• Asian-Australasian Journal of Animal Sciences
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• 제26권6호
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• pp.795-803
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• 2013

Various endometrial genes in ruminant ungulates are regulated by conceptus interferon tau (IFNT). However, the effect of each IFNT isoform has not been carefully evaluated. In this study, the effects of 2 IFNT isoforms, paralogs found in utero, and interferon alpha (IFNA) on uterine epithelial and Mardin-Darby bovine kidney (MDBK) cells were evaluated. Expression vectors of the bovine interferon (bIFNT) genes bIFNT1, bIFNTc1, and bIFNA were constructed, and recombinant bIFNs (rbIFNs) were produced by 293 cells. Bovine uterine epithelial or MDBK cells were cultured in the presence or absence of increasing concentrations of each rbIFN for 24, 48, or 72 h. Transcript levels of the IFN-stimulated genes (ISGs) ISG12, ISG15, MX1, and MX2 were analyzed using quantitative reverse transcription-polymerase chain reaction. These messenger RNAs were up-regulated by rbIFN in a time- and concentration-dependent manner. In the epithelial cells, the ISG12 transcript level increased at 48 h after rbIFN treatment but slightly decreased at 72 h, whereas the transcript level of ISG15 increased at 24 h and was maintained through 72 h. Expressions of MX1 and MX2 increased at 72 h after rbIFN treatment. MX1 expression increased in all treatment groups, but MX2 increased only by bIFNTc1. In MDBK cells, the expression of ISG12 was increased by bIFNT1 and bIFNTc1 after 24 and 72 h; however, it was unchanged by rbIFNA. ISG15 increased following the same pattern as that seen in uterine epithelial cells, and MX1 showed a similar expression pattern. MX2 expression was increased by bIFNTc1 treatment in uterine epithelial cells, and its expression was increased by both bIFNT1 and bIFNTc1 in MDBK cells. These results show that epithelial and MDBK cell responses to IFNs differ, suggesting that IFNs possess common functions, but may have acquired different functions following gene duplication.

• 한국식품위생안전성학회:학술대회논문집
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• 한국식품위생안전성학회 2001년도 The Asia-Pacific Conference on Reproductive Biology and Environmental Sciences
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• pp.219-222
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• 2001

• Clinical and Experimental Reproductive Medicine
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• 제25권1호
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• pp.25-33
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• 1998

Cyclooxygenase (COX) is an enzyme involved in the conversion of arachidonic acid to prostaglandins (PGs), and exists in two forms, COX-1 and COX-2. COX has been reported to be involved in early implantation by secretion of PGs which causes permeability of vessels and reaction of decidual cells around the implantation site. Recently, in mice and sheep studies, COX-1 and COX-2 expression in the endometrium has been reported to be different according to implantation and stages of the estrous cycle, but expression of COX-1 and COX-2 in human endometrium during the menstrual cycle has not yet been established. The purpose of this study was to observe the variances of COX-1 and COX-2 expression by immunohistochemical staining in endometrial samples obtained from human hysterectomy specimens and biopsies of women of reproductive age according to different stages of the menstrual cycle. Also, we attempted to observe COX-1 and COX-2 expression in the epithelial and stromal cells of the endometrium obtained during the mid-secretory phase, which were cultured separately. COX-2 showed a cyclic pattern of expression according to the different stages of the menstrual cycle and was strongly expressed particularly at the mid-secretory phase which corresponds to the time of implantation. However, COX-1 tended to be increased in the early proliferative, and mid- and late secretory phases, but was also expressed in the whole menstrual cycle showing no particular pattern. In the separately cultured cells COX-1 was expressed in epithilial cells and COX-2 in the stromal cells. The above results suggest that since COX-2 is expressed at the same time as implantation and cultured cells display a specific secretory pattern, COX-2 has inductive endocrine enzyme properties and has an important effect on endometrial cells during implantation. Also, COX-2 expression in endometrial cells may be utilized as a useful marker of endometrial maturation.

• 한국가축번식학회지
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• 제17권2호
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• pp.141-148
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• 1993

This studies were carried out to investigate the effects of co-culture with cumulus cells, oviduct epithelial cells and uterine endometrium cells on the in-vitro fertilization and cleavage rate of bovine follicular oocytes. The ovaries were obtained from slaughtered Korean native cows. The follicular oocytes surrounded with cumulus cells were recovered by aspirating follicular fluids from the visible follicles of diameter 3~5 mm. The follicular oocytes were cultured in TCM-199 medium containing hormones and 10% FCS for 24~48 hrs in a incubator with 5% CO2 in air at 38.5$^{\circ}C$ and then matured oocytes were again cultured for 12~18 hrs with motile capacitated sperm by preincubation. The results obtained in these experiments were summarized as follows: 1. The in vitro maturatin and fertilization rate of bovine oocytes co-cultured with cumulus cells in TCM-199 medium were 64.0~74.1% and 40.0~58.6% respectively. And in-vitro fertilization rate of cumulus-enclosed oocytes(55.4%) were significantly(p<0.05) higher than cumulus-denuded oocytes(23.1%). 2. The in-vitro maturatin and fertilization rate of bovine oocytes co-cultured with 1$\times$104 cells/ml, 1$\times$106 cells/ml, 1$\times$108 cells/ml and 1$\times$1015 cells/ml oviduct epithelial cells in TCM-199 medium were 59.3% and 40.7%, 64.0% and 48.0%, 58.3% and 37.5%, 52.0% and 32.0%, respectively. 3. The in-vitro maturation and fertilization rate of bovine oocytes co-cultured with 1$\times$104 cells/ml, 1$\times$106 cells/ml, 1$\times$108 cells/ml and 1$\times$1015 cells/ml uterine endometrium cells in TCM-199 medium were 56.0% and 36.0%, 60.7% and 42.9%, 59.3% and 37.0%, 52.0% and 36.0%, respectively. 4. When the in-vitro fertilized oocytes were co-cultured with cumulus cells, oviduct epithelial cells and uterine endometrium cells, the development rate to be blastocyst was 12.2%, 15.6% and 11.7%, respectively and rates were higher than that of control, 2.1%(P<0.05).

• 대한수의학회지
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• 제36권1호
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• pp.101-108
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• 1996

This study was desinged to investigate the effect of estrogen(Est) on the proliferating of progesterone(Prog) target cells. The spayed 13 rats(Wistar, approximately 300gm) were randomly alloted into 3 groups. One group was the control group and another Prog-treated group was injected with 1mg of Prog/rat/day for 2 consecutive days, and Estand Prog-treated group was injected intramuscularly with $17{\beta}$-estradiol $20{\mu}g/rat/day$ for 3 consecutive days and then with Prog for 2 days as above from 4th day. Rats were administrated intraperitoneally with bromodeoxyuridinc(Brdur,0.2mg/BW once) befero 2 hours of exanguination. In gross finding, the groups with more level of dimension and weight on the uterus were ordered as Est- and Prog-treated group, Prog-treated group and control group. The investigation by immunohistochemical methods using paraffin sections of the uteri was performed by using anti-Brdur antibody for labeling proliferating cells of Prog target cells. The groups with higher labeling index(LI) were ordered as Prog-treated grop, Est- and Prog- treated group and control group. The number of proliferating cells from Prog target cells in the rats were rather deceased by Prog injection following Est injection than prog injection only. The cell types with higher LI in the wall layers of all 3 groups were ordered as endometrial stromal cells, glandular epithelial cells, luminal epithelial cells, myometrial muscle cells and serosa methodelial cells, and the region with highest LI was functional zone of the endometrium and the region with lower LI was muscular layer and then those with lowest LI was serosa and also the considerable different LI from individual rat were observed.

• Clinical and Experimental Reproductive Medicine
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• 제21권3호
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• pp.315-323
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• 1994

Mammalian oviductal epithelial cells have been known to improve in vitro fertilization and embryonic development. Recently, co-cultured human embryos with the epithelial cells in human genital tract has been reported to improve the pregnancy rate. The purpose of the study was to investigate the effects of the epithelial cells of human genital tract on the development of mouse early embryos and human fertilized oocytes. The epithelial cells of human genital tract were collected from the fallopian tubes which were obtained during hysterectomy in fertile women and from the endometrium during endometrium biopsy. Collected human ampullary cells(HACs) and endometrial cells(HECs) were cultured for 10 days to establish primary monolayer. Second passaged HACs and HECs were obtained by trypsinization were cryopreserved in PBS with 1.5 M DMSO for later use. To investigate the effect when co-cultured with HACs and HECs, we tried to apply strict quality control on mouse embryo, from two cell to blastocyst prior to human trial. The results of quality control were as follows; In Group I (Ham's F10 with 10% FCS), Group IT (co-cultured with HACs) and Group ill (co-cultured with HECs), developmental rates to blastocyst were 63.3%(253/400), 76.0%(304/ 400),74.0%(296/400), respectively. Hatching rates were 36.8%(147/400), 41.80/0(167/400), 38.0%(152/400), respectively(p<0.05). To perform the human IVF, cryopreserved HACs were thawed at 37$^{\circ}C$ waterbath, seeded on the well dish and cultured for 48 hI'S. The pronuclear stage embryos were transferred to the seeded well dish. After 24 hRS, co-cultured embryos were examined and transferred to patient's uterus. The results of human IVF when co-cultured with HACs were that fertilization and developmental rates were 61.8% (256/414), 95.3% (244/256) as compared with 57.2% (279/488) and 94.6%(264/279) in Ham's F10 supplemented with 10% FCS(control). However, 62.9% (161/256) of co-cultured human embryos showed good embryos(no or slight fragmentation) as compared with 53.8 % (150/279) in control(p < 0.05). Pregnancy rate was 40.0% (12/30) when co-cultured with HACs whereas 30.6%(11/36) in control. In conclusions, co-culture system using HACs and HECs improved the developmental and hatching rates of mouse embryo. Also, in human IVF system when co-cultured with HACs, it improved both the quality of human embryos and the pregnancy rate.

• Clinical and Experimental Reproductive Medicine
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• 제37권3호
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• pp.207-216
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• 2010

목 적: Small interfering RNA (siRNA)를 이용하여 homeobox (HOXA) 10 유전자의 발현이 억제된 일차배양 자궁내막 세포를 이용하여 자궁내막 탈락막화 (decidualization)에 HOXA유전자를 포함한 세포 내 신호전달기전을 분석하고자 하였다. 연구방법: 본원 산부인과에서 자궁내막 질환 이외의 이유로 전자궁 적출술을 받은 환자의 자궁내막 조직을 채취한다. $37^{\circ}C$에서 20분간 Trypsin-EDTA를 처리하여 단일세포로 분리한 후 10% fetal bovine serum이 첨가된 DMEM/F12 배지를 이용하여 24시간 동안 $37^{\circ}C$ 5% $CO_2$ 배양기 안에서 배양한다. 배양된 자궁내막 세포를 HOXA10 siRNA로 첨가한 후 TGF-${\beta}1$을 10 ng/mL 농도로 48시간 첨가하여 탈락막화를 유도한다. 배양된 자궁내막 세포에서 reverse transcription polymerase chain reaction을 이용하여 HOXA10, prolactin, cyclooxygenase (COX)-2, peroxisome proliferator-activated receptor (PPAR)-$\gamma$ 및 wingless-type MMTV integration site family (Wnt)의 발현을 관찰하였다. 결 과: HOXA10의 경우 transforming growth factor (TGF)-${\bata}1$과 HOXA10 siRNA를 처리하지 않은 대조군에 비하여 TGF-${\beta}1$을 처리한 군에서 약 1.8배 가량 발현양의 증가를 보였다. 자궁내막 탈락막 표지인자로 알려져 있는 prolactin의 경우 TGF-${\beta}1$을 처리한 경우 대조군에 비하여 유의한 발현의 증가를 보였으며 HOXA10 siRNA를 처리한 군에 있어서는 TGF-${\beta}1$을 첨가하더라도 prolactin mRNA의 발현양의 증가를 관찰할 수 없었다. 또한 자궁내막 세포의 분화인자로 알려져 있는 COX-2의 발현 역시 HOXA10 siRNA를 처리한 군에 있어서 mRNA 발현양이 유의하게 감소하였으며 TGF-${\beta}1$을 처리하여도 발현의 증가를 관찰할 수 없었다. Wnt4의 경우 HOXA10 siRNA를 이용하여 HOXA10의 발현을 억제한 경우 대조군에 비하여 유의하게 mRNA의 발현양이 감소하였으며 이러한 발현양의 감소는 TGF-${\beta}1$을 처리하여도 증가됨을 관찰할 수 없었다. PPAR$\gamma$의 발현은 HOXA10 siRNA의 처리와 관계없이 TGF-${\beta}1$에 의하여 감소하는 것을 관찰할 수 있었다. 결 론: Progesterone에 의하여 자궁내막 상피세포에서 분비되는 것으로 알려져 있는 TGF-${\beta}1$에 의한 자궁내막 기질세포의 분화 (탈락막화)는 HOXA10 및 Wnt에 의하여 조절되는 것으로 생각된다.

• Clinical and Experimental Reproductive Medicine
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• 제27권3호
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• pp.245-251
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• 2000

Objective: To investigate the distribution of BCL-2, BAX proteins and DNA fragmented cells in the normal human endometrium during at each menstrual cycle in order to find out whether apoptosis regulates cyclic endometrial change. Methods: Normal endometrial tissues were obtained from 40 patients, $32{\sim}45$ year of age, all with regular menstrual cycle, who were undergoing abdominal hysterectomy for myoma of uterus or cervical intraepithelial neoplasia for the period from 1992 through 1997. Immunohistochemical staining was used to determine the expression of BCL-2 and BAX protein with paraffin-embedded tissues. Results: BCL-2 was expressed on the glandular epithelial cells and stromal cells during the proliferative phase. The intensity of BCL-2 was increased predominantly on the basal layer than the functional layer in late proliferative phase. However, BCL-2 immunoreactivity was decreased in the secretory phase. BAX was expressed predominantly during the secretory phase. The intesity was increased in late secretory phase rather than early secretory phase. DNA fragmented cells were detected in a few cells at each phase. However, it was increased during the late secretory phase. Conclusion: Apoptosis-related genes, BCL-2 and BAX, may play a role in the regulation of cyclic endometrial change.

• Molecules and Cells
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• 제22권3호
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• pp.262-268
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• 2006

During endometrial differentiation the extracellular matrix (ECM) changes dramatically to prepare for implantation of the embryo. However, the genes regulating the ECM build-up in the uterine endometrium during early pregnancy are not well known. Using the PCR-select cDNA subtraction method, dermatopontin was identified in the uterus of a pregnant mouse on day 4 of gestation. Dermatopontin mRNA increased dramatically on day 3, and was at its highest level at the time of implantation. Administration of RU 486 significantly inhibited mRNA expression by day 4 of gestation, but ICI 182,780 did not. Progesterone markedly induced dermatopontin expression in ovariectomized uteri within 4 h of administration, whereas estrogen had little effect. In silico analysis revealed progesterone receptor binding sites in the dermatopontin promoter region. Decidualization did not induce expression of dermatopontin; instead dermatopontin mRNA became strongly localized at the interimplantation site. In situ hybridization revealed that expression gradually decreased in the luminal epithelial cells as pregnancy progressed, whereas it increased in the stromal cells. The pattern of localization and the changes of intensity of dermatopontin mRNA coincided with those of collagen. Collectively, these results strongly suggest that dermatopontin expression is steroid-dependent. They also suggest that, at the time of implantation, dermatopontin expression is primarily regulated spatio-temporally by progesterone via progesterone receptors, and is modulated by the decidual response during implantation. Dermatopontin may be one of the regulators used to remodel the uterine ECM for pregnancy.

• Molecules and Cells
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• 제42권1호
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• pp.8-16
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• 2019

Mutations in the ${\beta}-catenin$ gene (CTNNB1) have been implicated in the pathogenesis of some cancers. The recent development of cancer genome databases has facilitated comprehensive and focused analyses on the mutation status of cancer-related genes. We have used these databases to analyze the CTNNB1 mutations assembled from different tumor types. High incidences of CTNNB1 mutations were detected in endometrial, liver, and colorectal cancers. This finding agrees with the oncogenic role of aberrantly activated ${\beta}-catenin$ in epithelial cells. Elevated frequencies of missense mutations were found in the exon 3 of CTNNB1, which is responsible for encoding the regulatory amino acids at the N-terminal region of the protein. In the case of metastatic colorectal cancers, in-frame deletions were revealed in the region spanning exon 3. Thus, exon 3 of CTNNB1 can be considered to be a mutation hotspot in these cancers. Since the N-terminal region of the ${\beta}-catenin$ protein forms a flexible structure, many questions arise regarding the structural and functional impacts of hotspot mutations. Clinical identification of hotspot mutations could provide the mechanistic basis for an oncogenic role of mutant ${\beta}-catenin$ proteins in cancer cells. Furthermore, a systematic understanding of tumor-driving hotspot mutations could open new avenues for precision oncology.