Endoplasmic reticulum stress in periimplantation embryos / 대한생식의학회지

Stress coping mechanisms are critical to minimize or overcome damage caused by ever changing environmental conditions. They are designed to promote cell survival. The unfolded protein response (UPR) pathway is mobilized in response to the accumulation of unfolded proteins, ultimately in order to regain endoplasmic reticulum (ER) homeostasis. Various elements of coping responses to ER stress including Perk, Ask1, Bip, Chop, Gadd34, Ire1, Atf4, Atf6, and Xbp1 have been identified and were found to be inducible in oocytes and preimplantation embryos, suggesting that, as a normal part of the cellular adaptive mechanism, these coping responses, including the UPR, play a pivotal role in the development of preimplantation embryos. As such, the UPR-associated molecules and pathways may become useful markers for the potential diagnosis of stress conditions for preimplantation embryos. After implantation, ER stress-induced coping responses become physiologically important for a normal decidual response, placentation, and early organogenesis. Attenuation of ER stress coping responses by tauroursodeoxycholate and salubrinal was effective for prevention of cell death of cultured embryos. Further elucidation of new and relevant ER stress coping responses in periimplantation embryos might contribute to a comprehensive understanding of the regulation of normal development of embryonic development and potentiation of embryonic development in vitro.

Effect of electromagnetic field exposure on the reproductive system / 대한생식의학회지

The safety of human exposure to an ever-increasing number and diversity of electromagnetic field (EMF) sources both at work and at home has become a public health issue. To date, many in vivo and in vitro studies have revealed that EMF exposure can alter cellular homeostasis, endocrine function, reproductive function, and fetal development in animal systems. Reproductive parameters reported to be altered by EMF exposure include male germ cell death, the estrous cycle, reproductive endocrine hormones, reproductive organ weights, sperm motility, early embryonic development, and pregnancy success. At the cellular level, an increase in free radicals and [Ca2+]i may mediate the effect of EMFs and lead to cell growth inhibition, protein misfolding, and DNA breaks. The effect of EMF exposure on reproductive function differs according to frequency and wave, strength (energy), and duration of exposure. In the present review, the effects of EMFs on reproductive function are summarized according to the types of EMF, wave type, strength, and duration of exposure at cellular and organism levels.

Effect of electromagnetic field exposure on the reproductive system / 대한생식의학회지

The safety of human exposure to an ever-increasing number and diversity of electromagnetic field (EMF) sources both at work and at home has become a public health issue. To date, many in vivo and in vitro studies have revealed that EMF exposure can alter cellular homeostasis, endocrine function, reproductive function, and fetal development in animal systems. Reproductive parameters reported to be altered by EMF exposure include male germ cell death, the estrous cycle, reproductive endocrine hormones, reproductive organ weights, sperm motility, early embryonic development, and pregnancy success. At the cellular level, an increase in free radicals and [Ca2+]i may mediate the effect of EMFs and lead to cell growth inhibition, protein misfolding, and DNA breaks. The effect of EMF exposure on reproductive function differs according to frequency and wave, strength (energy), and duration of exposure. In the present review, the effects of EMFs on reproductive function are summarized according to the types of EMF, wave type, strength, and duration of exposure at cellular and organism levels.

Effects of early prepubertal exposure to bisphenol A on the onset of puberty, ovarian weights, and estrous cycle in female mice / 대한생식의학회지

OBJECTIVE: Bisphenol A (BPA) is a chemical used extensively to manufacture plastics and epoxy resin liners for food and beverage cans. BPA, with properties similar to estrogen, has endocrine-disrupting effects. In the present study, we examined the effects of early prepubertal BPA exposure on the onset of puberty and reproductive parameters such as estrous cycle and reproductive organ weights in female mice. METHODS: Female mice were injected subcutaneously at postnatal day (PND) 8 with BPA (0.1, 1, 10, 100 mg/kg) in sesame oil or with sesame oil alone. Body weight was measured from PND 10 to 70. Vaginal opening and estrous cycle were monitored from PND 20 to 29. Animals were sacrificed at PND 25, 30, and 70, and the ovary and uterus weights were measured. RESULTS: Early prepubertal exposure to BPA (10 and 100 mg/kg) significantly decreased body weight from PND 18 to 30. BPA treated mice at testing dose levels showed early opening of the vagina compared to the control group. The number of estrous cycle and days of estrus were significantly decreased in high dose (100 mg/kg) BPA treated mice. The ovary weight at PND 25 and 30 was significantly decreased in all BPA treatment groups. CONCLUSION: Early prepubertal exposure to BPA accelerated the onset of puberty but decreased reproductive parameters in female mice.

Effect of Steroid Hormones on Expression and Localization of Aquaporin-4, -5 and -8 Genes in Mouse Uterine Endometrium / 대한불임학회지

No abstract available.

The Expression of p57(kip2) in Mouse Endometrium During Estrus Cycle and Pre- and Peri-implantation Periods / 대한산부인과학회잡지

OBJECTIVE: This study was to investigate the localization of CDK inhibitor, p57(kip2) in mouse endometrium during the estrus cycle and pre- and peri-implantation periods. METHODS: The p57(kip2) protein was immunostained from endometrium of mouse sacrificed at diestrus, proestrus, estrus, and metestrus cycle, and at day 1-6 post-coitum (p.c.). RESULTS: The staining in the luminal epithelium was very weak in comparison with glandular and stromal cells. In diestrus stage, immunoreactivity of p57(kip2) was heterogeneously strong in parts of decidualized or degenerated stromal cells. In proestrus stage, strong immunoreactivity p57(kip2) was largely found in stromal cells. But, p57(kip2) was showed low immunoreactivity in estrus stage. In metestrus stage, immunoreactivity of p57(kip2) was heterogeneously strong in decidualized stromal cells. In day 1-2 p.c., immunoreactivity of p57(kip2) was low in some endometrial stromal cells. In day 3-4 p.c., immunoreactivity of p57(kip2) was strong in some endometrial stromal cells. In day 5-6 p.c., immunoreactivity of p57(kip2) was strong in decidual cells. CONCLUSION: These suggest that p57(kip2) may play an essential role in endometrial differentiation for maintenance of implantation, especially decidualization of endometrial stromal cells.

The Expression of p27(kip1) and p57(kip2) in Mouse Endometrium / 대한산부인과학회잡지

OBJECTIVE: This study was to investigate the expression of CDK inhibitors, p27(kip1) and p57(kip2) in mouse endometrium during the estrus cycle and pregnant period. METHODS: Total RNA and protein were extracted from endometrium of mouse sacrificed at diestrus, proestrus, estrus, and metestrus cycle, and at day 1-6 post-coitum (p.c.), then semi-quantitative RT-PCR and western blotting of p27(kip1) and p57(kip2) was carried out. RESULTS: p27(kip1) and p57(kip2) mRNA was highly expressed in diestrus and proestrus stage than estrus and metestrus stage. In comparison with estrus cycle, p27(kip1) and p57(kip2) mRNA level was highly maintained in gestational endometrium (except p27(kip1) of day 5 p.c). p57(kip2) protein level was relatively low from day 1 p.c. to day 4 p.c. But it was significantly increased in day 5 p.c. and day 6 p.c. CONCLUSION: These results show that p27(kip1) and p57(kip2) may play a role in endometrial differentiation for regular estrus cycle and implantation, and especially p57(kip2) may play an essential role in endometrial differentiation for maintenance of implantation.

Granulocyte Macrophage-Colony Stimulating Factor Signaling in Development of Mouse Embryos / 대한불임학회지

OBJECTIVE Present study was aimed to verify the effect of granulocyte macrophage-colony stimulating factor (GM-CSF) in the preimplantation development of mouse embryos and the involvement of the mitogen activated protein kiase (MAPK) in the GM-CSF signaling. METHODS: Two-cell embryos were cultured for 96 h in the presence or absence of GM-CSF (0, 0.4, 2, 10 ng/ml) and PD98059, a MEK inhibitor (10 muM). Morphological development, cell number per blastocyst, and apoptotic nuclei, were eamined. MAPK activity of embryonic immunoprecipitate by MAPK (ERK1/2) antibody was measured by in vitro phosphorylation of myelin basic protein. RESULTS: At post hCG 122 h the embryonic development among the experimental groups was significantly different (p=0.018). The rate of blastocyst development and cell number per embryo were the highest in 2 ng/ml GM-CSF treatment group. The percent of apoptotic cells of the GM-CSF-treated embryos was the lowest among the group. in blastocysts, GM-CSF treatment transiently increased MAPK activity. PD098059 attenuated the effect of GM-CSF on the morphological development, increase in cell number per blastocyst, down regulation of apoptosis, and upregulation of MAPK activity, suggesting that activation of MAPK cascade possibly mediated the embryotropic effect of GM-CSF. CONCLUSION: This result suggested that GM-CSF potentiated the development of preimplantation mouse embryos by activation of MAPK.

The Expression of p27kip1 and p57kip2 in Mouse Placenta During Gestation / 대한산부인과학회잡지

OBJECTIVE: This study was to investigate the expression of CDK inhibitors, p27kip1 and p57kip2 during the growth and differentiation of mouse placenta. METHODS: Total RNA and protein were extracted from placenta of mouse sacrificed at day 12, 14, 16, 18 post-coitum (p.c.), then semi-quantitative RT-PCR and western blotting of p27kip1 and p57kip2 was carried out, respectively. RESULTS: p27kip1 mRNA was highly expressed in 18 days p.c. then other groups. But, p57kip2 mRNA expression was high in 12, 14, and 16 days p.c., then decreased in 18 days p.c. p27kip1 expression pattern was similar with mRNA. But, p57kip2 was higher in 14 days p.c. than other groups. CONCLUSION: This result shows that p27kip1 may play a role in late period of mouse placental development, and p57kip2 may play a role in middle period of mouse placental development.

Increase in Transepithelial Resistance Mouse Sertoli Cells by Leydig Cells Coculture / 대한남성과학회지

PURPOSE: To verify the regulation of transepithelial resistance (TER) of Sertoli cells by Leydig cells in mouse. MATERIALS AND METHODS: Primary culture of Sertoli cells was established on cell culture plate insert and monolayer culture was subjected to coculture in the Leydig cell culture. Changes in TER was monitored for 48 h using the conductivity meter equipped with two electrodes system. RESULTS: TER gradually increased according to the development of monolayer of Sertoli cells on the cell culture plate insert. Net changes in TER of Sertoli cells culture was significantly higher under the Leydig cells coculture compared to control after 48 h of coculture. CONCLUSIONS: It is the first report about the increase in TER of Sertoli cells by Leydig cells in vitro. Paracrine interaction between Leydig cells and Sertoli cells might be involved in the development of functional blood testis barrier which is made by tight junctions between Sertoli cells in mouse testis.

Expression of OB-R, Regulation of Mitogen Activated Protein Kinase Activity and Maturation by Leptin in Mouse Oocytes / 대한불임학회지

OBJECTIVE: To verify the expression of leptin receptor (OB-R) in oocytes and preimplantation embryos, the involvement of mitogen activated protein kinase (MAPK or Erk1/2) in the leptin signaling, and effect of leptin on the oocyte maturation in mice. METHOD: RT-PCR analysis of OB-R was conducted in germinal vesicle (GV)-intact and MII stage oocytes, and 1, 2, 8-cell embryos and blastocysts. Germinal vesicle breakdown (GVB), polar body extrusion, monitored in the presence or absence of leptin (1 microM). Following the leptin treatment, temporal changes in MAPK activity were verified by immunoprecipitation and in vitro kinase assay in MII oocytes. RESULTS: The expression of OB-R mRNA was found in GV and MII oocyte but not in the embryos. MAPK activity of the MII oocytes was significantly increased by brief incubation in the HTF supplemented with leptin (1 microM). Priming of PD098059, a MEK inhibitor to leptin treatment attenuated the activation of MAPK by leptin in MII oocytes. Following 24 hrs of culture of the GV oocytes, leptin significant increased the GVB and 1st polar body extrusion. CONCLUSION: This result suggested that functional interaction between leptin and OB-R resulted in potentiation of MAPK (Erk1/2) activity in MII oocytes through MEK activation and that leptin might be a local regulator of meiotic maturation of the mouse oocytes.

Expression of Mouse Homolog of Glucosamine-6-Phosphate Deaminase in Reproductive Organs / 대한남성과학회지

PURPOSE: To verify the expression of the mouse homolog of glucosamine-6-phosphate deaminase (GNPDA) in the testis and reproductive organs. MATERIALS AND METHODS: Expression of GNPDA was examined using a polyclonal antibody raised against a synthetic oliogopeptide of the N-terminus of the protein. RESULTS: In Western blots, an immunoreactive band of Mr 35 kDa was detected in the testis, ovary, and uterus extracts. Expression of GNPDA was greater in the adult than in the immature testis. With immunostaining, a positive signal was found in the cytoplasm of interstitial, Sertoli, and germ cells of adult testis. In the ovary, positive staining was found in the interstitial and luteal cells. Conclusion: The expression of GNPDA in many reproductive tissues suggests that the enzyme plays a housekeeping role in cell physiology and in the differentiation of the seminiferous tubules.

Analysis of Calcium-Binding Proteins of Mouse Spermatozoa by 2-Dimentional SDS-PAGE / 대한남성과학회지

PURPOSE: To verify the expression of calcium-binding proteins in mouse spermatozoa. MATERIALS AND METHODS: Mouse sperm proteins were subjected to 2-dimentional SDS-PAGE combined with calcium shift in the presence or absence of Ca2+, Zn2+ or EDTA. RESULTS: In the epididymal sperm extracts, 10 kinds of protein spots showed mobility shift in the gel containing Ca2+. Most of the CBPs disappeared after the acrosome reaction (AR) induced by Ca2+-ionophore A23187, suggesting that they originated from acrosome and/or the plasma membrane overlaying the acrosome. CONCLUSIONS: The CBPs might be involved in acrosome reaction of mouse spermatozoa. Protein database of sperm CBPs might be useful for diagnosis of male infertility.

Expression of Glucosamine-6-Phosphate Deaminase (GNPDA) in Mouse Epididymis / 대한남성과학회지

PURPOSE: To verify the expression of mouse homologue of glucosamine-6-phosphate deaminase (GNPDA) in mouse epididymis. MATERIALS AND METHODS: GNPDA expression was examined in mouse epididymis using the polyclonal antibody raised against a synthetic oligopeptide of N-terminus of mouse homologue of GNPDA. RESULTS: In Western blot, immunoreactive band of Mr. 33 kDa was detected in the cauda epididymal extracts. In immunostaining, positive signal was found in the cytoplasm and stereocillia of epithelial cells of cauda epididymal tubule, and interstitial cells of adult epididymis. CONCLUSION: Expression of GNPDA in epididymis suggests that it possibly involved in absorption and intracellular digestion of luminal contents, and the remodeling of epithelia in mouse epididymis.

Preimplantation Development and Apoptosis of Mouse Embryos in the Medium Containing Extracellualr Matrix / 대한불임학회지

OBJECTIVE: To verify the effect of Matrigel, a ECM complex from Engelbreth-Holm-Swarm(EHS) mouse sarcoma on the preimplantation development and apoptosis of mouse fertilized eggs. METHOD: Late Pronucleus stage eggs were cultured through the blastocyst stage in the presence of Matrigel (0.5%, v/v). Characteristics of apoptosis and cell number assessed by Hoecst staining and TUNEL labeling at the blastocyst stage, respectively. RESULTS: Morphological development, number of cells per embryo was significantly increased but rate and number of TUNEL positive nuclei of the embryo were decreased in the presence of Matrigel. CONCLUSION: This result suggested that at low concentration of Matrigel improves both viability and morphological development in the preimplantation mouse embryos.

Spermatozoa Characteristics of Streptozotocin-induced Diabetic Wistar Rat: Acrosome Reaction and Spermatozoa Concentration / 대한불임학회지

SUMMARY: Some of the information concerning sexual function in the male diabetes has been focused upon the problems of endocrine or semen parameters. However, the characteristics of acrosome reaction and spermatozoa concentration at the epididymis and vas deferens have scarcely been studied, and the causes of the infertility has not been critically identified. So, we designed to inspect the spermatozoa concentration and the characteristics of acrosome reaction at epididymis and was deferens of diabetic Wistar rat induced by streptozotocin (STZ, 70 mg/kg, ip). Experimental animal was sacrificed at 3 days and 14 days after the STZ injection. In the diabetes-induced rat, the levels of insulin and glucose had a pattern of inverse proportion. The spermatozoa concentrations in caput and corpus epididymis were significantly decreased in all diabetic condition. In cauda epididymis, however, there was significant decrease in sperm concentration at 14 days onward. In diabetic rat, the spontaneous reaction rate of spermatozoa of cauda and was deferens were significantly higher than the control group. The ARIC (acrosome reaction to ionophore challenge) value of caudal sperm was 28.7 at control, 22.1 at 3 days, and 8.3 at 14 days. In the present study the spermatozoa concentration was decreased and the spontaneous reaction rate was increased by diabetes. In ARIC-test, it is revealed that the fertility of spermatozoa of 14 days group was lower than control or 3 days group. Diabetes mellitus may be provoke the decreased fertilization rate and subsequent infertility.

The Effect of Uterine Environment during Peri-implantation Period on theUltrastructure of Zona Pellucida in Mouse Oocytes and Embryos / 대한불임학회지

In the studies on the hatching mechanisms in mammals, many investigators focused on the embryonic intrinsic factor(s) in in vitro culture, but the uterine environment as the extrinsic factor(s) is thought to play an important role in hatching mechanism. Therefore, to evaluate the effect of uterine environment on the hatching event in vivo, the immature(GV) and ovulated(MII) oocytes, and the late 2-cell embryos of mouse were transferred to pseudopregnant foster mother's uterus during peri-implantation period. So it was verified whether there would happen hatching by only uterine environment independently on embryonic stage. The ultrastructural changes of the zona surface of transferred group were compared with those of in vivo and vitro group by SEM. 36 hrs after transfer, the immature and ovulated oocytes almost degenerated, and the late 2-cell embryos developed to various embryonic stages. However, the embryos which didn't develop to blastula stage did not hatch. The ultrastructural network of ZP in transferred group seemed to be smoothed uniformly, which was different from in vitro group. In conclusion, it is suggested that the uterine environment during peri-implantation period enhances the embryo hatching by provoking the structural change of ZP.

Expression of Oscillin in Mouse Testis / 대한남성과학회지

PURPOSE: To verify the expression of the egg activator oscillin in mouse testis and adult organs. MATERIALS AND METHODS: Genomic PCR using primers for oscillin was conducted to confirm that the PCR product was derived from cDNA. Total RNA isolated from developing, immature, and mature testis was subjected to RT-PCR and restriction analysis. In situ hybridization of adult testis was performed to localize the oscillin transcript using cRNA probe. RESULTS: Genomic PCR using the primer for RT-PCR revealed no amplification product, suggesting that the oscillin gene consists of at least two exons. The RT-PCR product of oscillin mRAN was detected in testis beginning 2 weeks after birth. Oscillin mRAN was detected in both germ and somatic cells such as Sertoli and Leydig cells by in situ hybridization. The testis showed al high level of oscillin mRAN compared with other adult organs. CONCLUSION: Oscillin is not a testis-specific transcript and therefore may have another function intracellularly as well ad serving as a trigger for egg activation.

The Effect of Free Radical Scavenger, Superoxide Dismutase ( SOD ) in the Early Embryo Culture of Mice / 대한산부인과학회잡지

It is well known that developmental delay or arrest occurs before implantation in mamals, which have undergone in vitro culture. Recently, these phenomena are being attributed to oxygen free radicals, and successful cell culture are being obtained by lowering the oxygen environment of in vitro culture. This is due to the fact that the oxygen concentration in the fallopian tube is around 5%, which is lower than the room air 20% concentraition for in vitro culture. Superoxide dismutase(SOD), which are free radical scavenger, were added to early embryo mice culture and compared the free radical level at each stage of development with that of culture in which free radical scavenging agent was not added. The highest free radical level in control group observed in the late 2-cell stage and was markedly decreased if SOD added to culture media. Blastulation rate, hatching rate, and blastomere count as the parameters of embryo development was significantly increased in the experimental group. This study suggests the possibility of improvement in in-vitro embryo culture with media including free radical scavengers.

Effect of Seminal Vesicle Fluid Components on Acrosome Reaction of Mouse Epididymal Sperm / 대한불임학회지

This study aimed to evaluate the effect of seminal vesicle fluid (SVF) on the acrosome reaction (AR) occurred spontaneously or induced by Ca2+ ionophore A23187, follicular fluid, and progesterone in mouse epididymal sperm. SVF was divided into high (MW>10 kM)) and low (MW<10 kD) fractions by ultrafiltration. The low MW fraction of SVF decreased the rate of spontaneous AR, however the high MW fraction did not. It suggested that the low MW fraction of SVF might have contained decapacitation factor(s) responsible for prolonging of time need for capacitation. When sperm preincubated for 60 min in the presence of SVF, the rate of AR induced by A23187 was decreased, but prolongation of preincubation time for 120 min significantly potentiated the AR by A23187. It suggested that addition of SVF into sperm preincubation medium imposed the epididymal sperm a condition similar to ejaculation. AR induced by human follicular fluid or progesterone was also inhibited by SVF. It suggested that substance in SVF might have affected AR of mouse sperm by inhibiting the interaction between AR inducing ligands and sperm surface receptors involved in acrosomal exocytosis.