• Asian-Australasian Journal of Animal Sciences
• /
• 제2권4호
• /
• pp.595-598
• /
• 1989

Bovine embryos/ova obtained from in-vitro fertilization were either co-cultured on a monolayer of bovine cumulus cells or cultured in medium alone. Embryos/ova co-cultured with cumulus cells developed to 8-cell (30.9%), morula (29.8%) and blastocyst stages (26.6%) after 3-4, 5-6, and 7-8 days of culture, respectively, while embryos/ova cultured in medium alone failed to develop beyond 8-cell (0-13.3%), morula (0-1.5%) and blastocyst stages (0%). The results of this study demonstrated the beneficial effect of cumulus cells on the development of bovine embryos.

• 대한생식의학회:학술대회논문집
• /
• 대한불임학회 2003년도 제45차 추계학술대회
• /
• pp.150-151
• /
• 2003

• Fisheries and Aquatic Sciences
• /
• 제4권4호
• /
• pp.186-191
• /
• 2001

Vitellogenin (VTG) and VTG mRNA induction by $estradiol-17\beta\;(E_2)$ were examined in the primary cultures of hepatocyte in the rainbow trout. Hepatocytes were precultured for 2 days, then $E_2$ was added and cultured for another 5 days. Media and hepatocytes were then analyzed by electrophoresis and Northern blotting for VTG and VTG mRNA, respectively. The hepatocytes were formed a few aggregates within 5 days without further spreading to a monolayer. Cell viability and high DNA content were maintained during the incubation. The hepatocyte culture with E2 induced a weak VTG band at a molecular weight of 175kDa on Day 2 after $E_2$ addition. The relative amount of VTG was expressed in percentage of total protein concentrations. VTG was gradually increased as $1.9\%$ on Day 2, $6.3\%$ on Day 4 and $7.3\%$ on Day 5. VTG mRNA band was detected at about 6.6 kb in the culture with $E_2$ at day 1 of culture. The level of VTG mRNA expression linearly increased with time until Day 5 (r=0.97).

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2003년도 생물공학의 동향(XIII)
• /
• pp.335-339
• /
• 2003

본 연구에서는 연골조직으로부터 세포를 분리한 후 10%의 혈청 첨가 배지와 무혈청 배지에서의 세포 성장속도, GAGs 합성 및 Col. II의 발현을 확인하였다. EGF를 첨가한 무혈청 배지를 연골세포의 증식에 매우 효과적이었으나 GAGs 합성 및 Col. II 의 합성을 저해하였다. 또한 무혈청 배지에서 배양한 세포를 차후 신체내로 이식한다면 연골세포의 특성인 Col. II를 재합성할 수 있음을 간접적으로 확인하였다. 이는 무혈청 배지를 이용하여 평판배양 시 짧은 기간 내 연골세포 치료제로서 가능한 세포수를 얻기 위한 모델로서 유용할 것으로 생각된다. 또한 계대배양에 따른 분화 및 형태의 변화 등의 문제점들은 생분해성 지지체와 연계하여 해결할 수 있을 것으로 생각되며, 차후 위의 SFM을 이용한 3D 배양에 대한 연구를 수행할 예정이다.

• Macromolecular Research
• /
• 제11권6호
• /
• pp.488-494
• /
• 2003

We have fabricated gelatin-immobilized polyurethane foams (PUFG) by dipping polyurethane foam (PUF) in an aqueous solution containing gelatin and by subsequent reaction with glutaraldehyde after freeze-drying. Gelatin aqueous solutions of different concentrations were used as the dipping solutions to control the amount of immobilized gelatin. The average pore size of PUF decreased with an increase in gelatin concentration. It was found from the hepatocyte adhesion experiment that the amount of hepatocytes seeded on PUFG1, prepared by using a 1% aqueous gelatin solution, was higher than that on other PUFGs. The hepatocytes inoculated in PUFG1, were slightly aggregated as the incubation time increased. The cells inoculated in PUFG1 showed higher ammonia removal ability than those monolayer-cultured on a gelatin-immobilized polystyrene dish (PSG) after 1 and 4 days of incubation time. The inoculated cells exhibited higher albumin secretion relative to monolayer-cultured hepatocytes on PSG. Albumin secretion by hepatocytes seeded on PUFG1 was increased by the presence of serum and was further increased by both the presence of serum and cytokines. The results obtained from a 3-(3,4-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) assay indicated that PUFG can provide a better microenvironment for hepatocyte culture along with nutrition and metabolite transfer through the high porosity of PUF.

• 한국가축번식학회지
• /
• 제22권1호
• /
• pp.51-59
• /
• 1998

This study was undertaken to investigate the effects of ITS and EGF on embryonic development of in vitro matured and fertilized bovine oocytes in culture medium su, pp.ementing with or without calf serum. When fertilized oocytes were cultured in TCM-199 containing 0, 0.1, 0.5 and 1.0\ulcorner/ml ITS with 5% calf serum, the rates of development to blastocyst stage and the cell number of blastocysts were not significantly different among all treatments. And also monolayer of cumulus/granulosa cells prepared in containing calf serum and ITS were no beneficial effects of embryonic development. On the other hand, when EGF was su, pp.imented to TCM-199 containing calf serum or calf serum free, embryonic development rates(24.0 2.8% to 29.2 1.7% or 8% to 9%) and cell number of blastocysts(p<0.05) were significantly increased compared with EGF-free(22.1 2.1 or 1.0%, p<0.05). But when fertilized oocytes were cultured with cumulus/granurosa cells in TCM-199 containing EGF and calf serum, the rate of embryos development to the blastocyst stage and cell number of blastocysts were not significantly different compared with EGF-free and any concentrations. These results showed that ITS and EGF was not improved the development of bovine embryo in vitro matured and in vitro fertilized with calf serum and/or monolayer of cumulus/granulosa cells.

• Asian-Australasian Journal of Animal Sciences
• /
• 제15권3호
• /
• pp.340-345
• /
• 2002

In this experiment, oocytes were collected from goat ovaries available in slaughterhouse by follicle puncture method. Morphologically culturable type of oocytes which having compact, multilayered cumulus granulosa cell complex and evenly granulated cytoplasm, was separated under a stereozoom microscope. Oocytes were washed thoroughly in maturation medium containing TCM-199, $1{\mu}g/ml$ estradiol-$17{\beta}$, 0.5 ${\mu}g/ml$ FSH, $100{\mu}g/ml$ LH, 3 mg/ml BSA and 10% estrus goat serum. Washed oocytes were cultured into maturation medium on granulosa cell monolayer. Culture plate was then kept into $CO_2$ incubator at $38{\pm}1^{\circ}C$, maximum humidity and 5% $CO_2$ for 18 h. After maturation the oocytes were washed thoroughly with maturation medium containing polyvinyl alcohol (PVA) without serum and BSA and further cultured for 12 h for secretory proteins of oocytes. PVA medium was collected, pooled and concentrated by 5000 cut off centrisart. Secretory proteins were separated on 12.5% SDS-PAGE. A total number of 3.41 oocytes per ovary were obtained and 2.17 culturable oocytes per ovary were cultured into maturation medium. After 18 h of maturation, 4,567 oocytes (1.82 oocytes per ovary) were further cultured into serum and BSA free PVA medium for its secretory proteins. Four secretory proteins of oocytes with approximately molecular weight of 45, 55, 65 and 95 kDa were obtained on SDS-PAGE in silver staining and three proteins with approximately molecular weight of 45, 55 and 65 kDa in Coomassie brilliant blue staining. In conclusion, four secretory proteins with approximately molecular weight of 45, 55, 65 and 95 kDa was obtained from in vitro cultured oocytes of goats.

• 한국환경보건학회지
• /
• 제21권1호
• /
• pp.68-73
• /
• 1995

화합물질의 개발사용 및 환경오염 물질의 증가로 인하여 남성수태능력에 영향을 미치는 물질이 증가하고 있는 형편이다. 따라서 신속하고 경제적으로 남성 수태능력 영향물질을 평가하기 위하여 랫드의 고환세포를 이용한 남성 수태능력시험법을 개발하고자 본 시험을 실시하였다. 랫드의 고환 세포로부터 주요한 기능을 가진 Sertoli cell과 Leyding cell을 효소처리, 필터 그리고 percoll gradient과정을 거쳐 각각 분리하였다. 분리된 두 종류의 세포는 32$\circ$C에서 초기 배양한 후 37$\circ$C에서 계속 배양하였다. 본 배양조건에서 두 종류 세포는 양호한 발육을 보였으며 약 2주간의 배양이 가능하였다. Sertoli cell은 3주령의 랫드에서 분리가 잘되었고 Leydig cell은 85.7%의 생존율을 보였다. 이상의 결과와 같이 랫드의 고환세포중 Sertoli cell과 Leyding cell을 분리 배양하였으며, 앞으로 독성물질의 영향을 평가하기 위한 생화학적 및 구조적인 index 설정을 위한 연구가 필요하다고 사료된다.

• 한국수정란이식학회지
• /
• 제11권3호
• /
• pp.217-223
• /
• 1996

This study was conducted to investigate the effects of co-culture for the development rate to morula /blastocyst stages of early porcine embryos, derived from oocytes matured and fertilized in vitro, with porcine endometrial cell monolayers(PEM) in the two different media, respectively. The rates of embryos developed to 2-, 4-, 8~16-cell and morula /blastocyst stage were 49.6, 40.5, 28.2 and 15.3% in Ham's F-10 with PEM, and 55.3, 45.9, 32.7, and 17.6% in TCM-HEPES with PEM, respectively. The above development rates to morula /blastocyst stages were significantly higher than those of the embryos cultured in the Ham's F-10 and TGM-HEPES without PEM(P<0.05). The in vitro development rates to the morula /blastocyst stage of 1-cell embryos cultured in Ham's F-10 and TCM-HEPES without PEM were 0~1.2%. Especially, most of embryos were observed to arrest the development beyond 4-cell stages. As shown in the above results, the co-culture of in vitro produced porcine embryos with PEM in the two different media enhanced the development of fertilized eggs to morula /blastocyst stages in vitro. However, we didn't find out any differences for the in vitro development to morula /blastocyst stages between Ham's F-10 and TcM-HEPES media.

• 대한화장품학회:학술대회논문집
• /
• 대한화장품학회 2003년도 IFSCC Conference Proceeding Book I
• /
• pp.85-96
• /
• 2003

Normal human skin, constantly challenged by environmental micro-organisms, has an innate ability to fight invading microbes through antimicrobial peptides. These peptides, described in both plant and animal kingdoms are able to inactivate a broad spectrum of micro-organisms. Mammalian defensins constitute one of the most common antimicrobial peptide family. Among the three human beta-defensins hBD1, hBD2 and hBD3 produced in epithelia, only hBD2 and hBD3 are inducible and additionally have been described as expressed by differentiated keratinocytes at site of inflammation and infection. The aims of these studies were to define a cell culture model in which the basal production of hBD could be detected and up-regulated in order to enhance skin auto-protection against micro-organisms. A specific Polymerase Chain Reaction method have been developed for hBD2 and hBD3 mRNA detection in non-differentiated monolayer keratinocytes cell culture. We have been able to demonstrate that in vitro, hBD2 and hBD3 expression in normal human keratinocytes could be detected and enhanced by TNF-alpha and IFN-gamma, in hypercalcic culture conditions. This research opened the possibility of the development of cosmetic active compounds, able to induce the expression of skin natural antibiotic peptides responsible about microflora ecology of the skin.