• 제목/요약/키워드: CHO-K1 cell

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Molecular and Cellular Analyses of NCP, a Nuclear and Centrosomal Protein in Mouse Gametes and Early Embryos

  • Oh, Hwa-Soon;Youn, Hong-Hee;Lee, Kwang-Hee;Son, Chae-Ick;Lee, Sang-Ho
    • 한국동물번식학회:학술대회논문집
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    • 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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    • pp.97-97
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    • 2003
  • For many animals the centrosome consists of a pair of centrioles and surrounding pericentriolar materials (PCMs). PCMs have been known to play roles during cell division. It is known that centrioles are necessary to assemble centrosomal components. However, many types of oocytes undergo meiosis without centrioles. It is known that in nonmurine mammalian species, the sperm introduces an intact proximal centriole unlike sea urchin where two centrioles are introduced. In case of mouse sperm, the presence of centrosome is not clear In this study, a monoclonal antibody was developed to investigate centrosome during mouse germ cell and early embryo development. Results of immunostaining and Western blotting in CHO cells suggest that the monoclonal antibody recognizes a nuclear and centrosomal protein, thus called NCP. The NCP monoclonal antibody was used to screen a cDNA expression library prepared from 12.5 mouse brain to isolate NCP gene. Nucleotide size of NCP gene obtained from immunoscreening was about 5.5kb. It is determined that the NCP may be closely related with pericentriolar material -1 gene (Pcm-1) from the result of sequencing analysis. The molecular weight, 66kDa, calculated by known DNA sequence in database is consistent with that of detected from Western blotting using CHO cell lysates. Therefore, it is assumed that NCP may be alternative splicing form of Pcm-1 of which molecular weight is 228kDa. In mouse oocytes, NCP was distributed in nucleus as in CHO cells. It was shown that the NCP was localized around neck region, probably the centrosome in mouse neck region. Interestingly, dramatic change in distribution of NCP was also shown in male germ cell development. Finally, we observed the cellular distribution of NCP during early embryo development. NCP was detected in nucleus as well as centrosome foci. It is suggested that the centrioles reassembly we occurring in blastocysts and then affects the distribution of NCP.

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신경세포 배양법을 이용한 methamphetamine과 cadmium의 신경독성 평가 (Neurotoxicity Assessment of Methamphetamine and Cadmium Using Cultured Neuronal Cells of Long-Evans Rats)

  • 조대현;김준규;정용;이봉훈;김은엽;김정구;조태순;김진석;문화회
    • Toxicological Research
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    • 제12권1호
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    • pp.69-79
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    • 1996
  • Primary culture of cerebellar neuronal cells derived from 8-day old Long-Evans rats was used. Pure granule cells, astrocytes or mixed cells culture systems were prepared. These cells were differentiated and developed synaptic connections. And the astrocytes were identified by immunostaining with glial fibrillary acidic protein (GFAP). Methamphetamine (MAP), which acts on dopaminergic system and cadmium (Cd), a toxic heavy metal, were applied and biochemical assays and electrophysiological studies were performed. $LC_50$ values estimated by MTT assay of MAP and Cd were 3 mM and 2$\mu M$ respectively. Cells were treated with 1 mM or 2 mM MAP and 1$\mu M$ $CdCl_2$ for 48 hour, and the incubation media were analyzed for the content of released LDH. MAP (2 mM) and Cd significantly increased the LDH release. Cell viability was decreased in both groups and some cytopathological changes like cell swelling or vacuolization were seen. The cerebellar granule cells were used for measuring membrane currents using whole-cell clamp technique. Sodium and potassium currents were not affected by MAP neither Cd, but calcium current was significantly reduced by Cd but not affected by MAP. Therefore, in vitro neurotoxicity test system using neuronaI cells and astrocytes cultures were established and can be used in screening of potential neurotoxic chemicals.

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황기가 in vitro에서 자궁근종세포(子宮筋腫細胞)에 미치는 영향(影響) (Effects of Astragalus membranaceus Extract on Cell Cycle and Apoptosis of Human Leiomyoma Cell)

  • 박준규;이진무;이창훈;조정훈;장준복;이경섭
    • 대한한방부인과학회지
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    • 제21권2호
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    • pp.49-58
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    • 2008
  • Purpose: This study was conducted to investigate the effects of Astragalus membranaceus extract solution on cell cycle regulation and apoptosis of human leiomyomal cell. Methods: The leiomyoma cell of patients was used in the study, and we administered the extract solution of Astragalus membranaceus concentration at 1, $10mg/m{\ell}$ to the leiomyoma cell for 48 hours. We used flow cytometry and western blotting to confirm cell cycle and apoptosis. Results: In flow cytometry, G1 phase of the $1mg/m{\ell}$ and $10mg/m{\ell}$ group was shortened and S phase of the $1mg/m{\ell}$ and $10mg/m{\ell}$ group was increased. Cycline D1 expression increased in 1, $10mg/m{\ell}$ group. Bcl-2 expression decreased in 1, $10mg/m{\ell}$ groups than control group. And Bax expression that regulated cell apoptosis more increased in 1, $10mg/m{\ell}$ group than control group. VEGF expression rised in higher Astragalus membranaceus concentration group. Conclusion: This study suggest that Astragalus membranaceus might induce cell apoptosis of leiomyoma cell and shorten cell cycle. And Astragalus membranaceus would not have the regulation effect of cell cycle.

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Apoptotic Signaling Pathways: Caspases and Stress-Activated Protein Kinases

  • Cho, Ssang-Goo;Choi, Eui-Ju
    • BMB Reports
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    • 제35권1호
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    • pp.24-27
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    • 2002
  • Apoptotic cell death is an active process mediated by various signaling pathways, which include the caspase cascade and the stress-activated protein kinase pathways. The caspase cascade is activated by two distinct routes: one from cell surface and the other from mitochondria. Activation of the route from cell surface requires the cellular components that include membrane receptors, adaptor proteins such as TRADD and FADD, and caspase-8, while activation of the other from mitochondria requires Apaf-1, caspase-9, and cytosolic cytochrome c. On the other hand, persistent stimulation of the stress-activated protein kinase pathway is also shown to mediate apoptosis in many cell types. Gene-targeting studies with jnk- or jip-null mice, in particular, strongly suggest that this signaling pathway plays a pivotal role in the cellular machinery for apoptosis.

Synthesis and In Vitro Cytotoxicity of 1,3-Dioxoindan-2-Carboxylic Acid Arylamides

  • Jung, Jae-Kyung;Ryu, Jin-Hyeong;Yang, Sung-Il;Cho, Jung-Sook;Lee , Hee-Soon
    • Archives of Pharmacal Research
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    • 제27권10호
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    • pp.997-1000
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    • 2004
  • A series of 1,3-dioxoindan-2-carboxylic acid arylamides were synthesized and evaluated for in vitro cytotoxicity against four human cancer cell lines (HOP62, SK-OV-3, MD-MB-468 and T- 47D). The most active was compound 3e (1.2 ${\mu}M$ against SK-OV-3 cell line) bearing a 4- methyl substituent.

생물의약품 제조공정에서 Bovine Viral Diarrhoea Virus 정량 검출을 위한 Real-Time RT-PCR (Real-Time RT-PCR for Quantitative Detection of Bovine Viral Diarrhoea Virus during Manufacture of Biologics)

  • 조항미;이동혁;김현미;김인섭
    • 한국미생물·생명공학회지
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    • 제36권1호
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    • pp.34-42
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    • 2008
  • 소의 혈액, 세포, 조직, 기관 등 소 유래 물질을 원료로 사용한 생물의약품, 조직공학제제, 세포치료제의 경우 소 유래원료 물질에 다양한 바이러스가 오염된 사례가 있기 때문에 바이러스 안전성 검증이 필수적이다. BVDV는 동물세포주, 우혈청 등에 가장 흔하게 오염되는 바이러스이다. 소 유래물질을 원료로 하는 생물의약품, 조직공학제제, 세포치료제 등에서 BVDV안전성을 확보하기 위해, 원료물질, 제조공정, 완제품에서 BVDV를 정량적으로 검출하고, 제조공정에서 BVDV제거 검증을 위한 시험법으로 활용이 가능한 BVDV real-time RT-PCR시험법을 확립하였다. BVDV에 특이적인 primer를 선별하였으며, 형광염료 SYBR Green I을 사용하여 BVDV RNA 정량 검출 시험법을 최적화하였다. 세포배양법에 의한 감염역가와 비교한 결과 real-time RT-PCR 민감도는 $1TCID_{50}/mL$이었다. 확립된 시험법의 신뢰성(reliability)을 보증하기 위해 시험법 검증을 실시한 결과 특이성 (specificity)과 재현성(reproducibility)이 우수함을 확인하였다. 확립된 real-time RT-PCR을 생물의약품 제조공정 검증에 적용할 수 있는지 확인하기 위하여 인위적으로 BVDV를 오염시킨 CHO 세포주와 소 유래콜라겐에서 BVDV검출 시험을 실시하였다. BVDV를 감염시킨 CHO 세포와 세포배양 상청액에서 BVDV를 정량적으로 검출할 수 있었다. 소 유래 콜라겐에서도 $10TCID_{50}/mL$까지 정량적으로 검출할 수 있었다.

액화질소를 이용한 오픈 셀 실리콘 폼의 냉동 절삭조건 최적화 (Cryogenic Machining of Open-Cell Silicone Foam)

  • 황지홍;조광희;박민수
    • 한국생산제조학회지
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    • 제23권1호
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    • pp.32-37
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    • 2014
  • Open-cell silicon foam is difficult to cut using conventional machining processes because of its low stiffness. That is, open-cell silicon foam is easily pressed down when the tool is engaged, which makes it difficult to remove the material in the form of chip. This study proposes an advanced method of machining open-cell silicon foam by freezing the material using liquid nitrogen. Furthermore, the machining conditions are optimized to maximize the efficiency of material removal and minimize the usage of liquid nitrogen by conducting experiments under various machining conditions. The results show that open-cell silicone foam products with free surface can be successfully machined by employing the proposed method.

Butyrate처리된 차이니즈 햄스터 난소세포에서 Hepatitis B 바이러스 인간화항체의 생산 (Production of Humanised Anti-hepatitis B Antibody in Butyrate-Treated Chinese Hamster Ovary Cells)

  • 박세철;이재선;이병규;강희일
    • 한국미생물·생명공학회지
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    • 제34권1호
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    • pp.47-51
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    • 2006
  • Sodium butyrate (NaBu) is used as an enhancer for the production of recombinant proteins in Chinese hamster ovary (CHO) cells. However, NaBu is well-known for its cytotoxic effect, thereby inducing apoptosis. CHO cells which had been engineered to express a humanised anti-HBV antibody were cultured using serum-free medium, Ex-cell 301. From a seeding density of $2{\times}10^5$ cells/ml, CHO cells grown with serum-free medium reached a maximum cell density of $1.3{\times}10^6$ cells/ml after 9 days in culture and produced a maximal antibody concentration of 130 mg/l after 13 days in culture. In the perfusion culture system, CHO cells producing anti-HBV antibody grown in an 7.5 1 bioreactor seeded with $2{\times}10^5$ cells/ml reached a maximal antibody concentration of 85 mg/1 after 720 h in culture. The addition of 0.3 mM NaBu and lowering culture temperature to $33^{\circ}C$ elongated the culture period to 60 days and increased the production yield by 2-fold, compared to control culture.

Non-Dura Based Intaspinal Clear Cell Meningioma

  • Ko, Jun-Kyeung;Choi, Byung-Kwan;Cho, Won-Ho;Choi, Chang-Hwa
    • Journal of Korean Neurosurgical Society
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    • 제49권1호
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    • pp.71-74
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    • 2011
  • A 34-year-old female patient was presented with leg and hip pain for 6 months as well as voiding difficulty for 1 year. Magnetic resonance imaging revealed a well-demarcated mass lesion at L2-3. The mass was hypo-intense on T1- and T2-weighted images with homogeneous gadolinium enhancement. Surgery was performed with the presumptive diagnosis of intradural extramedullary meningioma. Complete tumor removal was possible due to lack of dural adhesion of the tumor. Histologic diagnosis was clear cell meningioma, a rare and newly included World Health Organization classification of meningioma usually affecting younger patients. During postoperative 2 years, the patient has shown no evidence of recurrence. We report a rare case of cauda equina clear cell meningioma without any dural attachment.