• Molecules and Cells
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• 제45권4호
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• pp.180-192
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• 2022

Nuclear receptor coactivator 6 (NCOA6) is a transcriptional coactivator of nuclear receptors and other transcription factors. A general Ncoa6 knockout mouse was previously shown to be embryonic lethal, but we here generated liver-specific Ncoa6 knockout (Ncoa6 LKO) mice to investigate the metabolic function of NCOA6 in the liver. These Ncoa6 LKO mice exhibited similar blood glucose and insulin levels to wild type but showed improvements in glucose tolerance, insulin sensitivity, and pyruvate tolerance. The decrease in glucose production from pyruvate in these LKO mice was consistent with the abrogation of the fasting-stimulated induction of gluconeogenic genes, phosphoenolpyruvate carboxykinase 1 (Pck1) and glucose-6-phosphatase (G6pc). The forskolin-stimulated inductions of Pck1 and G6pc were also dramatically reduced in primary hepatocytes isolated from Ncoa6 LKO mice, whereas the expression levels of other gluconeogenic gene regulators, including cAMP response element binding protein (Creb), forkhead box protein O1 and peroxisome proliferator-activated receptor γ coactivator 1α, were unaltered in the LKO mouse livers. CREB phosphorylation via fasting or forskolin stimulation was normal in the livers and primary hepatocytes of the LKO mice. Notably, it was observed that CREB interacts with NCOA6. The transcriptional activity of CREB was found to be enhanced by NCOA6 in the context of Pck1 and G6pc promoters. NCOA6-dependent augmentation was abolished in cAMP response element (CRE) mutant promoters of the Pck1 and G6pc genes. Our present results suggest that NCOA6 regulates hepatic gluconeogenesis by modulating glucagon/cAMP-dependent gluconeogenic gene transcription through an interaction with CREB.

• 한국수의병리학회:학술대회논문집
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• 한국수의병리학회 2001년도 추계학술대회 및 정기총회
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• pp.13-15
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• 2001

발암성시험연구에 사용되고 있는 형질전환 동물들은 랫드와 마우스 등이 있는데, 그 중 c-Ha-ras proto-oncogene 마우스 (ras H2 mice), v-Ha-ras 형질전환 마우스 (Tg.AC mice), pim-1 형질전환 마우스 및 p53 knockout 마우스 등이 발암유발물질에 감수성이 높아 현재 중기발암성시험에 이용되고 있다. (중략)

• Journal of Microbiology and Biotechnology
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• 제27권8호
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• pp.1529-1538
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• 2017

Klebsiella pneumoniae is an opportunistic and clinically significant emerging pathogen. We investigated the relative roles of Toll-like receptor (TLR) 2 and TLR4 in initiating host defenses against K. pneumoniae. TLR2 knockout (KO), TLR4 KO, TLR2/4 double KO (DKO), and wild-type (WT) mice were inoculated with K. pneumoniae. Mice in each group were sacrificed after either 12 or 24h, and the lungs, liver, and blood were harvested to enumerate bacterial colony-forming units (CFU). Cytokine and chemokine levels were analyzed using enzyme-linked immunosorbent assay and real-time PCR, and pneumonia severity was determined by histopathological analysis. Survival was significantly shortened in TLR4 KO and TLR2/4 DKO mice compared with that of WT mice after infection with $5{\times}10^3CFU$. TLR2 KO mice were more susceptible to infection than WT mice after exposure to a higher infectious dose. Bacterial burdens in the lungs and liver were significantly higher in TLR2/4 DKO mice than in WT mice. Serum $TNF-{\alpha}$, MCP-1, MIP-2, and nitric oxide levels were significantly decreased in TLR2/4 DKO mice relative to those in WT mice, and TLR2/4 DKO mice showed significantly decreased levels of $TNF-{\alpha}$, IL-6, MCP-1, and inducible nitric oxide synthase mRNA in the lung compared with those in WT mice. Collectively, these data indicate that TLR2/4 DKO mice were more susceptible to K. pneumoniae infection than single TLR2 KO and TLR4 KO mice. These results suggest that TLR2 and TLR4 play cooperative roles in lung innate immune responses and bacterial dissemination, resulting in systemic inflammation during K. pneumoniae infection.

• 한국식품영양과학회지
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• 제43권11호
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• pp.1627-1634
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• 2014

본 연구는 전통 음청류인 수정과의 세계화 일환으로 대체당을 사용하여 관능성이 탁월한 기능성 수정과 음료를 개발한 후, 개발된 수정과 음료의 항산화 및 산화적 DNA 손상억제 효능을 살펴보고자 하였다. 고콜레스테롤식이 공급으로 고콜레스테롤혈증이 유발된 10주령의 ApoE KO 마우스를 대조군인 설탕물 섭취군(Control), 설탕 첨가 수정과 음료 섭취군(Sucrose), 스티비아 첨가 수정과 음료 섭취군(Stevia), scFOS 첨가 수정과 음료 섭취군(scSFO), 그리고 양성대조군으로 시판 수정과를 섭취시킨 군(Positive control)으로 나누어 실험 식이를 6주간 공급한 후, 혈장 TRAP, 혈장 및 간의 TBARS, 백혈구, 간세포 및 비장세포의 내재적 또는 산화적 스트레스로 인한 DNA 손상 정도를 측정하였다. 모든 군에서 체중 증가량, 식이 섭취량, 조직 무게는 유의적 차이가 없었다. 간 TBARS 농도는 설탕물을 섭취한 대조군에 비해 설탕의 농도를 달리하여 제조한 수정과 음료군인 sucrose군, stevia 첨가군, 그리고 scFOS 첨가군의 TBARS 농도가 유의적으로 감소하여 수정과의 지질과산화 억제 효과를 확인할 수 있었다. 설탕을 줄이는 대신 stevia를 첨가하여 제조한 기능성 수정과 섭취군에서 설탕물만을 섭취한 대조군에 비해 간, 비장세포의 내재적(endogenous) 또는 $H_2O_2$로 유도된 산화적 DNA 손상에 대한 억제 효과가 있는 것으로 나타났다. 백혈구의 DNA 손상의 경우 stevia군에서 대조군에 비해 감소하는 경향을 보여주었으나 통계적 유의성은 없었다. 시판 수정과의 경우 간세포의 내재적 DNA 손상만을 억제하는 효능이 있는 것으로 나타났으며, 설탕만으로 제조한 수정과나 scFOS가 첨가된 수정과 섭취군의 경우 내재적 또는 $H_2O_2$로 유도된 산화적 DNA 손상이 대조군에 비해 감소하는 경향을 보여주었으나 유의성은 없었다. 이상 본 연구의 결과, 고콜레스테롤혈증이 유발된 ApoE KO 마우스에서 수정과의 보충 섭취는 간 지질과산화를 개선시키고 stevia 첨가 수정과의 경우 간 및 비장에서 DNA 손상 억제효과가 있음을 확인할 수 있었다. 본 연구 결과가 수정과의 세계화에 기여할 수 있기를 기대하며, 향후 본 연구를 기반으로 한 분자생물학적 수준의 기전 연구가 필요하다고 사료된다.

• 한국환경성돌연변이발암원학회:학술대회논문집
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• 한국환경성돌연변이발암원학회 2002년도 Current Trends in Toxicological Sciences
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• pp.36-42
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• 2002

Diesel exhaust (DE) has been recognized as a noxious mutagen and/or carcinogen, because its components can form DNA adducts. Mechanisms governing the susceptibility to DE and the efficiency of such DNA adduct formation require clarification. The transcription factor Nrf2 is essential for inducible and/or constitutive expression of a group of detoxification and antioxidant enzymes, and we hypothesized that the nrf2 gene knockout mouse might serve as an excellent model system for analyzing DE toxicity. To address this hypothesis, lungs from nrf2(-/-) and nrf2(+/-) mice were examined for the production of xenobiotic-DNA adducts after exposure to DE (3 $mg/m^{3}$ suspended particulate matter) for 4 weeks. Whereas the relative adduct levels (RAL) were significantly increased in the lungs of both nrf2(+/-) and nrf2(-/-) mice upon exposure to DE, the increase of RAL in the lungs from nrf2(-/-) mice exposed to DE were approximately 2.3-fold higher than that of nrf2(+/-) mite exposed to DE. In contrail, cytochrome P4501Al mRNA levels in the nrf2(-/-)mouse lungs were similar to those in the nrf2(+/-) mouse lungs even after exposure to DE, suggesting that suppressed activity of phase II drug-metabolizing enzymes is important in giving ise to the increased level of DNA adducts in the Nrf2-null mutant mouse subjected to DE. Importantly, severe hyperplasia and accumulation of the oxidative DNA adduct 8-hydroxydeoxyguanosine were observed in the bronchial epidermis of nrf(-/-) mite following DE exposure. These results demonstrate the increased susceptibility of the nrf2 germ line mutant mouse to DE exposure and indicate the nrf2 gene knockout mouse nay represent a valuable model for the assessment of respiratory DE toxicity.

• 대한약학회:학술대회논문집
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• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.1
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• pp.110.1-110.1
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• 2002

• 대한약학회:학술대회논문집
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• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.1
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• pp.168.1-168.1
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• 2002

• Laboraroty Animal Research
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• 제34권4호
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• pp.279-287
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• 2018

Placenta specific 8 (PLAC8, also known as ONZIN) is a multi-functional protein that is highly expressed in the intestine, lung, spleen, and innate immune cells, and is involved in various diseases, including cancers, obesity, and innate immune deficiency. Here, we generated a Plac8 knockout mouse using the CRISPR/Cas9 system. The Cas9 mRNA and two single guide RNAs targeting a region near the translation start codon at Plac8 exon 2 were microinjected into mouse zygotes. This successfully eliminated the conventional translation start site, as confirmed by Sanger sequencing and PCR genotyping analysis. Unlike the previous Plac8 deficient models displaying increased adipose tissue and body weights, our male Plac8 knockout mice showed rather lower body weight than sex-matched littermate controls, though the only difference between these two mouse models is genetic context. Differently from the previously constructed embryonic stem cell-derived Plac8 knockout mouse that contains a neomycin resistance cassette, this knockout mouse model is free from a negative selection marker or other external insertions, which will be useful in future studies aimed at elucidating the multi-functional and physiological roles of PLAC8 in various diseases, without interference from exogenous foreign DNA.

• BMB Reports
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• 제45권3호
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• pp.153-158
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• 2012

Geft is a guanine nucleotide exchange factor, which can specifically activate Rho family of small GTPase by catalyzing the exchange of bound GDP for GTP. Geft is highly expressed in the excitable tissue as heart and skeletal muscle and plays important roles in many cellular processes, such as cell proliferation, migration, and cell fate decision. However, the in vivo role of Geft remains unknown. Here, we generated a Geft conditional knockout mouse by flanking exons 5-17 of Geft with loxP sites. Cre-mediated deletion of the Geft gene in heart using Mef2c-Cre transgenic mice resulted in a dramatic decrease of Geft expression. Geft knockout mice develop normally and exhibit no discernable phenotype, suggesting Geft is dispensable for the development of the second heart field in mouse. The Geft conditional knockout mouse will be a valuable genetic tool for uncovering the in vivo roles of Geft during development and in adult homeostasis.

• IMMUNE NETWORK
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• 제2권4호
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• pp.233-241
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• 2002

Background: Monoclonal antibodies (mAb) of rodent origin are produced with ease by hybridoma fusion technique, and have been successfully used as therapeutic reagents for humans after humanization by genetic engineering. However, utilization of these antibodies for therapeutic purpose has been limited by the fact that they act as immunogens in human body causing undesired side effects. So far, there have been several attempts to produce human mAbs for effective in vivo diagnostic or therapeutic reagents including the use of humanized xenomouse that is generated by mating knockout mice which lost Ig heavy and light chain genes by homologous recombination and transgenic mice having both human Ig heavy and light gene loci in their genome. Methods: Genomic DNA fragments of mouse Ig heavy and light chain were obtained from a mouse brain ${\lambda}$ genomic library by PCR screening and cloned into a targeting vector with ultimate goal of generating Ig knockout mouse. Results: Through PCR screening of the genomic library, three heavy chain and three light chain Ig gene fragments were identified, and restriction map of one of the heavy chain gene fragments was determined. Then heavy chain Ig gene fragments were subcloned into a targeting vector. The resulting construct was introduced into embryonic stem cells. Antibiotic selection of transfected cells is under the progress. Conclusion: Generation of xenomouse is particularly important in medical biotechnology. However, this goal is not easily achieved due to the technical difficulties as well as huge financial expenses. Although we are in the early stage of a long-term project, our results, at least, partially contribute the successful generation of humanized xenomouse in Korea.