference but abnormal neck and tail were significant difference between the over 30mm and under 30mm group in sperm penetration distance.

• Clinical and Experimental Reproductive Medicine
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• 제46권4호
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• pp.173-177
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• 2019

Objective: We investigated the clinical characteristics of men with testosterone replacement therapy (TRT)-induced hypogonadism and its effect on assisted reproductive technology (ART) in infertile couples. Methods: This study examined the records of 20 consecutive male patients diagnosed with azoospermia or severe oligozoospermia (< 5 × 10⁶/mL) who visited a single infertility center from January 2008 to July 2018. All patients were treated at a primary clinic for erectile dysfunction or androgen deficiency symptoms combined with low serum testosterone. All men received a phosphodiesterase 5 inhibitor and TRT with testosterone undecanoate (Nebido^®) or testosterone enanthate (Jenasteron^®). Patients older than 50 years or with a chronic medical disease such as diabetes were excluded. Results: The mean age of patients was 37 years and the mean duration of infertility was 16.3 ± 11.6 months. At the initial presentation, eight patients had azoospermia, nine had cryptozoospermia, and three had severe oligozoospermia. Serum follicle-stimulating hormone levels were below 1.0 mIU/mL in most patients. Three ongoing ART programs with female factor infertility were cancelled due to male spermatogenic dysfunction; two of these men had normal semen parameters in the previous cycle. After withholding TRT, serum hormone levels and sperm concentrations returned to normal range after a median duration of 8 months. Conclusion: TRT with high-dose testosterone can cause spermatogenic dysfunction due to suppression of the hypothalamic-pituitary-testicular axis, with adverse effects on infertility treatment programs. TRT is therefore contraindicated for infertile couples attempting to conceive, and the patient's desire for fertility must be considered before initiation of TRT in a hypogonadal man.

• Clinical and Experimental Reproductive Medicine
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• 제18권1호
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• pp.63-71
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• 1991

To establish the normal fertile range in the results of the sperm zona-free hamster ova penetration assay (SPA) in Korean male, SPA using the low temperature ($4^{\circ}C$) capacitation in TEST-yolk buffer (TYB) was performed in 67 fertile and 26 infertile men. Sperm parameters in routine semen analysis were also checked and compared with the results of SPA. Sperm concentration, motility and motility index (MI) were significantly higher in fertile group compared with infertile group: $96.0{\pm}46.6$ vs $43.6{\pm}31.9{\times}10^6/ml$, $65.5{\pm}14.8%$ vs $45.8{\pm}23.6%$ and $46.31{\pm}13.29$ vs 27.40{\pm}17.98$, respectively. In fertile group, the hamster ova penetration rate (PR) was $98.5{\pm}5.0%$ (80%-100%), and the penetration index (mean penetrations per ovum, PI) was $9.59{\pm}6.35$(3.1-29.0). All the fertile men showed PI>3.0. In infertile group, PR was $24.6{\pm}24.8%$ (0%-70%), and PI was $0.40{\pm}0.42$ (0-1.3). Both PR and PI were significantly lower in infertile group. There was a significant correlation beween PI and sperm motility or MI, respectively, in fertile group whereas there was no correlation in infertile group. These data suggest that SPA using the low temperature capacitation in TYB can be a valuable diagnostic tool for the assessment of male fertility in vitro and provide an important supplement to the traditional tests of sperm quality.

• 한국수정란이식학회지
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• 제28권3호
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• pp.297-302
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• 2013

Cryopreservation of epididymal spermatozoa offers a potential tool for rescuing genetic material from males of genetically elite populations. Castration, catastrophic injury, sudden death or any other event that makes semen collection or mating impossible may prematurely terminate a stallion reproduction. Stallion epididymal spermatozoa vary widely in the loss of progressive motility, acrosomal integrity, and viability during freezing and thawing. The objective of this work was to investigate the effect of (1) freezing package types on cryopreservation efficiency, (2) thawing temperatures (37, 56 or $70^{\circ}C$) on Computer Assisted Sperm Analysis (CASA) parameters and (3) post-thawing incubation time (0, 1, 2 or 4h) on castrated stallion epididymis. Post-thawed sperm motility ranged between 59.69% and 64.28% ($56^{\circ}C$ and $37^{\circ}C$) in various thawing temperatures. When stallion epididymis sperm was frozen, straw was better than freezing tube on VCL (Velocity of Curvilinear Line) and VAP (Velocity of Average Path) parameter. Higher percentage of motility was observed at $37^{\circ}C$ thawing temperature even though no significant difference was observed among various temperatures. The motility, VCL, ALH (Amplitude of Lateral Head displacement), VAP, BCF (Beat-Cross Frequency) and STR (Straightness index) parameter of post-thawed sperm were significantly decreased by increasing the incubation time for all thawing temperatures. The present study showed that type of freezing package (Straw vs. Freezing tube) was not significantly different on cryopreservation efficiency. Furthermore, stallion epididymal spermatozoa frozen-thawed at $37^{\circ}C$ for 1 min resulted the highest proportion of motility and velocity movement. In addition, motility and viability of frozen-thawed stallion epididymal spermatozoa were also decreased by incubation.

• 대한약침학회지
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• 제17권3호
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• pp.31-39
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• 2014

Objectives: This study was performed to analyze the toxicity and to find the lethal dose of the test substance Water-soluble Carthami-flos pharmacopuncture (WCF) when used as a single intravenous-dose in 6-week-old, male and female Sprague-Dawley rats. Methods: The experiment was conducted at Biotoxtech according to Good Laboratory Practices. 20 female and 20 male Spague-Dawley rats were divided into 4 groups of 5 female and 5 male animals per group. The rats in the three experimental groups received single intravenous injections with 0.125-mL, 0.25-mL and 0.5-mL/animal doses of WCF, Groups 2, 3, and 4, respectively, and the control group, Group 1, received a single intravenous injection with a 0.5-mL dose of normal saline. Clinical signs were observed and body weight measurements were carried out for 14 days following the injections. At the end of the observation period, hematology, clinical chemistry, histopathological tests and necropsy were performed on the injected parts. Results: No deaths occurred in any of the groups. Also, no significant changes in body weight, hematological parameters or clinical chemistry test results between the control group and the experimental groups were observed. Visual inspection after necropsy showed no abnormalities. Histopathological tests on the injected parts showed no significant differences, except for Group 1 females; however, the result was spontaneous generation and had no toxicological meaning because it was not dose-dependent. Therefore, this study showed that WCF had no effect on the injected parts in terms of clinical signs, body weight, hematology, clinical chemistry, and necropsy. Conclusion: As a result of single intravenous-dose tests of the test substance WCF in 4 groups of rats, the lethal dose for both males and females exceeded 0.5 mL/animal. Therefore, WCF is a relatively safe pharmacopuncture that can be used for treatment, but further studies should be performed.