• Reproductive and Developmental Biology
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• 제29권2호
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• pp.83-91
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• 2005

Somatic cell nuclear transfer in cattle has limited efficiency in terms of production of live offspring due to high incidence of fetal failure after embryo transfer to recipients. Such low efficiency of cloning could possibly arise from abnormal and poorly developed placenta. In the present study the placental proteome in late pregnancy established from in vitro fertilization (IVF) and nuclear transfer (NT) was analysed. Proteome alternation was tested using two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI- TOF). Comparing placenta from NT embryos to those from IVF counterparts, significant changes in expression level were found in 18 proteins. Of these proteins 12 were not expressed in NT placenta but expressed in IVF counterpart, whereas the expression of the other 6 proteins was limited only in NT placenta. Among these proteins, cytokeratin 8 and vimentin are considered to be involved in regulation of post-implantation development. In particular, cytokeratin 8 and vimentin may be used as makers for placental development during pregnancy because their expression levels changed considerably in NT placental tissue compared with its IVF counterpart. Data from 2-DE suggest that protein expression was disorientated in late pregnancy from NT, but this distortion was eliminated with progression of pregnancy. These findings demonstrate abnormal placental development during late pregnancy from NT and suggest that alterations of specific placental protein expression may be involved in abnormal function of placenta.

• Reproductive and Developmental Biology
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• 제34권1호
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• pp.33-40
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• 2010

Equine chorionic gonadotropin (eCG) is a heavily glycosylated glycoprotein composed of non-covalently linked $\alpha$- and $\beta$-subunits. To study the function and signal transduction of tethered recombinant-eCG (rec-eCG), a single chain eCG molecule was constructed, and the rec-eCG protein was prepared. In this study, we constructed 5 mutants (${\Delta}1$, ${\Delta}2$, ${\Delta}3$, ${\Delta}4$, and ${\Delta}5$) of rec-eCG using data about known glycoprotein hormones to analyze the role of specific follicle stimulating homone (FSH)-like activity. Three amino acids of certain specific sites were replaced with alanine. The expression vectors were transfected into CHO cells and subjected to G418 selection for 2~3 weeks. The media were collected and the quantity of secreted tethered rec-eCGs was quantified by ELISA. The LH- and FSH-like activities were assayed in terms of cAMP production by rat LH/CG and rat FSH receptors. Then, the metabolic clearance rate analyzed by the injection of rec-eCG (5 IU) into the tail vein was analyzed. The mutant eCGs (${\Delta}l$, ${\Delta}4$, and ${\Delta}5$) were transcripted, but not translated into proteins. Rec-eCG A2 was secreted in much lower amounts than the wild type. Only the rec-eCG ${\Delta}3$ ($\beta$-subunit: $Gln^{94}-Ile^{95}-Lys^{96}{\rightarrow}Ala^{94}-Ala^{95}-Ala^{96}$) was efficiently secreted. Although activity is low, its LH-like activity was similar to that of tethered $eCG{\beta\alpha}$. However, the FSH-like activity of rec-$eCG{\beta\alpha\Delta}3$ was completely flat. The result of the analysis of the metabolic clearance rate shoed the persistence of the mutant in the blood until 4 hours after the injection. After then, it almost disappeared at 8 hours. Taken together, these data suggest that 94~96 amino acid sequences in eCG $\beta$-subunit appear to be of utmost importance for signal transduction of the FSH receptor.

• 생명과학회지
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• 제19권9호
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• pp.1309-1313
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• 2009

내분비교란물질은 일상생활에서 쉽게 발견되는 화학물질이다. Bisphenol A, DDT, benzophenone, phenylphenol 등과 같은 화학물질들은 플라스틱 식품용기나 살충제 등을 통해서 체내에 흡수된다. 내분비교란물질은 내분비계의 정상적인 기능을 방해하기 때문에 인체에 해로우며, 매우 독성이 강한 것으로 알려져 있다. 이러한 내분비교란물질은 심혈관계, 생식계, 신경계뿐만 아니라 인체의 여러 기관에 심각한 타격을 주는 것으로 알려져 있다. 따라서 본 연구에서는 benzophenone과 같은 내분비교란물질이 계배의 발생과정에 영향을 미치고, 계배의 기형을 유도하는지를 알아보고자 하였다. Benzophenone $1{\mu}g$/egg에서 $500{\mu}g$/egg까지를 투여한 후 부화기에서 발생을 유도한 후 각각 6. 9. 12. 15. 18. 21일째마다 확인하였다. 체장의 경우에는 중용량($40{\mu}g$/egg $\sim$ $60{\mu}g$/egg)에서 감소가 시작되는 것을 확인할 수 있었다. 고용량의 실험군($80{\mu}g$/egg $\sim$ $500{\mu}g$/egg)에서는 높은 사망률을 보였으며, 부리 이상이나 안구 기형과 같은 기형의 징후를 발견할 수 있었다. 결론적으로 benzophenone은 계배의 기형을 유도하며, 발생단계에서 심각한 영향을 미치는 것을 확인할 수 있었다.

• 한국응용곤충학회지
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• 제42권4호
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• pp.367-381
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• 2003

곤충의 분산다형성이란 비상능력과 관련된 다형성으로 그 구체적 내용으로는 시다형성, 비상근다형성 그리고 비상행동변이성 등과 이들과는 별개로 개체군 밀도의존적인 상변이성이 있다. 분산다형성은 시간적으로나 공간적으로 이질적인 서식처 환경에 대응하기 위하여 이동형인 “유시형이나 장시형”과 정주형인 “무시형이나 단시형”을 생활사에 적절히 짜넣은 적응적 형질이다. 점변태곤충류에서는 유충과 성충의 생태학적지위가 중복되여 있어 유충과 성충이 생활공간과 그 밖의 요구조건을 달리하고 있는 완전변태류나 반변태류에 비하여 분산다형성의 예가 대단히 많다. 무시형 또는 단시형곤충은 같은 종의 유시형 또는 장시형곤충에 비하면 초산연령이 빠르고 총산란수도 많은 것이 보통이여서 자연증가율(r)이 크다. 단시형과 관련된 환경요인으로는 서식처의 시간적 영속성이나 공간적 이질성, 먹이조건, 개체군밀도, 온도, 일장 기타 여러 가지가 알려지고 있다 서식처의 환경조건에 대한 분산다형성발현상은 종에 따라 다를 뿐 아니라 암수간에도 차가 있고 같은 종에서도 계통간에 차가 있는 극히 탄력적인 현상이다. 분산다형성의 문제는 생리학, 유전학 그리고 생태학등에 걸친 폭넓은 학문분야로 특히 생태유전학이나 정량유전학분야치 연구는 분산다형성의 유전적본질 구명에 중요하다 하겠다.

• 대한의생명과학회지
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• 제10권4호
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• pp.391-395
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• 2004

The demand for the production of gene-defective mice from embryonic stem (ES) cells is increasing to clarify decisive gene function in vivo. Although blastocyst injection is widely used to generate ES cell-mediated knockout mice, coculture method has been alternatively used because of several advantages, such as low cost and simple procedure. Thus, this experiment was designed to demonstrate the feasibility of the coculture method using J1 ES cells, which are known to be efficient for blastocyst injection. Eight-cell embryos were harvested from 2.5 days post-coitum (dpc), denuded with acid tyrode's solution, and transferred onto trypsinized J1 ES cells. Aggregation was carried out following two typical methods, which are simple coculture method and aggregation in groove prepared by aggregation needle. Successfully aggregated-embryos were developed to blastocysts for 24 h and transferred into uterus of pseudo-pregnant foster mother. Chimeric offspring was judged by coat pigmentation. In this study, we could obtain chimeric mice from all the two aggregation methods, but the chimera production efficiencies in coculture using groove were three times higher at least than those in the other group. In conclusion, these observations suggest that coculture method should be available for production of knockout mice from J1 ES cells. Presently, the germ-line transmission rates of the chimeras produced from the two methods are under investigation.

• 한국발생생물학회지:발생과생식
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• 제2권1호
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• pp.21-27
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• 1998

본 연구에서는 흰쥐 자궁에서 pituitary adenylate cyclase-activating polypeptide (PACAP)과 그 수용체 유전자들이 발현되는가와 각각 어떠한 유형의 transcript들이 발현되는가를 조사하였고, 이를 위해 역전사 중합효소 연쇄반응 (RT-PCR)을 시행하였다. 시상하부와 정소에서 공통적으로 존재함이 알려진 PACAP common exon 부위에 해당되는 primer를 사용하여 PCR을 시행한 결과 자궁을 포함한 모든 조직에서 에상대로 297 bp의 band가 확인되었다. 흰쥐 자궁에서 발현되는 PACAP mRNA에 정소 특이적인 exon 1이 포함되는가를 조사하기 위해 정소 특이적인 primer를 사용하여 PCR을 시행한 결과, 정소에서만 예상대로 586 bp의 band가 검출되었고 자궁을 포함한 다른 조직에서는 발견되지 않았다. 흰쥐의 PACAP 수용체 PCR에서는 자궁에서 923 bp와 839 bp크기의 band를 확인할 수 있었으며, 이는 알려진 PACAP type I수용체의 splicing variant 들 가운데 hip-hop (923bp)과 hip- 또는 nop-type (839bp)의 예상 크기와 일치하였다. 인위적으로 성적인 성숙을 유도한 PMSG 주사모델하에서 자궁내 PACAP transcript 수준은 주사 24 시간 후 실험군에서 증가하여 생식주기상 proestus 시기에 해당되는 주사 48 시간까지 증가하였다가 72 시간후에는 control 보다 낮은 수준을 보였는데, 이는 자궁의 PACAP 유전자발현이 생리적으로 조절됨을 나타내는 것이다. 본 연구는 흰쥐의 자궁에서 PACAP과 그 수용체 isoform들의 유전자가 발현됨을 최초로 보고한 것으로 자궁 자체에서 발현되는 PACAP이 autocrine 또는 paracrine하게 자궁의 생리 및 기능 조절을 담당함을 시사한다.

• Asian-Australasian Journal of Animal Sciences
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• 제30권5호
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• pp.643-652
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• 2017

Objective: Prostaglandins (PGs) function in various reproductive processes, including luteolysis, maternal pregnancy recognition, conceptus development, and parturition. Our earlier study has shown that PG transporters ATP-binding cassette, subfamily C, member 4 (ABCC4) and solute carrier organic anion transporter family, member 2A1 (SLCO2A1) are expressed in the uterine endometrium in pigs. Since several other PG transporters such as ABCC1, ABCC9, SLCO4C1, and SLCO5A1 are known to be present in the uterine endometrium, this study investigated the expression of these PG transporters in the porcine uterine endometrium and placenta. Methods: Uterine endometrial tissues were obtained from gilts on day (D) 12 and D15 of the estrous cycle and days 12, 15, 30, 60, 90, and 114 of pregnancy. Results: ABCC1, ABCC9, SLCO4C1, and SLCO5A1 mRNAs were expressed in the uterine endometrium, and levels of expression changed during the estrous cycle and pregnancy. Expression of ABCC1 and ABCC9 mRNAs was localized mainly to luminal and glandular epithelial cells in the uterine endometrium, and chorionic epithelial cells during pregnancy. Conceptuses during early pregnancy and chorioallantoic tissues from mid to late pregnancy also expressed these PG transporters. $Estradiol-17{\beta}$ increased the expression of ABCC1 and SLCO5A1, but not ABCC9 and SLCO4C1 mRNAs and increasing doses of $interleukin-1{\beta}$ induced the expression of ABCC9, SLCO4C1, and SLCO5A1 mRNAs in endometrial explant tissues. Conclusion: These data showed that several PG transporters such as ABCC1, ABCC9, SLCO4C1, and SLCO5A1 were expressed at the maternal-conceptus interface, suggesting that these PG transporters may play an important role in the establishment and maintenance of pregnancy by regulating PG transport in the uterine endometrium and placenta in pigs.

• Reproductive and Developmental Biology
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• 제35권2호
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• pp.151-157
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• 2011

Clinical arthritis is typically divided into rheumatoid arthritis (RA) and osteoarthritis (OA). Arthritis-induced muscle weakness is a major problem in aged people, leading to a disturbance of balance during the gait cycle and frequent falls. The purposes of the present study were to confirm fiber type-dependent expression of muscle atrophy markers induced by arthritis and to identify the relationship between clinical signs and expression of muscle atrophy markers. Mice were divided into four experimental groups as follows: (1) negative control (normal), (2) positive control (CFA+acetic acid), (3) RA group (CFA+acetic acid+type II collagen), and (4) aging-induced OA group. DBQA/1J mice (8 weeks of age) were injected with collagen (50 ${\mu}g/kg$), and physiological (body weight) and pathological (arthritis score and paw thickness) parameters were measured once per week. The gastrocnemius muscle from animals in each group was removed, and the expression of muscle atrophy markers (MAFbx and MuRF1) and myosin heavy chain isoforms were analyzed by reverse transcription-polymerase chain reaction. No significant change in body weight occurred between control groups and collagen-induced RA mice at week 10. However, bovine type II collagen induced a dramatic increase in clinical score or paw thickness at week 10 (p<0.01). Concomitantly, the expression of the muscle atrophy marker MAFbx was upregulated in the RA and OA groups (p<0.01). A dramatic reduction in myosin heavy chain (MHC)-$I{\beta}$ was seen in the gastrocnemius muscles from RA and OA mice, while only a slight decrease in MHC-IIb was seen. These results suggest that muscle atrophy gene expression occurred in a fiber type-specific manner in both RA- and OA-induced mice. The present study suggests evidence regarding why different therapeutic interventions are required between RA and OA.

• Reproductive and Developmental Biology
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• 제35권4호
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• pp.479-483
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• 2011

The objective of this study was to determine the effects of E. coli isolated from porcine semen on sperm viability, motility, and semen pH. Semen samples were prepared using commercial extender, $Seminark^{Pro}$ (Noahbio Tech, Korea) that did not contain antibiotics. And 4 different levels of E. coli were artificially innoculated to semen with following concentrations; 4,000 of sperms with 1 of E. coli (T1), 400 with 1 (T2), 40 with 1 (T3), and 4 with 1 (T4). Semen samples were preserved at $17^{\circ}C$ for 5 days in semen storage box until analyzed by flowcytometer. Aliquots were subjected to measure the sperm viability (Live/$Dead^{(R)}$ stain), motility (mitochondrial function), and semen acidity (pH) from day 0 (day of semen collection) to day 5. Sperm motility and viability were significantly decreased (p<0.05) on day 0 (4 hrs after preservation at $17^{\circ}C$) in T3 and T4 compared to control groups and were significantly decreased (p<0.05) in all groups from day 3. Sample pH was acidic in T3 (6.90~6.86) and T4 (6.86~6.65) from day 3 to day 5 (p<0.05). On the other hand, sample pH was maintained 7.0~7.1 in control, T1, and T2 during the experimental period. Sperm motility and viability were significantly decreased from day 0 to day 5 compared to control in samples contaminated with E. coli above a value of 40:1 ($20{\times}10^6$ sperm cells/ml : $5{\times}10^5$ cfu/ml). Even on day 1 in T4 and on day 3 in T3, semen pH was acidic probably due to the acidification of dead spermatozoa. These results suggest that E. coli contamination has a concentration-dependent detrimental effect on extended porcine semen quality.

• 생명과학회지
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• 제23권8호
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• pp.1064-1072
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• 2013

일주기 리듬은 모든 살아있는 유기체의 생리현상을 지배하는 호르몬의 변화에 의해서 조절된다. 포유동물에서 송과체의 주된 기능은 시상 하부 시교차 상핵에서 발생되는 일주기 리듬을 주로 어두울 때 증가하는 순환성 멜라토닌의 리듬 신호로 변화시키는 것이다. 송과체는 직접적인 광감도는 없지만, 망막신경절세포로 하부조직을 포함하는 멀티 시냅스 경로를 통하여 빛에 반응한다. 주기적인 리듬 조절은 주위환경의 빛과 멜라토닌 생성의 리듬조절 효소인 arylalkylamine-N-acetyltransferase (AANAT)의 발현과 긴밀한 관계를 통해 이루어진다. 이전 실험에서 AANAT 단백질이 어두울 때의 발현이 전사 조절, 전사 후 조절, 번역 후 조절 메커니즘으로 설명되었다. AANAT 단백질 발현에 관한 분자적 기전은 멜라토닌의 일주기 리듬에 대한 새로운 견해를 제공한다. 광범위한 동물 연구에서 많은 포유류의 계절 리듬을 위한 송과체 멜라토닌은 일주기 리듬의 조절과 수면 조절에 관련이 있는 것으로 알려졌다. 이것은 시차증이나 교대 근무 수면 장애와 같은 일주기 리듬 수면 장애를 치료하는 데 있어서 가치가 있다. 또한 멜라토닌은 다른 영역에도 영향을 미치는데 특히 몸의 생리적 기능을 조절하는데 영향을 미친다. 게다가 정신의학적 질환뿐 만 아니라 생식기 질환, 심혈관 질환, 면역 조절 질환도 이 호르몬에 의해 영향을 받는 것으로 밝혀졌다.