• Proceedings of the KSAR Conference
• /
• 2001.03a
• /
• pp.16-16
• /
• 2001

Apoptosis로 일컬어지는 예정된 세포사멸(programmed cell death)은 개별 세포의 입장에서는 곧바로 사멸을 의미하지만, 정상적인 고등 생물의 입장에서는 개체의 발생과 분화하는데 프로그램된 과정이다. 자발적 세포사멸은 다른 조직에 비해 생식 조직인 난소나 정소에서 복잡한 apoptosis 기작들을 가지리라 사료된다. 본 연구는 Bcl-2 family중 apoptotic protein인 Bax에 대해 suppression하는 유전자를 yeast system을 활용하여 돼지 정소와 난소로부터 각각 cDNA library를 구축한 후 탐색하였다. 탐색에 활용된 cDNA library는 돼지의 정소와 난소로부터 mRNA를 분리하여 yeast vector인 pAD-GAL4-2.1에 구축하였고, 마우스 bax 유전자는 gal 1 promoter의 조절 하에 glucose 배지에서는 유도되지 않고, galactose 배지에서만 선택적으로 Bax를 발현할 수 있는 효모 vector(pL19-bax)를 구축하였다. Bax에 의한 apoptosis suppressor를 탐색하기 위해 우선 효모 W303에 pL19-bax를 transform하여 glucose 배지에서 Bax의 발현을 억제하였다. pL19-bax를 가진 효모에 정소와 난소로부터 구축된 cDNA library를 transform 시키고, transform된 효모는 각각 Bax에 의한 toxicity를 저해하는 유전자를 찾기 위해 스크린되었다. 이러한 방법으로 정소 cDNA library 탐색에서는 5 $\times$ $10^{6}$ transformant중 39개, 난소cDNA library 탐색에서는 2 $\times$ $10^{6}$ transformant중 26개의 콜로니가 생존하였다. 이들 콜로니로부터 유전자를 분리하여 분석해 본 결과 여러 그룹으로 분류할 수 있었다. 각 그룹의 관련 유전자는 protein synthesis/degradation 12종, oxidation/reductation 5종, detoxin/ cell cycle promoter 3종, signal transduction/growth factor 5종, 그리고 알려지지 않은 유전자 9종이었다. 그 중, bax-toxicity inhibition에 강력한 survival phenotype을 가지는 유전자(pSEDL)를 동정하였다. 이것은 T3-4-1 콜로니로부터 분리하였는데 140개 아미노산으로 이루어진 인간 SEDL(GenBank, XM_013096) 유전자와 매우 유사한 homology를 가지며, bax와 관련된 기능은 밝혀져 있지 않다. 이외에도 분리된 유전자에는 NADH, thioreduction, 그리고 cytochrome oxidase와 같은 positive 유전자 군이 크로닝되어, Bax를 이용한 효모에서 apoptosis suppressor에 관련된 유전자를 손쉽게 스크린하는 것이 가능하고, 분리된 유전자의 기능을 예측할 수 있어 지금까지 보고된 유전자 크로닝법 보다는 강력한 수단으로 활용될 수 있다는 사실을 시사하였다. 그러나, ORF에 관계없이 Bax 발현에 저항하는 유전자군이 선발된다든지 하는 문제점은 금후 검토가 필요하리라 사료된다.

• Korean Journal of Veterinary Service
• /
• v.32 no.3
• /
• pp.189-200
• /
• 2009

The purpose of this study was to evaluate the effect of a subsequent infection of PCV2 on piglets with PEDV. In clinical signs, the signs observed in dual-infected with PEDV and PCV2 piglets and alone infected with PEDV piglets ranged from diarrhoea to vomiting and dehydration. Dual-infected piglets developed signs of anorexia, vomiting and watery diarrhoea within 12 hpi. Nevertheless alone -infected piglets caused pasty diarrhea at first. In mortality, dual infections showed 25%, but alone -infections showed 8.3%, respectively. In gross findings, piglets dual-infected with PEDV and PCV2 appeared the severe findings of congestion, distension of lumen, milder curdes of undigested milk in stomach than those of single-infected piglets. In histopathological findings, piglets of dual-infection group appeared the more severe findings of villous atrophy and fusion, congesion, exfoliation, vacuolation, squamation, loss of cilia and proliferation of crypt. Significant (P<0.05) decrease in VH:CD ratio in dually infected piglets compared to piglets from alone-PEDV infections. In immunohistochemical findings, strong hybridization signals in dual-infected piglets observed moderate to severe villous atrophy or vacuolation with positive cells arranged continuously over the villi. In the lumen, exfoliated enterocytes were strongly positive in dual-infected piglets. A number of PEDV-positive cells in dual-infected pigs were significantly higher than that in alone PEDV-infected piglets.

• Journal of the Korea Academia-Industrial cooperation Society
• /
• v.18 no.10
• /
• pp.551-558
• /
• 2017

To study and validate tissue-specific promoters and vectors, it is important to develop cell culture systems that retain the tissue and species specificity. Such systems are attractive alternatives to transgenic animal models. This study established a line of porcine mammary gland epithelial cells (PMECs) from a primary culture based on the cellular morphology and mRNA levels of porcine beta-casein (CSN2). The selected PMECs were stained with the cytokeratin antibody, and were shown to express milk protein genes (CSN2, lactoferrin, and whey acidic protein). In addition, to confirm the acini structure of PMEC932-7 in 3D culture, live cells were stained with SYTO-13 dye, which binds to nucleic acid. The acini of these PMECs on matrigel were formed by the aggregation of peripheral cells and featured a hollow lumens. The system was demonstrated by testing the effects of the culture conditions to cell culture including cell density and matrigel methods of the PMECs. These results suggest that PMECs possess the genetic and structural features of mammary epithelial cells.

• Journal of Biomedical Engineering Research
• /
• v.10 no.2
• /
• pp.165-172
• /
• 1989

In this paper, the temperature dependence of tissue is estimated by measuring the attenuation coefficient and the propagation velocity of ultrasonic reflection signal. And, on the basis of expeiments, the possibility of non- invasive temperature estimation is considered. Specimens in the experiment are acryl 1)late. muscle, fat and liver of pig. The temperature of specimen is controlled by water bath which is able to adjust temperature a quarter of a degree. Through the series of experiments, we conformed that the ultrasonic parameters have lin earity to a certain extents with the change of tissue's temperature. And we expect that noninvasive temperature estimation of tissue can be realized after several preconditions be satisfied with the standard experiment conditions and a great number of base data.

• Journal of the korean veterinary medical association
• /
• v.33 no.1
• /
• pp.22-28
• /
• 1997

12마리의 개들을 12주동안 6종류의 규정식으로 각각 급여시켰다. 규정식들은 단지 단백질원 즉 닭고기, 새끼양의 고기, 물고기, 쇠고기 및 콩이란 것에서만 차이가 있었다. 개들은 CBC 즉, 혈청화학프로필, 요분석, TSH반응시험 및 피부생검을 통하여 평가하였다. 피부생검은 조직학적으로 하는 것과 피부지방산 농도의 측정을 통한 평가, 두가지 방법으로 평가하였다. 평가된 지방산들은 리롤레산, 감마-리롤렌산, 알파-리롤렌산, 아라키돈산, 아이코사테트라에노이산 및 아이코사펜타에노이산 등이었다. 개들은 주관적으로 생검채취부위에서 털의 재성장과 비늘(피부의 얇은 조각)의 존재유무로 평가하였다. 결과에서 보면 각종의 규정식을 급여시킨 개들 사이에서 CBS, 조직적 소견 또는 피부지방산 수치에서 차이가 나타나지 않았다. 쇠고기 규정식을 급여시킨 12마리중 3마리의 개에서 고콜레스테롤 혈증이 있었고, 콩 규정식을 급여시킨 12마리중 9마리의 개에서 알카리성 오줌과 돼지고기를 급여시킨 12마리중 4마리가 비늘이 주관적 증가가 있었고, 털의 재성장이 감소된 것으로 나타났다.

• Journal of Chest Surgery
• /
• v.37 no.4
• /
• pp.297-306
• /
• 2004

Current artificial heart valves have several disadvantages, such as thromboembolism, limited durability, infection, and inability to grow. The solution to these problems would be to develop a tissue-engineered heart valves containing autologous cells. The aim of this study was to optimize the protocol to obtain a porcine acellular matrix and seed goat autologous endothelial cells on it, and to evaluate the biological responses of xenograft and xeno-autograft heart valves in goats. Material and Method: Fresh porcine pulmonic valves were treated with one method among 3 representative decellularization protocols (Triton-X, freeze-thawing, and NaCl-SDS). Goat venous endothelial cells were isolated and seeded onto the acellularized xenograft leaflets. Microscopic examinations were done to select the most effective method of decellularizing xenogeneic cells and seeding autologous endothelial cells. Two pulmonic valve leaflets of. 6 goats were replaced by acellularized porcine leaflets with or without seeding autologous endothelial cells while on cardiopulmonary bypass. Goats were sacrificed electively at 6 hours, 1 day, 1 week, 1 month, 3 months, and 6. months after operation. Morphologic examinations were done to see the biological responses of replaced valve leaflets. Result: The microscopic examinations showed that porcine cells were almost completely removed in the leaflets treated with NaCl-SDS. The seeded endothelial cells were more evenly preserved in NaCl-SDS treatment. All 6 goats survived the operation without complications. The xeno- autografts and xenografts showed the appearance, the remodeling process, and the cellular functions of myofibroblasts, 1 day, 1 month, and 3 months after operation, respectively. They were compatible with the native pulmonary leaflet (control group) except for the increased cellularity at 6 months. The xenografts revealed the new endothelial cell lining at that time. Conclusion: Treatment with NaCl-SDS was most effective in obtaining decellularized xenografts and facilitate seeding autologous endothelial cells. The xenografts and xeno-autografts were repopulated with myofibroblasts and endothelial cells in situ serially. Both of grafts served as a matrix for a tissue engineered heart valve and developed into autologous tissue for 6 months.

• Analytical Science and Technology
• /
• v.20 no.4
• /
• pp.347-354
• /
• 2007

A simple and rapid high-performance liquid chromatography assay for the determination of residual novobiocin levels in bovine, porcine, chicken, flatfish and japanese eel muscle has been developed and validated. The separation condition for HPLC/UV was optimized with phenyl hexyl ($4.6{\times}150mm$, $5{\mu}m$) column with 10 mM monobasic sodium phosphate buffer (pH 2.5)/acetonitrile (50/50, v/v) as the mobile phase at a flow rate of 1.0 mL/min and detection wavelength was set at 254 nm. Residues were extracted from tissue by blending with methanol and lipid materials were removed with n-hexane. Then, the methanol extract was evaporated to dryness under a nitrogen stream, reconstituted in the mobile phase. Aliquot of the organic extract was decanted and filtered through $0.45{\mu}m$ syringe filter. The $20{\mu}L$ of the resulting solution was injected into the HPLC system. The calibration ranges were $0.5{\sim}5{\mu}g/g$ and calibration curves were linear with coefficients of correlation better than 0.95. The limits of quantification were $0.5{\mu}g/g$ for all muscles. The recoveries of bovine, porcine, chicken, flatfish and japaneseel muscles were 99.8%, 102.4%, 91.0%, 104.0% and 93.0%, respectively. The procedures were validated according to the CODEX guideline, determining specificity, linearity, accuracy, precision, quantitation limit and recovery.

• Journal of Animal Science and Technology
• /
• v.49 no.5
• /
• pp.593-598
• /
• 2007

The current study was undertaken to determine the effects of sex steroid hormones(estrogen, testosterone and 19-nortestosterone) on differentiation and proliferation of pig preadipocytes. The preadipocytes were isolated from the backfat of new-born female pigs by collagenase digestion. 10－8M and 10－7M sex steroid hormones were treated to the cultured preadipocytes. Sex steroid hormones treated during the early stage of cell growth did not affect differentiation and proliferation of preadipocytes. However, testosterone and 19- nortestosterone treated during the late stage of cell growth stimulated differentiation of pig preadipocytes.

• Journal of Life Science
• /
• v.19 no.5
• /
• pp.587-592
• /
• 2009

A dramatic morphological change of embryos occurs at peri-implantation. Maternal and embryonic cross-talk during this period, initiated by signals from embryo(s), provides signals for maternal recognition of pregnancy and establishing and maintaining the pregnancy. However, the cellular, biochemical and genetic processes that direct embryo remodeling in mammalian species are not well studied or understood. In order to identify potential genes responsible for morphological change and cross-talk between embryo and uterus, an initial EST analysis was performed. A catalog of expressed genes (Transcriptome) from the d12 peri-implanting porcine embryos was constructed. Six clones were chosen from the initial ESTs for elucidation of their expression patterns during embryogenesis in early pregnancy. A number of these genes demonstrated unique expression profiles in a tissue, cell-type, and temporal fashion, indicating dynamic regulation of embryonic and endometrial gene expressions at different stages of pregnancy. Cross-talk between the embryo and endometrium of the pregnant uterus has provided a suitable micro-environment for the embryo's rapid and dramatic morphological changing process at the peri-implantation stage.

• Journal of Animal Science and Technology
• /
• v.52 no.1
• /
• pp.17-22
• /
• 2010

The current study was undertaken to determine the effects of sex steroid hormones (estrogen, testosterone and 19-nortestosterone) on proliferation and differentiation of preadipocytes of female and male pigs. The preadipocytes were isolated from the backfat of new-born female and male pigs by collagenase digestion and cultured in the $CO_2$ incubator. The concentration of $10^{-7}M$ and 10-6M sex steroid hormones were treated to the cultured preadipocytes. Regarding the effects on preadipocytes proliferation, high concentration ($10^{-6}M$) of all the three hormones increased proliferation of female preadipocytes,and only estrogen and testosterone increased proliferation of male preadipocytes. Regarding the effects on preadipocyte differentiation, all the three hormones increased differentiation of pig preadipocytes, regardless of hormone concentrations and sex of preadipocytes. The degree of stimulation of cell differentiation by sex steroid hormones was greater than that of cell proliferation.