• 한국수정란이식학회지
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• 제9권1호
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• pp.15-21
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• 1994

This experiment was investigated the effect of cell stage of embryos at 48 hours post-insemination On in vitro development of IVF embryos. The ovaries of Korean native cows or heifers were obtained from an abattoir and kept on 25 to 28$^{\circ}C$ and transported to laboratorty within 2 hrs. The oocytes were matured in vitro(IVM) for 24 hrs. in TGM-199 supplemented with 35 $\mu$g/$m\ell$ FSH, 10 $\mu$g/$m\ell$ LH, 1 $\mu$g/$m\ell$ estradiol-17$\beta$ and granulosa cells at 39$^{\circ}C$ under 5% $CO_2$ in air. They were fertilized in vitro(IVF) by epididymal spermatozoa treated with heparin for 24 hrs. , and then the zygotes were co-cultured in vitro(IVC) with bovine oviductal epithelial cells for 7 to 9 days. At 48 hrs. post-insemination, the embryos were classfied into 5 to 8-cell, 3 to 4-cell or 2-cell stage and then were co-cultured in vitro(IVC) with bovine oviductal epithelial cells until the embyos reached blastocyst stage. Embryos developed to blastocyst stage were stained with Hoechst 33342 for cell counting. The embryos of 5 to 8-cell stage at 48 hrs. post-insemination with grade I oocytes were significantly (P<0.05) better developed to blastocysts(63.0%) than 3 to 4-cell(42.0%) and 2-cell stage(2.7%) embryos which delayed in the early cleavage, and those embryos cleaved faster in the very early stage seemed to develop to blastocysts earlier. These results indicate that the embryos cleaved faster at 48 hrs. post-insemination seemed to develop to blastocysts earlier.

• 한국수정란이식학회지
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• 제9권1호
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• pp.117-125
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• 1994

In vitro development of parthenogenetic embryo was examined after ethanol treatment of follicular oocytes matured in vitro for 42, 48, 54 and 60h in the pig. The follicular oocytes were matured in TCM 199 containing 15% FCS and gonadotrophins in an atmosphere of 39 $^{\circ}C$ 5% $CO_2$. The cumulus-free oocytes were activated by 10% ethanol treatment in M2+4mg /ml BSA for 10 min. The ethanol-activated oocytes were washed and further cultured in TCM199+20%FCS containing granulosa cell monolayer. Maturation rates at 42, 48, 54 and 60h of IVM were 75.0, 86.5, 81.6 and 87.9%, respectively. Thus the oocytes maturated in vitro for longer periods did not improve nuclear maturation much. Pronuclear formation rates at 18h post-activation in ethanol-activated oocytes were 21.9, 25.0, 47.4 and 32.6%. The cytoplasmic maturation leading to pronuclear formation upon activation increased when the I VM period was extended from 42 to 54h. When the activated oocytes were cultured for 96~120h to analyse early development of the activated oocytes, the rates of embryonic development upto $\leq$ 5~8 cell were 5.3, 5.8, 12.0 and 11.7% among the cultured embryos. The result indicate that earlier development of activated porcine occyte is dependent on the duration of oocyte maturation, and that better development could be obtained from the oocyte matured for 54h.

• 한국수정란이식학회지
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• 제12권3호
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• pp.283-291
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• 1997

In vitro development of bovine embryos is affected by many factors such as energy substrates, amino acids, and some growth factors. It has been reported that mRNA of insulin, PDGF and their receptors are detected in cow embryos, and that some chelating agents such as EDTA and transferrin have beneficial role on mouse and bovine embryos. The author hypothesized that insulin, transferrin arid PDGF added to a culture medium increase in vitro development of bovine embryos by chelating toxic substance(s) or increasing cell growth and metabolism. Immature oocytes from slaughtered ovaries of Holstein cows and heifers were matured for 24 hours in a TCM199 containing 10% fetal calf serum, FSH, LH and estradiol with granulosa cells in vitro. Matured oocytes were coincubated with sperm for 30 hours in a modified Tyrode's medium (IVF). Embryos cleaved to 2- to 4-cell at 30 hours after IVF were selected and cultured in a 30-$\mu$l drop of a synthetic oviduct fluid medium (SOFM) containing 0.8% BSA, Minimum Essential Medium essential and non-essential amino acids, and insulin, transferrin or PDGF for 9 days. Supplementation of a SOFM with insulin, and /or transferrin did not increase develop-mental rate to expanding and hatching blastocyst of 2- to 4-cell bovine embryos compared with control. The highest developmental rate to hatching blastocyst was shown when PDGF was added at the concentration of 10 ng /ml among the supplementing doses tested in the present study (p<0.05). Addition of PDGF without insulin to a SOFM could not increase embrye development, but combined addition of PDGF with insulin significantly increased (p<0.05) embryo development to hatching blastocyst (50%) compared with control (38%). In conclusion, insulin and PDGF supplemented to a SOFM may act synergistically and have beneficial effect on in vitro development of 2- to 4-cell bovine embryos matured and fertilized in vitro.

• 한국발생생물학회지:발생과생식
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• 제24권3호
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• pp.177-185
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• 2020

Although many aquaporin (AQP) transcripts have been demonstrated to express in the female reproductive tract, the defined localizations and functions of AQP subtype proteins remain unclear. In this study, we investigated the expression of AQP1, AQP3, AQP5, AQP6, and AQP9 proteins in female reproductive tract of mouse and characterized their precise localizations at the cellular and subcellular levels. Immunofluorescence analyses for AQP1, AQP3, AQP6, and AQP9 showed that these proteins were abundantly expressed in female reproductive tract and that intense immunoreactivities were observed in mucosa epithelial cells with a subtype-specific pattern. The most abundant aquaporin in both vagina and uterine cervix was AQP3. Each of AQP1, AQP3, AQP6, and AQP9 exhibited its distinct distribution in stratified squamous or columnar epithelial cells. AQP9 expression was predominant in oviduct and ovary. AQP1, AQP3, AQP6, and AQP9 proteins were mostly seen in apical membrane of ciliated epithelial cells of the oviduct as well as in both granulosa and theca cells of ovarian follicles. Most of AQP subtypes were also expressed in surface epithelial cells and glandular cells of endometrium in the uterus, but their expression levels were relatively lower than those observed in the vagina, uterine cervix, oviduct and ovary. This is the first study to investigate the expression and localization of 5 AQP subtype proteins simultaneously in female reproductive tract of mouse. Our results suggest that AQP subtypes work together to transport water and glycerol efficiently across the mucosa epithelia for lubrication, proliferation, energy metabolism and pH regulation in female reproductive tract.

• 대한한방부인과학회지
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• 제24권1호
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• pp.27-41
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• 2011

Objectives: This study was designed to investigate the effect of Ojeok-san(OJS) on the estradiol valerate(EV)-induced polycystic ovaries(PCO) in rats. Methods: PCO was induced by single intramuscular injection with estradiol valerate(EV)(4mg) in female rats. Normal group(n=8) were injected with sesame oil and orally administrated distilled water for sixty days. PCO control group (n=8) were injected with EV and orally administrated distilled water for sixty days. OJS treated group(n=8) were injected with EV and orally administrated OJS for sixty days. Then, we measured weights of body, ovaries, adrenal glands and uterus of rats. The histomorphometrical changes of ovaries were also evaluated. And we observed the NGF and CRF expression by immunohistochemistry. Results: The results were as follows - The weights(mg) of ovaries in OJS treated group($57.4{\pm}9.4$) were significantly increased(p < 0.05) compared with PCO control group($42.3{\pm}8.5$). - The numbers of mature follicles in OJS treated group($9.3{\pm}2.5$) were significantly increased(p < 0.05) compared with PCO control group($6.1{\pm}2.1$). - The expressions of NGF-immunoreactive cells in the ovarian granulosa cells in OJS treated group were weaker than PCO control group. Conclusion: From the these results, we concluded that Ojeok-san(OJS) has inhibitory effect on the development of EV-induced polycystic ovary. And it's effect maybe related with decreased NGF activities in the ovary.

• 대한수의학회지
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• 제23권1호
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• pp.57-67
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• 1983

The present study was carried out to observe the histopathological changes in the mammary gland of lactating rats and rabbits injected with dexamethasone. White rats were intramuscularly injected with 0.25mg, 0.5mg or 1.0mg of dexamethasone sodium phosphate (containing $9{\alpha}$-fluoro-$16{\alpha}$-methylprednisolone, 5.0mg/ml) daily for 3 to 10 days on the 3rd day after parturition and white rabbits were intramammary infused with 4mg or 20mg of dexamethasone daily for 4 days on 7th day after parturition. The histopathological changes of the mammary glands, ovaries and adrenal glands of rats and rabbits were observed with light microscope. In the mammary glands of rats, the microscopic findings encountered were decrease of the milk in the alveolar lumina, necrosis and desquamation of epithelial cells, atrophy of alveoli, proliferation of fibroblasts and thickness of alveolar walls, destruction of alveoli, presence of fat droplets within the glandular epithelial cells, infiltration of mononuclear cells and proliferation of adipose tissue, which were relative to the dose and duration of injection. Especially, in the cases of the administration of large doses or long duration, there were severe fibrosis and focal necrosis of glandular tissue. In the mammary glands of rabbits, the morphological changes were similar to those findings in the rats. The milk in the alveolar lumina was decreased gradually according to the dose and duration of injection, while milk fat concentration regarded to increase. In the histological findings of ovaries, necrosis of granulosa cellos, vacuolization and necrosis of luteal cells, atrophy and necrotic foci in the corpora lutes were observed. In the adrenal glands, hyperemia, hemorrhage, vacuolization of adrenal cortical cells, necrotic foci and atrophy of adrenal cortex were observed.

• Animal Bioscience
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• 제34권11호
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• pp.1766-1775
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• 2021

Objective: The oxidative stress status and changes of chicken ovary tissue after shading were studied, to determine the mechanism of the effect of shading on follicular development. Methods: Twenty healthy laying hens (40 weeks old) with uniform body weight and the same laying rate were randomly divided into two groups (the shading group and normal light group). In the shading group, the cage was covered to reduce the light intensity inside the cage to 0 without affecting ventilation or food intake. The normal lighting group received no additional treatment. After 7 days of shading, oxidative stress related indicators and gene expression were detected. Results: Analysis of paraffin and ultrathin sections showed that apoptosis of ovarian granulosa cells (GCs) increased significantly after light shading. Enzyme linked immunosorbent assay results revealed that the levels of total antioxidant capacity, malondialdehyde, superoxide dismutase (SOD), glutathione, catalase (CAT), and other substances in the sera, livers, ovaries, and follicular GCs of laying hens increased significantly after shading for 7 days; and reactive oxygen species (ROS) levels in the livers of laying hens also increased significantly. ROS in the serum, ovarian and GCs also increased. After shading for 7 days, the levels of 8-hydroxy-2 deoxyguanosine in the sera and ovarian tissues of laying hens increased significantly. Cell counting kit-8 detection showed that the proliferation activity of GCs in layer follicles decreased after shading for 7 days; the expression level of the anti-apoptotic gene B-cell lymphoma-2 in ovarian tissue and follicular GCs was significantly reduced, and the expression levels of pro-apoptotic caspase 3 (casp3), and SOD, glutathione peroxidase 2 (GPX2), and CAT were all significantly increased. Conclusion: Oxidative stress induced by shading light has a serious inhibitory effect on follicular development during reproduction in laying hens.

• Clinical and Experimental Reproductive Medicine
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• 제46권2호
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• pp.76-86
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• 2019

Objective: This study was performed to explore the possibility that each oocyte and its surrounding cumulus cells might have different genetic expression patterns that could affect human reproduction. Methods: Differential gene expression analysis was performed for 10 clusters of cumulus cells obtained from 10 cumulus-oocyte complexes from 10 patients. Same procedures related to oocyte maturation, microinjection, and microarray analyses were performed for each group of cumulus cells. Two differential gene expression analyses were performed: one for the outcome of clinical pregnancy and one for the outcome of live birth. Results: Significant genes resulting from these analyses were selected and the top 20 affected pathways in each group were analyzed. Circadian entrainment is determined to be the most affected pathway for clinical pregnancy, and proteoglycans in cancer pathway is the most affected pathway for live birth. Circadian entrainment is also amongst the 12 pathways that are found to be in top 20 affected pathways for both outcomes, and has both lowest p-value and highest number of times found count. Conclusion: Although further confirmatory studies are necessary, findings of this study suggest that these pathways, especially circadian entrainment in cumulus cells, may be essential for embryo development and pregnancy.

• 한국수정란이식학회지
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• 제33권4호
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• pp.305-311
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• 2018

Gonadotropin releasing hormone (GnRH) centrally plays a role in control of the hypothalamic-pituitary-gonadal axis-related hormone secretions in the reproductive neuroendocrine system. In addition, hormone receptors like luteinizing hormone receptor (LHR) are important element for hormones to take effect in target organ. However, ageing-dependent changes in terms of the distribution of GnRH neurons in the brain and LHR expression in the acyclic ovary have not been fully understood yet. Therefore, we comparatively investigated those ageing-dependent changes using young (1-5 months), middle (11-14 months) and old (21-27 months) aged female mice. Whereas a number of GnRH positive fibers and neurons with monopolar or bipolar morphology were abundantly observed in the brain of the young and middle aged mice, a few GnRH positive neurons with multiple dendrites were observed in the old aged mice. In addition, acyclic ovary without repeated development and degeneration of the follicles was shown in the old aged mice than others. LHR expression was localized in theca cells, granulosa cell, corpora lutea and atretic follicle in the ovaries from young and middle aged mice, in contrast, old aged mice had few positive LHR expression on the follicles due to acyclic ovary. However, the whole protein level of LHR was higher in the ovary of old aged mice than others. These results are expected to be used as an important basis on the relationship between GnRH and LHR in old aged animals as well as in further research for reproduction failure.

• 한국동물생명공학회지
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• 제37권3호
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• pp.155-161
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• 2022

Pregnancy-associated plasma protein-A (PAPP-A) is known as an important biomarker for fetal abnormality during first trimester and has a pivotal role in follicle development and corpus luteum formation. And also, it is being revealed that an expression of PAPP-A in various cells and tissues such as cancer and lesion area. PAPP-A is the major IGF binding protein-4 (IGFBP-4) protease. Cleavage of IGFBP-4 results in loss of binding affinity for IGF, causing increased IGF bioavailability for proliferation, survival, and migration. Additionally, PAPP-A can be used as a promising therapeutic target for healthy longevity. Despite growing interest, almost nothing is known about how PAPP-A expression is regulated in any tissue. This review will focus on what is currently known about the zinc metalloproteinase, PAPP-A, and its role in cells and tissues. PAPP-A is expressed in proliferating cells such as fetus in uterus, granulosa cells in follicle, dermis in wound, cancer cells, and Sertoli cells in testis. They have common characteristics of proliferation faster than normal cells with stimulating IGFs action and inhibiting IGFBPs. The PAPP-A functions and expression studies in livestock have not yet been conducted much. Further studies are needed to use PAPP-A as a marker for healthy longevity in animal science.