• 한국동물학회지
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• 제34권2호
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• pp.209-216
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• 1991

신경세포의 분화에 미치는 insulin의 영향을 알아보기 위하여 계배의 단뇌 신경아세포를 serum-free defined medium에서 배양하였다. Immunofluorescence실험을 통하여 신경특이단백질인 MAP-2는 신경아세포의 세포체와 신경돌기에 존재하는 것으로 나타났다. 또한, 배양액내에 insulin의 농도를 증가하면 신경아세포의 신경돌기 형성이 증가할 뿐 아니라 MAP-2의 합성도 증가하였다. 따라서, 신경아세포의 형태적분화와 생화학적 분화는 서로 밀접하게 연관되어 일어나는 것으로 추측되며, insulin은 신경특이단백질의 합성을 촉진시킴으로써 신경아세포가 신경돌기를 형성하며 분화되어 가는데 결정적인 역할을 하는 것으로 보인다.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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• pp.71-71
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• 2003

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a multifunctional cytokine that has been implicated in the regulation of pre-implantation embryo development across several species. The aim of this study was to determine the effects of mouse granulocyte-macrophage colony-stimulating factor (mGM-CSF) on development of porcine parthenotes and nuclear transferred embryos, and on their expression of implantation-related genes. In the presence of bovine serum albumin, mGM-CSF did not increase the percentage of oocytes that developed to the blastocyst stage and at day 7 did not increase oocyte cell number. Addition of 10 mM GM-CSF to protein-free culture medium significantly increased the compaction and blastocoel formation of 1- to 2-cell parthenotes and cloned embryos developing in vitro. However, cell number was not increased when they were cultured in the presence of GM-CSF. Semi-quantitative reverse transcripts polymerase chain reaction (RT-PCR) revealed that mGM-CSF enhances mRNA expression of the leukemia inhibitory factor receptor, but does not influence interleukin-6 or sodium/glucose co-transporter protein gene expression in blastocyst stage parthenotes. These results suggest that mGM-CSF may enhance viability of porcine embryos developing in vitro in a defined culture medium.

• Journal of Applied Biological Chemistry
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• 제57권1호
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• pp.61-63
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• 2014

Aged black garlic (ABG) was extracted with 20% ethanol and water (crude extracts) and fractionated into three categories (>10, 3-10, and <3 kDa). The effect of crude extract supplements on anti-My-10 hybridoma cell growth and IgG1 antibody production was investigated in suspension culture with a chemically defined protein-free medium. We observed that supplementation of ABG to the cell culture medium stimulated anti-My-10 hybridoma cell growth and production of IgG1 antibody, particularly with fractionated ABG of low molecular weight. The stimulation depended upon the concentration and the size of the fractionated ABG. We also found that the growth-promoting activity was not correlated with high antibody production. These results suggest that fractionated ABG is a novel and promising alternative as an animal cell culture supplement.

• Archives of Pharmacal Research
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• 제28권8호
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• pp.963-969
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• 2005

Many reports have stated that some of the pathogenic bacteria can obtain iron from ferroproteins, such as cytochrome C, ferritin, hemin, hemoglobin, and myoglobin. These reports prompted us to determine if an opportunistic pathogenic fungus, Candida albicans, can utilize ferroproteins to circumvent the iron-regulatory effect of transferrin. The following assays were carried out to measure in vitro growth stimulation by the ferroproteins: as an initial step, C. albicans was cultured in iron-free (pretreated with apotransferrin for 24h) culture medium. Once Candida albicans yeast cell growth reached stasis from iron starvation, individual ferroproteins were added to the culture media. Results showed that hemin, hemoglobin, and myoglobin supported a partial growth recovery. Additional studies with haptoglobin, a serum protein that interacts with the globin moiety of certain ferroproteins, established that C. albicans could obtain iron from the haptoglobin-ferroprotein complexes. These data indicate that the heme part of the ferroproteins is the source of iron. This implies that heme oxygenase, CaHMX1 might be involved in bringing about dissociation of heme-containing protein for iron-acquisition. In addition, anticandidal activity of transferrin takes place not only by the process of iron regulation, but also by direct interaction with the yeast cells.

• 약학회지
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• 제55권3호
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• pp.203-212
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• 2011

Adipocytes have been known to secrete a number of important proteins called adipokines with roles in energy metabolism, reproduction, cardiovascular function and immunity. In this study we have attempted to identify intensively secretory proteins from 3T3-L1 adipocytes. 3T3-L1 preadipocytes were differentiated into mature adipocytes and then the cells were left in serum-free medium. The supernatant was filtrated and dialyzed. Lyophilized secretome was fractionated by two different methods, 1-D SDS PAGE and RP-FPLC. The tryptic peptides from the gel slices and the FPLC fractions were analyzed by nanoLC/ESI-MS/MS. We identified a total of 303 identical proteins from two methods, 251 proteins from 1-D gel and 184 proteins from RP-FPLC. 86 of them were listed as a secretory protein Finally, we identified many known or unknown secreted proteins existed in the low level including adiponectin, angiotensinogen, bone morphogenetic protein-1 (BMP-1), macrophage migration inhibitory factor (MIF), insulin like growth factor-II (IGF-II), interleukin-6 (IL-6), follistatin-related protein-1, minecan, and resistin. The existence of some of secreted proteins has been confirmed in RNA level. This proteomic experiment is useful for the intensive screening of secretory proteins in many kinds of other cells.

• Clinical and Experimental Reproductive Medicine
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• 제21권2호
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• pp.177-182
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• 1994

ICR female mice aged 3 to 4 weeks, were stimulated with 7.5 IU PMS injection. At 48-52h post-PMS injection, ovaries were dissected out and oocytes-cumulus complexes(OCCs) were divided into three groups, cumulus-free oocytes(O), cumulus-free oocyte cocultured with cumulus cells(O+C) and OCC. The oocyte were cultured in TCM199 containing various protein sources, FCS, BSA or PVP with gonadotropins(Gns) for 24h. Spermatozoa were collected from cauda epididymis and capacitated in T6 + BSA for 2h. After oocyte maturation in vitro(IVM) in different experimental groups, matured oocytes were inseminated with the capacitated spermatozoa in T6 + BSA for 6h. In the groups of IVM in TCM + BSA or PVP, fertilization(IVF) did not occur efficiently. However, increased fertilization was found in TCM+ FCS group. The oocytes groups, with cumulus cells showed decreased polyspermy in FCS group (O; 31.8 %, O + C; 12.2 %, OCC; 16%), the addition of Gns did not prevent polyspermy in all three groups. The rates of fertilization increased in zona-free oocytes in PVP group. This results showed that culture system for IVM and IVF could be improved. Furthermore, PVP can be used for the substitution of protein source during maturation, and its low rate of fertilization has been found due to zona hardening which occurred in FCS-free medium.

• Journal of Nutrition and Health
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• 제17권3호
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• pp.210-216
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• 1984

카제인 8%, 13% , 18% 수준의 동열량 함유 식이를 8%를 기준으로 하여 1일동일 열량 섭취하도록 급식시킨 이유쥐의 성장, 지방축적, 에너지 소비를 관찰하였다. 30일 급식시켰을 때 낮은 수준의 단백식이 (8%)군의 몸무게는 높은 수준(18%)의 단백섭취군 보다 43.01g더 가벼운 것으로 나타났다(최종 몸무게증가 ; ${85.57}{\pm}{7.5g}$ 대 ${128.58}{\pm}{11.64g}$, p<0.001). 도살체당 지방축적량은 8%단백식이군의 작은 몸의 크기에도 불구하고, 13%,18% 단백식이군과 비교할 때 차이를 보이지 않았다. 이와 반대로, 체질소 함량은 13%와 18% 단백식 군들이 8% 보다 훨씬 높았다. 총 실험기간(32일)동안 8%, 13%, 18%군의 에너지 소비는 각기 4,576.61kJ, 5,440.80kJ, 5,6067.67kJ로 추정되었다. 낮은 수준의 단백 식이군이 섭취한 과다에너지는 손실된 것으로 보인다. 간세포의 malic enzyme활성은 식이변화에 따라 변동이 없는 것으로 나타났다. 이러한 실험결과들로 부터, 낮은수준의 단백식이의 성장지연은 성장에 필요한 열량부족에서 온 것이 아니라, 질소공급의 부족에서 초래된 것이며, 단백질 급여 제한은, 에너지 섭취를 같게한 고단백군들과 비교할 때, 지방이외 무게는 감소시키는 반면, 체지방 합성과 축적능력에는 영향을 미치지 않는다는 결론을 얻었다.

• Asian-Australasian Journal of Animal Sciences
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• 제32권2호
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• pp.257-264
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• 2019

Objective: Dairy cattle nutrient requirement systems acknowledge amino acid (AAs) requirements in aggregate as metabolizable protein (MP) and assume fixed efficiencies of MP used for milk protein. Regulation of mammary protein synthesis may be associated with AA input and milk protein output. The aim of this study was to evaluate the effect of nanoemulsified methionine and cysteine on the in-vitro expression of milk protein (casein) in bovine mammary epithelial cells (MAC-T cells). Methods: Methionine and cysteine were nonionized using Lipoid S 75 by high-speed homogenizer. The nanoemulsified AA particle size and polydispersity index were determined by dynamic light scattering correlation spectroscopy using a high-performance particle sizer instrument. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed to determine the cytotoxicity effect of AAs with and without nanoionization at various concentrations (100 to $500{\mu}g/mL$) in mammary epithelial cells. MAC-T cells were subjected to 100% of free AA and nanoemulsified AA concentration in Dulbecco's modified Eagle medium/nutrient mixture F-12 (DMEM/F12) for the analysis of milk protein (casein) expression by the quantitative reverse transcription polymerase chain reaction method. Results: The AA-treated cells showed that cell viability tended to decrease (80%) in proportion to the concentration before nanogenesis, but cell viability increased as much as 90% after nanogenesis. The analysis of the expression of genetic markers related to milk protein indicated that; ${\alpha}_{s2}$-casein increased 2-fold, ${\kappa}$-casein increased 5-fold, and the amount of unchanged ${\beta}$-casein expression was nearly doubled in the nanoemulsified methionine-treated group when compared with the free-nanoemulsified methionine-supplemented group. On the contrary, the non-emulsified cysteine-administered group showed higher expression of genetic markers related to milk protein ${\alpha}_{s2}$-casein, ${\kappa}$-casein, and ${\beta}$-casein, but all the genetic markers related to milk protein decreased significantly after nanoemulsification. Conclusion: Detailed knowledge of factors, such nanogenesis of methionine, associated with increasing cysteine and decreasing production of genetic markers related to milk protein (casein) will help guide future recommendations to producers for maximizing milk yield with a high level of milk protein casein.

• Journal of Plant Biology
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• 제38권2호
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• pp.143-151
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• 1995

Addition of plant growth hormones [0.01 mg/L NAA and 0.2mg/L benzyladenine (BA)] to a woody plant medium stimulated the adventitious bud formation of poplar explants during culture. Endogenous IAA content increased rapidly at the initial culture stage and then decreased, being followed by rapid increment again at the late culture. But the content of trans-zeatin riboside (t-ZR) increased continuously during the culture. Cytoplasmic soluble proteins were analyzed by one- and two-dimensional SDS-PAGE. Increased amount of 40 kD band was detected by one-dimensional electrophoresis using Coomassie Blue staining during the culture and two distinctive proteins whose mol wt is 40,000 were detected by two-dimensional electrophoresis using autoradiography and these proteins were synthesized continuously prior to the adventitious bud formation. When the midvein segments were transferred to the actinomycin D-containing medium, the spots of adventitious bud-related proteins(ABRPs) did not disappeared but weakened in intensity. So, it is concluded that genes coding for the ABRPs are regulated to some degree at the transcriptional level. Also, they were not observed in BA-free medium, suggesting that these proteins be regulated by cytokinin, which made then possible to form the adventitious bud.

• Journal of Microbiology and Biotechnology
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• 제18권1호
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• pp.183-187
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• 2008

Vascular endothelial growth factors (VEGFs) are a family of proteins that mediate angiogenesis. $VEGF_{165}$ is a VEGF-A isoform and has been extensively studied owing to its potential use in therapeutic angiogenesis. This study established Chinese hamster ovary (CHO) cells overexpressing recombinant human $VEGF_{165}$ $(rhVEGF_{165})$ protein. The production rate of the established CHO cells was over 80mg/l of $rhVEGF_{165}$ protein from a 7-day batch culture process using a 7.5-l bioreactor with a 5-l working volume and serum-free medium. The $rhVEGF_{165}$ protein was purified to homogeneity from the culture supernatant using a two-step chromatographic procedure that resulted in a 48% recovery rate. The purified $rhVEGF_{165}$ protein was a glycosylated homodimeric protein with a higher molecular weight (MW) than the protein expressed from insect cells, suggesting that the glycosylation of the $rhVEGF_{165}$ protein in CHO cells differed from that in insect cells. The purified $rhVEGF_{165}$ protein in this study was functionally active with a half-maximal effective concentration of 3.8ng/ml and specific activity of $2.5{\times}10^5U/mg$.