• 한국가축번식학회지
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• 제27권1호
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• pp.53-59
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• 2003

본 연구에서는 한우 태아의 시기별로 35일령, 50일령, 70일령 및 90일령의 fetal fibroblast cell line을 생산하였고, bovine-specific primer와 Y chromosome-specific primer를 이용하여 PCR에 의해 성을 판별하여 각각 암수 2 line의 한우 fetal fibroblast cell line을 확립하였다. 이들 cell line을 계대배양하여 passage number가 10 이상에서 염색체 분석을 실시하였는데 모두에서 80%이상의 세포가 60개의 정상 염색체수의 나타내어 계대배양이 karyotype에 영향을 미치지 않는 것으로 나타났다. Serum starvation과 confluent 배양 방법을 이용하여 Go 상태로 유도되었는지 확인하기 위해 PCNA antibody를 이용하여 Western blotting 분석을 실시하였는데 PCNA 발현이 현저히 감소되는 것을 확인할 수 있었고, 다시 정상 medium으로 환원시켰을 때 세포분열이 재개되어 Go상태로 유도되었음을 확인할 수 있었다. 또한 serum stravation 방법이 conflent한 배양방법보다 PCNA 발현양이 적은 것으로 나타나 좀더 효율적인 Go 상태 세포 주기 조절방법으로 판명되었다.

• Archives of Plastic Surgery
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• 제32권3호
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• pp.347-356
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• 2005

The wound healing process in fetus is quite different form that of adult. Regeneration plays an important role and scarless wound healing is possible in early gestational fetal period. Recently, the various effects of the hypoxia and reoxygenation in the wound healing process have been investigated by many researchers. The hypoxic state is known to alter protein synthesis and gene expression of TGF-${\beta}$, VEGF. The authors hypothesize there may be differences between fetal and adult fibroblast and this difference may play a possible role in the mechanism of scarless fetal wound healing. In this study, we investigated the growth of fibroblast, the amount of collagen deposition, the amount of protein synthesis and gene expression in TGF-${\beta}$(transforming growth factor-${\beta}$), VEGF(vascular endothelial growth factor) under the various hypoxic and reoxygenation conditions. Through these processes, we tried to determine the relationships between scarless fetal wound healing and hypoxic condition. In control group, fetal and adult fibroblasts were cultured under normoxic condition. The experimental groups were allocated into four different groups. The differences in TGF-beta, VEGF under 24, 48, 72 hours were statistically investigated. Compared to adult fibroblast group, there was a statistically significant increase (p<0.01) in the rates of protein synthesis in TGF-beta and VEGF of fetal fibroblast. In this study, these results may reflect the possibility that fetal fibroblast are more susceptible to change in oxygen and has a superior rate of angiogenesis through increased VEGF expression. The possible superiority of angiogenesis in fetal fibroblast may play an important role in scarless wound healing.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2004년도 제4회 발생공학 국제심포지움 및 학술대회
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• pp.51-52
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• 2004

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2004년도 제4회 발생공학 국제심포지움 및 학술대회
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• pp.26-31
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• 2004

This work was undertaken in order to study the developmental competence of nuclear transfer cat embryo with fetal fibroblast and adult skin fibroblast as donor nuclei. Oocytes wererecovered by mincing the ovaries in Hepes-buffered TCM199 and selected the cumulus oocyte complexes (COCs) with compact cumulus cell mass and dark. Homogenous ooplasm were cultured for maturation in TCM199 + 10% fetal bovine serum (FBS) for 12 hours and used as a source of recipient cytoplast for exogenous somatic nuclei. In Experiment 1, we evaluated the effect donor cell types on the reconstruction and development of cloned embryos. Fusion, first cleavage and blastocyst developmental rate was not different between fetal fibroblast and adult skin cell (71.2 vs. 66.8; 71.0 vs. 57.6; 4.0 vs. 6.1 %, P<0.05). In Experiment 2, cloned embryos were surgically transferred into the oviducts of recipient queens. One of seven recipient queens was delivered naturally 2healthy cloned cats and 1 stillborn from fetal fibroblast cell of male origin after 65 days embryo transfer. One of three recipient queens was delivered naturally 1 healthy cloned cat from adult skin cell of female after 65 days embryo transfer. The cloned cats showed genotypes identical to the donor cell lines, indicating that adult somatic cells can be used for feline cloning.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2001년도 발생공학 국제심포지움 및 학술대회 발표자료집
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• pp.60-61
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• 2001

To produce reconstituted rabbit embryos with fetal fibroblasts, the present study was evaluated the efficiencies of the activation conditions as assessments of subsequent development and chromosome in the embryos. New Zealand White rabbits were used throughout the study. Fetal fibroblasts collected from 22-d of fetuses were cultured in DMEM＋10% FBS in 5% CO₂ in air. The culture was maintained for 10 passages. In every passage half of cell suspension were kept in frozen. (omitted)

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2002년도 춘계학술발표대회 발표논문초록집
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• pp.37-37
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• 2002

The success of embryo cloning depends on numerous factors; interaction between recipient ooplasm and donor nucleus, nuclear reprogramming, oocyte activation, and donor cell cycle and type. In this study, the cell cycle and apoptosis of bovine fetal fibroblast as a donor cell for embryo cloning were evaluated following different activation treatments. (omitted)

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2001년도 발생공학 국제심포지움 및 학술대회 발표자료집
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• pp.60-61
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• 2001

• Reproductive and Developmental Biology
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• 제32권4호
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• pp.249-253
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• 2008

Interspecies somatic cell nuclear transfer (iSCNT) is a valuable tool for studying the interactions between an oocyte and somatic nucleus. The object of this study was to investigate the developmental competence of in vitro-matured porcine oocytes after transfer of the somatic cell nuclei of 2 different species (goat and rabbit). Porcine cumulus oocytes were obtained from the follicles of ovaries and matured in TCM-199. The reconstructed embryos were electrically fused with 2 DC pulses of 1.1kV/cm for $30{\mu}s$ 0.3M mannitol medium. The activated cloned embryos were cultured in porcine zygote medium-3 (PZM-3), mSOF or RDH medium for 7 days. The blastocyst formation rate of the embryos reconstructed from goat or rabbit fetal fibroblasts was significantly lower than that of the embryos reconstructed from porcine fetal fibroblast cells. However, a significantly higher number of embryos reconstructed from goat or rabbit fetal fibroblasts cultured in mSOF or RDH, respectively, developed to the morular stage than those cultured in PZM-3. These results suggest that goat and bovine fetal fibroblasts were less efficacious than porcine-porcine cloned embryos and that culture condition could be an important factor in iSCNT. The lower developmental potential of goat-porcine and porcine-bovine cloned embryos may be due to incompatibility between the porcine oocyte cytoplasm and goat and bovine somatic nuclei.

• Reproductive and Developmental Biology
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• 제33권2호
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• pp.113-117
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• 2009

Animals produced by somatic cell nuclear transfer (SCNT) using genetically modified cells are almost always transgenic, implying that this method is more efficient than the traditional pronuclear microinjection method. Most somatic cells for SCNT in animals are fetus-derived primary cells and successful gene integration in somatic cells will depend on transfection condition. The objective of this study is to evaluate the efficiency of electroporation (Microporator) and liposome reagents (F-6, F-HD, W-EX, W-Q, W-M) for tissue-type plasminogen activator (tPA) gene transfection and to estimate the overall efficiency of transfection of Korean native pig fetal fibroblast cells (KNPFF). Electroporation showed significantly higher transfection efficiency than liposome reagents with regard to the transfection of in vitro cultures in the early stages of development (41.7% with Microporator vs. 18.3% with F-6, 20.0% with F-HD 18.5% with W-EX, 5.0% with W-M and 6.3% W-Q,). Colonies identified as tPA-positives were treated once more with G418 for 10 to 14 days and growing colonies were selected again. When the cells of newly selected colonies were subjected to single-cell PCR, reselection of colonies following second round of G418 selection increased the rate of transgene integration per each colony. These results suggest that transfection with electroporation is the most efficient and the second rounds of G418 selection may be an effective method for transfection of porcine fetal fibroblast cells.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2004년도 춘계학술발표대회
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• pp.220-220
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• 2004

The purpose of this study was to development of transgenic cow using the nuclear transfer. To secrete hFSH in urea, the vector was constructed with UPII promoter. The fetal fibroblast cells (KbFF) were constructed from pregnant day 45 male fetus. The hFSH genes were cotransfected with pcDNA3 (neo) vector to KbFF cells by electroporation. (omitted)