• 대한수의학회지
• /
• 제37권3호
• /
• pp.647-659
• /
• 1997

To overcome the difficulties of collecting and culture of primary cell from genital tract on embryonic development, the present study was carried out to investigate the critical effect of cell lines, such as BRL and Vero cell and its conditioned medium on the development of early Korean native cattle embryos in vitro. Oocytes collected from slaughterhouse ovaries were matured in TCM199 containing FSH, estradiol-$17{\beta}$ and FBS with granulosa cell monolayer for 24 hours and then fertilized in vitro using frozen-thawed, heparin-treated spermatozoa in TALP for 30 hours. And then early embryos (1-2cell) were cultured in TCM199 containing 10% FBS with BOEC, Granulosa, BRL, Vera cell monolayers and conditioned medium for 2~3 days. Development to morulae and blastocysts were recorded, also examined the number of blastomeres presented a valuable parameter for the evaluation of embryonic development. The early cleavage rates of in vitro fertilized embryos co-cultured, there was no differences between primary cell and cell lines(p<0.05). The rate of development to the later stage, coculture of BRL cell was significantly higher than that of the primary cell(p<0.05). The rates of development to morula and blastocyst were significantly higher in vero cell than BRL, Granulosa, Oviduct epithelial cell conditioned medium. In the result of effect of serum on development of early bovine embryos, the use of media containing serum were significantly higher than the use of not containing one on development of early and later stage of embryos. The result of number of blastomeres in blastocysts, there is no differences between primary cell and cell lines. The blastocysts from coculture were higher than from conditioned medium in blastomere cells. In summary, these experments have proved that the culture system in TCM199 with BRL, Vero cell monolayers is effective on in vitro development of early bovine embryos, In addition, it is effective to development of bovine embryos that containing serum in conditioned medium, or in co-culture rather than in conditioned medium alone. The use of cell lines opponent to primary cells is effective in bovine embryo culture.

• 한국수정란이식학회지
• /
• 제12권3호
• /
• pp.293-300
• /
• 1997

Supplementation of maturation medium with additional granulosa cells has beneficial effect on in vitro maturation of bovine follicular oocytes and their subsequent cleavage and development in vitro. However, maturation system using granulosa cells have some disadvantages that collection of granulosa cells is cumbersome and metabolic activity of the cells is variable according to ovarian cycle or follicular size. We hypothesized that bovine immsture oocytes matured without granulosa cell coculture can fertilize and develop normally if the medium volume per oocyte is reduced during in vitro maturation. Immature oocytes were matured for 24 hours in a TCM199 containing 10% fetal calf serum, anterior pitultary hormone (0.02 AU /ml Antrinⓡ) and estradiol with or without granulosa cells in vitro. In Group 1, 35 to 40 oocytes were matured in a well of 4-well plastic dish containing 500 $\mu$l of maturation medium and granulosa cells, and 9 to 10 oocytes were matured in a 50-$\mu$l drop of maturation medium without granulosa cells in Group 2. After maturation, oocytes were coincubated with sperm for 30 hours in a modified Tyrode's medium (IVF). Inseminated oocytes were cultured in a microdrop (30 $\mu$l) of a synthetic oviduct fluld medium (SOFM) containing BSA, Minimum Essential Medium essential and non-essential amino acids for 9 days. As a preliminary experiment, we investigated the beneficial effect of granulosa cells during maturation on subsequent cleavage and development using the same type of culturedishes (4-well dish). Granulosa cells could not increase embryo cleavage after fertilization but significantly improved (p<0.05) embryo development to expanding blastocyst (Table1 and 2). In Group 1, 68 and 80% of inseminated oocytes have cleaved at 30 hours and 2 days after IVF, respectively, which is similar (p>0.05) to the result of Group 2 (69% at 30 hours and 78% at 2 days after IVF). The oocytes in Group 2 showed 21 and 11% of developmental rates to expanding and hatching blastocysts, respectively, which was not significantly different (p>0.05) from those (20 and 10%, respectively) of oocytes in Group 1. In conclusion, it has been clarified that a microdrop culture system without granulosa cells for in vitro maturation can support bovine embryonic development to blastocyst in vitro as readily as a granulosa cell coculture system.

• Journal of Ginseng Research
• /
• 제48권1호
• /
• pp.77-88
• /
• 2024

Background: Lung inflammation occurs in many lung diseases, but has limited effective therapeutics. Ginseng and its derivatives have anti-inflammatory effects, but their unstable physicochemical and metabolic properties hinder their application in the treatment. Panaxadiol (PD) is a stable saponin among ginsenosides. Inhalation administration may solve these issues, and the specific mechanism of action needs to be studied. Methods: A mouse model of lung inflammation induced by lipopolysaccharide (LPS), an in vitro macrophage inflammation model, and a coculture model of epithelial cells and macrophages were used to study the effects and mechanisms of inhalation delivery of PD. Pathology and molecular assessments were used to evaluate efficacy. Transcriptome sequencing was used to screen the mechanism and target. Finally, the efficacy and mechanism were verified in a human BALF cell model. Results: Inhaled PD reduced LPS-induced lung inflammation in mice in a dose-dependent manner, including inflammatory cell infiltration, lung tissue pathology, and inflammatory factor expression. Meanwhile, the dose of inhalation was much lower than that of intragastric administration under the same therapeutic effect, which may be related to its higher bioavailability and superior pharmacokinetic parameters. Using transcriptome analysis and verification by a coculture model of macrophage and epithelial cells, we found that PD may act by inhibiting TNFA/TNFAR and IL7/IL7R signaling to reduce macrophage inflammatory factor-induced epithelial apoptosis and promote proliferation. Conclusion: PD inhalation alleviates lung inflammation and pathology by inhibiting TNFA/TNFAR and IL7/IL7R signaling between macrophages and epithelial cells. PD may be a novel drug for the clinical treatment of lung inflammation.

• Biotechnology and Bioprocess Engineering:BBE
• /
• 제5권2호
• /
• pp.92-98
• /
• 2000

Mesangial cell has several key roles in thee control of glomerular function: it partocipates in the regulation of glomerular filtration rate, macromolecular clearance, and as both a source and target of numerous hormones and autocrines. Many of these insights into mesangial cell function have been obtained by studying mesangial cells in culture. However, no suitble cell lines have established yet. We here reported the immortalization of rat kidney glomeruar mesangial cell by transfection of E6 and E7 genes of human papillomavirus type 16 (HPV-16) via electroporation and lipofection. The reslts showed that only electroporation could transfect the genes to mesangial cells and the transfected cells maintained the viability for longer than 6 months. Fluorescence microscopic observation showed that cellular contractility and phagocytosis, which are the two main phenotypes of mesangial cells with rat glomerular epithelial cells showed that the growth of mesangial dells was suppressed by epithelial cell, but the growth of epithelisl cells was enhanced by mesangial cells. Moreover, Such results may imply that the glomerular cell-cell interaction plays an important role in the regulation of cell proliferation and differentiation.

• Journal of Microbiology and Biotechnology
• /
• 제18권3호
• /
• pp.545-551
• /
• 2008

When cultivated aerobically, Aspergillus niger hyphae produced extracellular glucoamylase, which catalyzes the saccharification of unliquified potato starch into glucose, but not when grown under anaerobic conditions. The $K_m\;and\;V_{max}$ of the extracellular glucoamylase were 652.3 mg/l of starch and 253.3 mg/l/min of glucose, respectively. In mixed culture of A. niger and Saccharomyces cerevisiae, oxygen had a negative influence on the alcohol fermentation of yeast, but activated fungal growth. Therefore, oxygen is a critical factor for ethanol production in the mixed culture, and its generation through electrolysis of water in an electrochemical bioreactor needs to be optimized for ethanol production from starch by coculture of fungal hyphae and yeast cells. By applying pulsed electric fields (PEF) into the electrochemical bioreactor, ethanol production from starch improved significantly: Ethanol produced from 50 g/l potato starch by a mixed culture of A. niger and S. cerevisiae was about 5 g/l in a conventional bioreactor, but was 9 g/l in 5 volts of PEF and about 19 g/l in 4 volts of PEF for 5 days.

• 생명과학회지
• /
• 제19권5호
• /
• pp.676-679
• /
• 2009

미생물들 사이에 존재하는 수평적 유전자 전달을 이용하여 Jurkat T cell에 대하여 새로운 항생물질을 생산하기 위해 토양박테리아 AY2000과 여러 종류의 항생물질 생산 균주인 Streptomyces griseus의 공배양을 수행하였다. MTT assay를 수행하여 세포 독성을 실험을 하였을 때 공배양 상등액은 각각 배양 하였을 때보다 높은 세포독성을 보였고 또한 48시간 배양 하였을 때 가장 높은 활성을 나타내는 것으로 나타났다. 더욱이 DAPI 염색을 하였을 때 Jurkat T cell 세포의 세포핵의 변화도 관찰 되었다. 이런 결과는 공배양 상등액에 새로운 항생물질이 생성되었음을 보여주었고 이런 방법으로 새로운 항생물질 생산에 이용되어 질수 있음을 의미한다.

• 한국수정란이식학회지
• /
• 제11권3호
• /
• pp.241-248
• /
• 1996

This study was conducted to improve the production efficiency of in vitro produced (IVP) embryos in Korean Native cows. The optimal conditions and procedures for in vitro maturation(IVM), in vitro fertilization(IVF) and in vitro culture(IVC) of bovine follicular oocytes and IVP embryos were evaluated. Immature follicular oocytes were collected fiom the follicles of bovine ovaries obtained from abattoirs. The oocytes of Grade I and II for IVM were cocultured with monolayered bovine oviductal epithelial cells(BOEG) or granulosa cells in TCM-199 solution supplemented with follicle stimulating hormone, lutenizing hormone, estradiol-17$\beta$ and heat inactivated fetal calf serum at 39$^{\circ}C$ under 5% $CO_2$ in air for 14 to 24 hours. Most of the oocytes(93%) matured to metaphase II in 24 hours. The cocultured IVM oocytes were fertilized in vitro at significantly(P<0.05) higher rate with BOEC(83.8%) and with granulosa cells(84.6%) than the non-cocultured IVM oocytes(73.6%). The IVM-IVF embryos developed to morula and blastocyst at significantly(P<0.05) higher rate in coculture with BOEC(41.2%) than with granulosa cells(23.1%) or conditioned medium(23.4%).

• The Korean Journal of Physiology and Pharmacology
• /
• 제16권4호
• /
• pp.243-247
• /
• 2012

Recent studies suggest that immunization with autologous dendritic cells (DCs) results in protective immunity and rejection of established tumors in various human malignancies. The purpose of this study is to determine whether DCs are generated from peripheral blood mononuclear cells (PBMNs) by using cytokines such as F1t-3 ligand (FL), granulocyte macrophage-colony stimulating factor (GM-CSF), IL-4, and TNF-${\alpha}$, and whether cytotoxic T cells activated against the thyroid cancer tissues by the DCs. Peripheral blood was obtained from 2 patients with thyroid cancer. DCs were established from PBMNs by culturing in the presence of FL, GM-CSF, IL-4, and TNF-${\alpha}$ for 14 days. At day 14, the differentiated DCs was analyzed morphologically. The immunophenotypic features of DCs such as CDla, CD83, and CD86 were analyzed by immunofluorelescence microscopy. At day 18, DCs and T cells were incubated with thyroid cancer tissues or normal thyroid tissues for additional 4 days, respectively. DCs generated from the PBMNs showed the typical morphology of DCs. Activated cytotoxic T lymphocytes (CTLs) were observed also. DCs and the CTLs were attached to the cancer tissues on scanning electron microscope. The DCs activated the CTLs, which able to specifically attack the thyroid cancer. This study provides morphologic evidence that the coculture of T cells/cancer tissues activated the T cells and differentiated CTLs. The CTLs tightly adhered to cancer tissues and lysed cancer tissues vigorously. Therefore DCs could be used as potential vaccines in the immunotherapy.

• Journal of Microbiology and Biotechnology
• /
• 제17권11호
• /
• pp.1758-1764
• /
• 2007

Synthesis of oligosaccharides during milk fermentation was attempted by inoculating Leuconostoc citreum with Lactobacillus casei, Lb. delbrueckii subsp. bulgaricus, and Streptococcus thermophilus as starters. Dextransucrase of Ln. citreum worked as a catalyst for the transglycosylation reaction of sugars; sucrose was added as the glucose donor, and lactose or maltose acted as the acceptor compound for the reaction. When 4% sucrose was added in milk, glucosyl-lactose was synthesized (about 1%, w/v) after 1-2 days of fermentation at 15 or $25^{\circ}C$. Alternatively, when sucrose and maltose (2% each, w/v) were added, panose (about 1 %, w/v) and other isomaltooligosaccharides were made in a day at $15-35^{\circ}C$. Growth patterns of lactobacilli and streptococci starters were not affected by the coculture of leuconostoc starter, but the rate of acid synthesis was slightly slowed at every temperature. Addition of sugars in milk did not give any adverse effect on the lactate fermentation. Accordingly, the use of leuconostoc starter and addition of sugars in milk allowed the production of oligosaccharides-containing fermented milk, and application of this method will facilitate the extensive development of synbiotic lactate foods.

• 한국정보통신학회:학술대회논문집
• /
• 한국정보통신학회 2015년도 춘계학술대회
• /
• pp.943-946
• /
• 2015

탈수초화를 위해 쥐의 배아(임신 16일)의 슈반세포와 뉴런 세포가 척수신경절로 부터 in vitro 시스템에서 배양되었다. 항 유사분열제로 첨가된 분리 정제된 척수신경절 세포와 분리 정제된 슈반세포가 공동배양 배양되었고 배양되었고 수초화 과정이 구축되었다. 이렇게 형성된 공동 배양에 M. leprae-specific phenolic glycolipid-1 (PGL-1)을 처리하고 myelin basic protein (MBP)의 항체를 이용하여 탈수초화가 형성되었음을 확인하였다.