• Clinical and Experimental Reproductive Medicine
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• v.23 no.1
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• pp.87-94
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• 1996

For evaluating the suitability of human follicular fluid(HFF) as protein supplement in ART, this preliminary study was performed to examine the maturation promoting activity of HFF on mouse follicular oocytes in vitro. Mouse follicular oocytes were collected from ovaries of 21-28 day old ICR mice by puncturing the antral follicles with fine needle at 48 hours after PMSG injection. The oocytes were rinsed and cultured in modified Whittingham's $T_6$ medium containing purines or dbcAMP to maintain meiotic arrest, and different concentrations of HFF were added into the culture medium to examine the effect of HFF on releasing the oocytes from the suppressive influence of the meiotic inhibitors. As a control for HFF, the maturation promoting activity of human fetal cord serum(HFCS) was investigated and compared with the activity of HFF. While HFF was separated, by molecular weight(M.W), into high M.W. fraction(M.W>30,000) and low M.W. fraction(M.W<30,000) and the effects of the fractions on meiotic resumption were investigated in the presence of the meiotic inhibitors. Also hormone analysis was performed to compare the content of hormones in HFF with that in HFCS. Same concentrations of HFF and HFCS induced similar germinal vesicle break down(GVBD) rates of the oocytes meiotic arrested by purines(4mM hypoxanthine+0.75mM adenosine), but the extrusion rate of 1st polar body(PB) of the oocytes cultured in HFF(65.0%, P<0.05) was significantly higher than that(51.6%) in HFCS. While, in the presence of 200 M dbcAMP, the maturation promoting activity of HFF (GVBD: 70.5%, $p<10^{-6}$; 1st PB extrusion: 67.1%, $p<10^{-3}$) was significantly greater than that of HFCS(GVBD: 35.2%; 1st PB extrusion: 41.1%). The oocytes cultured in the fraction of HFF containing high M.W. components showed higher meiotic maturation rates than the oocytes cultured in the low M.W. fraction of HFF. Gonadotropins and $E_2$ were known to improve the completion of maturation changes, and the levels of these hormones were higher in HFF than in HFCS. Therefore, HFF was more effective than HFCS to use for promoting meiotic resumption of mouse oocytes in vitro.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.3
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• pp.225-233
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• 2001

Objective : To evaluate the efficacy of low-dose aspirin on IVF outcome and endometrium in patients undergoing IVF-ET. Materials and Methods : From February, 2001 to Jun, 2001, 60 infertile patients were randomly divided into study group (28 cycles) and control group (32 cycles). The study group received a daily oral dose of 25 mg of aspirin for at least 2 weeks from first visiting day. Controlled ovarian hyperstimulation was initiated in all patients with the GnRH agonist starting in the midluteal phase of the previous cycle. Results: There were no significant differences in age of the patients, basal serum E2, LH, FSH level and endometrial thickness among two groups. There were no statistically significant differences between the study group and the control group respectively in dosage ($26.5{\pm}4.8$ vs $26.2{\pm}5.3$ amples) and duration ($10.4{\pm}4.2$ vs $9.8{\pm}5.3$ days) of gonadotropin administration, serum E2 level on the hCG administration day ($1823{\pm}342$ vs $1854{\pm}543$), LH ($14.5{\pm}2.7$ vs $14.8{\pm}3.1$), FSH ($16.7{\pm}3.4$ vs $18.3{\pm}4.7$), the number of follicles > 15 mm ($13.2{\pm}6.3$ vs $12.8{\pm}5.9$), the number of oocytes retrieved ($9.2{\pm}2.4$ vs $8.4{\pm}1.7$), the number of embryos transferred ($4.7{\pm}2.0$ vs $4.7{\pm}2.0$), fertilization rate (68.4% vs 64.5%), implantation rate (21.3% vs 17.6%), and clinical pregnancy rate (28.4% vs 26.2%). The endometrial thickness and the percentage of endometrial trilaminar pattern on hCG day were significantly higher in study group than control group ($12.9{\pm}3.7mm$ vs $10.4{\pm}2.8mm$, 78.3% vs 64.5%). Conclusion: Many reports suggest that low-dose aspirin improve ovarian response, implantation rate, fertilization rate, implantation rate, and pregnancy rate by increasing the blood flow, but we couldn't prove the significant effect of low-dose aspirin on the IVF outcome except on endometrium. This may be affected by dose of aspirin, duration, and number of patients studied. This trial is small, so our results highlight the need for a large randomized controlled trial to identify the effect of low-dose as pirin on IVF-ET outcome.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.4
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• pp.225-234
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• 2004

Objectives: To evaluate the efficacy of GnRH antagonist cetrorelix in women undergoing controlled ovarian hyperstimulation (COH) for in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) and to determine changes in serum hormone concentrations during cetrorelix administration. Methods: We performed a clinical trial on 30 patients undergoing COH with highly purified follicular stimulating hormone (HP-FSH) and gonadotropin releasing hormone antagonist (GnRHant), cetrorelix. FSH was administrated from day 2 or 3 of cycle with fixed dose and adjusted according to individual response. 0.25 mg of cetrorelix was injected daily subcutaneously from stimulation day 5 until the day of hCG administration. Daily ultrasound monitoring was performed for growing follicles and serum levels of luteinizing hormone (LH), estradiol ($E_2$) and progesterone were measured daily during cetrorelix administration. Up to 4 embryos were transferred. Results: Mean age of enrolled patients was $32.0{\pm}3.4$ years (mean $\pm$ S.D.). All of 30 patients underwent oocyte pick-up, and embryo transfer was done in 28 patients. The total and mean numbers of received oocytes were 196 and $6.5{\pm}4.7$, the number of fertilized eggs was 111, and the fertilization rate was 56.6%. Total duration of FSH administration was $9.2{\pm}2.2$ days and mean of $24.3{\pm}7.7$ ampules of HP-FSH was administered. Total duration of cetrorelix administration was $5.7{\pm}1.9$ days. Serum LH and progesterone levels were maintained in the range of $1.4{\sim}2.9\;mIU/mL$ and $0.3{\sim}0.6\;ng/mL$, which respectively reflected effective prevention of premature LH surge. Clinical pregnancies were achieved in 9 patients, and overall clinical pregnancy rate was 30.0% per oocyte retrieval, and 32.1% per embryo transfer. Conclusion: GnRH antagonist is safe and convenient for COH for IVF-ET and effective with optimal pregnancy rate.

• Development and Reproduction
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• v.12 no.2
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• pp.125-132
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• 2008

Melatonin (N-acetyl-5-methoxytryptamine), a major hormone of pineal gland in vertebrates, is known to be associated with regulation of the dynamic physiological functions in general and has some functions on reproduction in the ovarian follicles in particular. And its antioxidant properties as a scavenger are also reported. The aim of this study was to investigate the effect of melatonin on the in vitro maturation of mouse germinal vesicle (GV)-stage oocytes. Oocyte maturation, apoptosis, and mRNA expression of melatonin receptor were analyzed in the cumulus cell-enclosed oocytes (CEOs) cultured with melatonin for 18 h. The CEOs were obtained from 3 wk-old ICR female mice cultured in media with 0, 0.1 nM, 10 nM, or 1,000 nM melatonin for 18 h. And then the extrusion of the first polar body was assessed to evaluate the maturation rate. The apoptosis and mRNA expression of melatonin receptor (Mtnr1-a and Mtnr1-b) in cumulus cells of each group were measured by TUNEL assay, ELISA, and real time RT-PCR after in vitro maturation(IVM). The addition of melatonin in the IVM medium significantly improved nuclear maturation of the mouse GV oocytes and the highest maturation rate were obtained from the group treated with 1,000 nM melatonin. Apoptosis was not detected in IVM oocytes, but detected in cumulus cells. And cumulus cells treated with 1,000 nM melatonin exhibited significantly lower apoptosis. In the group treated with 1,000 nM melatonin, the expression of melatonin receptor mRNA was decreased in CEOs. In conclusion, melatonin has a potentially important role for regulating oocyte maturation and reduces the apoptosis of cumulus cells in vitro.

• Korean Journal of Animal Reproduction
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• v.14 no.1
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• pp.1-8
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• 1990

These experiments were carried out to investigate the effects of cumulus cells for in vitro fertilization and development of bovine follicular oocytes matured in vitro. The bovine ovaries were obtained at a slaughter house and the follicular oocytes surrounded by cumulus cells were collected by puncturing follicles with 2~6 mm of diameter. Bovine oocytes were matured in vitro for 24~26 hours in a CO2 incubator with 5% CO2 in air at 39$^{\circ}C$. The medium used for maturation was TCM-199 supplemented with hormones, pyruvate, FCS and antibiotics. Epididymal spermatozoa were capacitated by in vitro culture for 2~3 hours in BO solution containing BSA(5mg/ml) and caffeine(2.5mM). Insemination was made by introducing about 10~15 matured oocytes into the suspension of capacitated spermatozoa. Six hour after insemination the eggs were transferred to TCM-199 supplemented with FCS(10%) and HEPES(25mM), cultured for 7~8 days with 10~15 eggs/well in 4-well multidishes(Nunc Co.) forming cumulus cell monolayer. The results obtained in these experiments were summarized as follows ; 1. The majority of the follicular oocytes with compacted cumulus cells existed in GV stage while those with dispersed or denuded cumulus cells existed GVBD and M II stage. 2. After 24~26 hours maturation, the maturation rates of the follicular oocytes cultured in TCM-199 containing hormones were slightly higher than those of oocytes cultured in medium without hormones, and the frequency of cumulus compacted or denuded oocytes reaching M II stage cultured in medium containing hormones was 75.7% or 51.7%, respectively(P<0.05). 3. After 20 hours in vitro insemination, percentages of ova fertilized were 61.4% or 51.4%, respectively, for cumulus oophorus intacted or removed, and increased frequency of ova with both male and female pronuclei was found when cumuli were present(P<0.05). 4. The rates of embryos developed to 2-, 4-, 8-, 16-cell and morula or blastocyst stage after cocultured with cumulus cells were 65.0%, 45.3%, 34.7%, 28.0% and 22.7%, respectively. The results for momla or blastocyst stage were significantly higher than those of the embryos cultured in the basic medium(P<0.05).

• International Journal of Oral Biology
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• v.36 no.1
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• pp.37-42
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• 2011

The working mechanism of bisphosphonate on bone cells is unclear despite its powerful inhibitory activity on bone resorption. The differentiation and activation of osteoclasts are essential for bone resorption and are controlled by the stimulatory RANKL and inhibitory OPG molecules. Teeth exhibit a range of movement patterns during their eruption to establish their form and function, which inevitably accompanies peripheral bone resorption. Hence, the mandible, which contains the teeth during their eruption processes, is a good model for revealing the inhibitory mechanism of bisphosphonate upon bone resorption. In the present study, RANKL and OPG expression were examined immunohistochemically in the mandible of rats with developing teeth after alendronate administration (2.5 mg/kg). The preeruptive mandibular first molars at postnatal days 3 to 10 showed the developing stages from bell to crown. No morphological changes in tooth formation were observed after alendronate administration. The number of osteoclasts in the alveolar bone around the developing teeth decreased markedly at postnatal days 3, 7 and 10 compared with the control group. RANKL induced strong positive immunohistochemical reactions in the dental follicles and stromal cells around the mandibular first molar. In particular, many osteoclasts with strongly positive reactions to RANKL appeared above the developing mandibular first molars at postnatal days 3 and 10. Immunohistochemical reactions with RANKL after alendronate administration were weaker than the control groups. However, the immunohistochemical reactivity to OPG was stronger after alendronate administration, at postnatal days 3 and 10. These results suggest that alendronate may decrease bone resorption by regulating the RANKL/OPG pathway in the process of osteoclast formation, resulting in a delay in tooth eruption.

• The korean journal of orthodontics
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• v.29 no.6 s.77
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• pp.663-672
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• 1999

A case is described, where an adolescent boy developed alopecia areata and alopecia totalis during the course of routine orthodontic treatment for the resolution of a dentoalveolar Class II division 1 malocclusion. The orthodontic treatment lasted 22 months, with a successful outcome. However, within eight months of the onset of treatment the patient lost all his hair Exhaustive medical tests and differential diagnosis determined that the etiolgy of the patient's alopecia was psychological stress evoked by the orthodontic treatment. Numerous reports suggest that psychological stress can cause alopecia by affecting the immune system. Therefore, it appears reasonable to assume that in the case of this patient, alopecia had resulted from stress effects on the immune system, leading to autoimmune disease-like conditions in tissues surrounding the scalp hair follicles. The alopecia condition was successfully reversed by daily oral and topical applications of vitamin D. It is concluded that the immune system plays a pivotal role in tissue remodeling around the teeth and elsewhere in the body, and that any conditions capable of affecting this system may cause unfavorable outcomes, such as alopecia.

• Development and Reproduction
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• v.4 no.1
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• pp.125-131
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• 2000

This study has used differential display-PCR, to amplify genes transcribed during the ovarian maturation induced by human chorionic gonadotropin (hCG). The cDNA expressed at the times of acquisition of oocyte maturational competence in red seabream (Pagrus major) following treatment with hCG was amplified and cloned. A full-length of cDNA for p. major was isolated using differential display-PCR and 5'RACE. This cDNA clone contained 2,662 nucleotides including the open reading frame that encoded 434 amino acids. Homology analyses, using the GenBank and EMBL general database searches, indicated that the nucleotides sequence of the cDNA does not have high homology with any other genes. This cDNA was judged to be a gene, which induction of maturational competence coincides with increase of mRNA related ovarian maturation. Consensus sequences which were consistent with protein kinase C phosphorylation sites and casein kinase II phosphorylation sites were identified. in vitro, the transcription level of mRNA related ovarian maturation increased between 9hr and 24hr following treatment of ovarian follicles with hCG. It was also increased after GtH-II (300 ng/ml) stimulation. Furthermore, in vivo, mRNA related ovarian maturation was rarely expressed prior to the acquisition of oocyte maturational competence, but was strongly expressed after the acquisition of oocyte maturational competence, suggesting that the hCG induction of maturational competence is brought about by the de novo synthesis of the mRNA related ovarian maturation in p. major.

• Development and Reproduction
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• v.11 no.2
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• pp.97-104
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• 2007

This study describes the spermatogenesis and sperm ultrastructure of the spiny top shell, Batillus cornutus using light and electron microscopy. The spiny top shells were collected by divers in the coastal water of Wandogun, Cheollanamdo, Korea(N $34^{\circ}13'$, E $126^{\circ}47'$) at May 2003. Spiny top shells of $60.0{\sim}69.9\;mm$ in shell height were used in this study. The testis comprises many spermatogenic follicles which contains germ cells in different developmental stages. The primary spermatocytes in the pachytene stage were characterized by synaptonemal complexes. The early spermatids were characterized by appearance of Golgi complex, increased karyoplasmic electron density and tubular mitochondria. In early spermatid the mass of proacrosomal granules consists of numerous heterogeneous granules with high electron density. From the mid-stage of spermiogenesis the well-developed mitochondria aggregate posterior to the nucleus, and surround the proximal and distal centrioles. In this stage, proacrosomal granules are condensed and form a acrosome with thin envelope. During the late spermiogenesis, the acrosome begins to elongate and then became conical. The sperm consists of head, mid-piece and tail. The head comprises a round nucleus and a conical acrosome. Acrosomal rod of microfibrous is observed between nucleus and acrosome. Five mitochondria observed in mid-piece. And tail has the typical "9+2" microtubular system originates from the centrioles.

• Journal of Life Science
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• v.26 no.11
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• pp.1345-1354
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• 2016

Alopecia areata (AA) is a common and tissue-specific autoimmune disease of hair follicle resulting in the loss of hair on the scalp and elsewhere on the body. Hair follicles is a unique organ because it has its own immune system and hormonal milieu and has a different immune state at each hair cycle stage. The collapses of anagen-dependent hair follicle immune privilege arise autoimmune attack, inducing ectopic MHC class I expression in the hair follicle epithelium and autoantigen presentation to autoreactive CD8+T cells, which results in AA. Clinical and experimental studies have pointed that psychological stress may also influence the hair follicle immune/hormone systems and contribute to the induction of AA. The key pathogenesis of AA is associated with immune privilege guardians (including ACTH, ${\alpha}-MSH$, and $TGF-{\beta}$), natural killer group 2D-positive (NKG2D+) cells (including NK and CD8+T cells), and stress hormones (including CRH and substance P). Effective treatments for AA are still demanded. One of the future targets of treatment will be the modification of hair follicle immune privilege including stress. Recent studies have reported that JAK inhibitors and immunomodulators used in other autoimmune disease, such as psoriasis, atopic dermatitis, and rheumatoid arthritis, Tregs, platelet-rich plasma therapy, statins, and prostaglandin anaolgues are effective for AA. Here the article reviews the recent understanding in the pathogenesis associated with perifollicular endocrine/immunology and new treatments of AA.