• Journal of Microbiology and Biotechnology
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• 제23권8호
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• pp.1116-1122
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• 2013

Cell line development is the most critical and also the most time-consuming step in the production of recombinant therapeutic proteins. In this regard, a variety of vector and cell engineering strategies have been developed for generating high-producing mammalian cells; however, the cell line engineering approach seems to show various results on different recombinant protein producer cells. In order to improve the secretory capacity of a recombinant tissue plasminogen activator (t-PA)-producing Chinese hamster ovary (CHO) cell line, we developed cell line engineering approaches based on the ceramide transfer protein (CERT) and X-box binding protein 1 (XBP1) genes. For this purpose, CERT S132A, a mutant form of CERT that is resistant to phosphorylation, and XBP1s were overexpressed in a recombinant t-PA-producing CHO cell line. Overexpression of CERT S132A increased the specific productivity of t-PA-producing CHO cells up to 35%. In contrast, the heterologous expression of XBP1s did not affect the t-PA expression rate. Our results suggest that CERT-S132A-based secretion engineering could be an effective strategy for enhancing recombinant t-PA production in CHO cells.

• Journal of Microbiology and Biotechnology
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• 제26권4호
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• pp.684-692
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• 2016

Acute respiratory virus infectious diseases are a growing health problem, particularly among children and the elderly. Much effort has been made to develop probiotics that prevent influenza virus infections by enhancing innate immunity in the respiratory tract until vaccines are available. Lactobacillus plantarum GB-LP2, isolated from a traditional Korean fermented vegetable, has exhibited preventive effects on influenza virus infection in mice. To identify the molecular basis of this strain, we conducted a whole-genome assembly study. The single circular DNA chromosome of 3,284,304 bp was completely assembled and 3,250 protein-encoding genes were predicted. Evolutionarily accelerated genes related to the phenotypic trait of anti-infective activities for influenza virus were identified. These genes encode three integral membrane proteins, a teichoic acid export ATP-binding protein and a glucosamine - fructose-6-phosphate aminotransferase involved in host innate immunity, the nonspecific DNA-binding protein Dps, which protects bacteria from oxidative damage, and the response regulator of the three-component quorum-sensing regulatory system, which is related to the capacity of adhesion to the surface of the respiratory tract and competition with pathogens. This is the first study to identify the genetic backgrounds of the antiviral activity in L. plantarum strains. These findings provide insight into the anti-infective activities of L. plantarum and the development of preventive probiotics.

• 한국건축시공학회지
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• 제16권6호
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• pp.561-569
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• 2016

콘크리트는 현재까지 사용되는 건축재료 양호한 품질을 지니고 있으며, 시멘트의 화학조성이 지표상의 화학조성과 유사하여 가장 쉽게 얻을 있는 재료 중 하나이다. 하지만 약한 결합력과 낮은 결합에너지로 변형에 취약하여 균열이 발생하게 된다. 건조수축 균열 저감을 위해 본 연구에서는 AE제, 감수제, 수축저감제 등을 각각 사용하는 기존연구와 달리 작업의 편리성 향상을 위하여 수축저감제 및 감수제의 혼합 형태인 수축저감형 혼화제(Superplasticizer for Reduction of Shrinkage, 이하 SRS로 칭함)를 이용하였다. 현재 레미콘사에서 일반적으로 사용되고 있는 콘크리트와 모르타르의 배합으로 계획하였으며, SRS 혼입률 변경과 혼화재 치환에 따른 기초적 실험을 진행하였다. 강도와 길이변화율을 고려하였을 때 SRS 혼입률 2%가 적정한 혼입률이라고 판단된다. 또한 대량치환과 노출콘크리트용 혼화제 적용에 관한 연구를 진행할 예정이다.

• Journal of Acupuncture Research
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• 제34권1호
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• pp.23-30
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• 2017

Objectives : In this study, the roles of Interleukin (IL)-4 and Signal transducer and activator of transcription 6 (STAT6), which have been reported to play a role in the pathogenesis of inflammation and cancer, were evaluated in snake venom toxin (SVT)-induced apoptosis. Methods : Inflammation was induced in human HaCaT kerationocytes, by lipopolysaccharide (LPS; $1{\mu}g/mL$) or tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), followed by treatment with SVT (0, 1, or $2{\mu}g/mL$). Cell viability was assessed by MTT assays after 24 h, and the expression of levels of IL-4, STAT6, and the apoptosis-related proteins p53, Bax, and Bcl-2 were evaluated by western blotting. Electro mobility shift assays (EMSAs) were performed to evaluate the DNA binding capacity of STAT6. Results : MTT assays showed that inflammation-induced growth of HaCaT cells following LPS or TNF-${\alpha}$ stimulation was inhibited by SVT. Western blot analysis showed that p53 and Bax, which promote apoptosis, were increased, whereas that of Bcl-2, an anti-apoptotic protein, was decreased in a concentration-dependent manner in LPS- or TNF-${\alpha}$-induced HaCaT cells following treatment with SVT. Moreover, following treatment of HaCaT cells with LPS, IL-4 concentrations were increased, and treatment with SVT further increased IL-4 expression in a concentration-dependent manner. Western blotting and EMSAs showed that the phosphorylated form of STAT6 was increased in HaCaT cells in the context of LPS- or TNF-${\alpha}$-induced inflammation in a concentration-dependent manner, concomitant with an increase in the DNA binding activity of STAT6. Conclusion : SVT can effectively promote apoptosis in HaCaT cells in the presence of inflammation through a pathway involving IL-4 and STAT6.

• 한국식품영양과학회지
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• 제44권10호
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• pp.1583-1587
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• 2015

본 연구에서는 어린감과 성숙감의 탄닌 함량 및 담즙산과의 결합능력을 비교 조사하였다. 또한 사람의 장내 세포인 Caco-2 cell에 어린감과 성숙감 추출물을 처리한 후 고농도의 DCA로 자극시켜 생성되는 NO의 수준과 TEER의 차이를 확인하였다. 탄닌 함량과 담즙산의 결합능력은 성숙감에 비해 어린감에서 유의적으로 높은 것으로 나타났으며 어린감 산성 에탄올 추출물에서 유의적으로 가장 높은 탄닌 함량 및 담즙산 결합능력을 보였다. NO 생성 억제능으로 확인한 염증반응에서도 어린감 산성 에탄올 추출물이 유의적으로 가장 높은 효과를 보였으며 DCA에 의해 유도된 장세포 간극의 느슨함을 유의하게 막아줄 수 있음이 확인되었다. 본 연구 결과를 토대로 어린감의 산성 에탄올 추출물은 고지방 식사를 통해 고농도로 축적되는 담즙산에 의해 형성된 유해한 장내 환경을 건강하게 유지시켜줄 수 있는 기능성 소재로 개발할 가능성이 있음이 확인되었다.

• Journal of Microbiology and Biotechnology
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• 제9권3호
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• pp.235-242
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• 1999

We investigated the growth-inhibitory mechanism of Helicobacter pylori by omeprazole (OMP) and its activated sulfenamide (OAS). Using dithiothreitol (DTT) and 5,5'-dithio-bis[2-nitrobenzoic acid] (DTNB; Ellman's reagent), we first determined the relationship between the binding capacity of these compounds to H. pylori membrane and its significance to membrane P-type ATPase activity. After incubation of the intact H. pylori cells with either OMP or OAS, the residual quantity of free SH-groups on the cell membrane was measured, and, the resulting values were plotted as a function of time. From this experiment, we found that there was a considerable difference in the membrane-binding rates between OMP and OAS. At neutral pH, the disulfide bond formation on H. pylori membrane was completed within 2 min of incubation of the intact cells with OAS. By OMP, however, it was gradually formed, exceeding 10 min of incubation for completion, whereby, the extent of P-type ATPase inhibition appeared to be proportional to the disulfide forming rate. From this data, it was suggested that the disulfide formation might directly affect enzyme activity. Since OMP per se cannot yield a disulfide bond with cysteine, it is predicted that the enzyme inactivation must be caused by the OAS form. Accordingly, we postulated that, under the neutral pH, OMP could be converted to OAS in the course of transport. By extrapolating the inhibitory slopes, we could evaluate K₁ values, relating to their minimal inhibitory concentrations (MICs) for H. pylori growth. In these MIC ranges, H. pylori uptake or vesicular export of nutrients such as peptides were totally prohibited, but their effect in Escherichia coli were negligible. From these observations, we strongly suggest that the P-type ATPase activity is essential for the survival of H. pylori cells in particular.

• Toxicological Research
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• 제28권3호
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• pp.159-164
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• 2012

Allergic skin inflammation such as atopic dermatitis (AD) is characterized by edema and infiltration with various inflammatory cells such as mast cells, basophils, eosinophils and T cells. Thymic stromal lymphopoietin (TSLP) is produced mainly by epidermal keratinocytes, as well as dermal fibroblasts and mast cells in the skin lesions of AD. Omega-3 polyunsaturated fatty acids in fish oil can reduce inflammation in allergic patients. Fermentation has a tremendous capacity to transform chemical structures. The antiinflammatory effects of fish oil have been described in many diseases, but the beneficial effects by which fermented olive flounder oil (FOF) modulates the allergic response is poorly understood. In this study, we produced FOF and tested its ability to suppress the various allergic inflammatory responses. The ability of FOF to modulate the immune system was investigated using a mouse model of AD. The FOF-treated group showed significantly decreased immunoglobulin E (IgE) and histamine in serum. Also, the increased TSLP expression was significantly inhibited in the FOF group; the FOF-treated group was not appreciably different from the hydrocort cream treatment group. In addition, FOF treatment resulted in a smaller spleen size with reduced the thickness and length compared to the induction group. Splenocytes from mice treated with FOF produced significantly less IFN-${\gamma}$, IL-4, T-box transcription factor (T-bet) and GATA binding protein 3 (GATA3) expression compared with the induction group. These results suggest that FOF may be effective in treating the allergic symptoms of AD. 5.

• 생약학회지
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• 제47권1호
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• pp.29-37
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• 2016

Anti-diabetic potential of the leaves of A. elata through the inhibitory activity on PTP1B and ${\alpha}$-glucosidase has not been reported. In this study, the EtOAc fraction of methanolic extract from the leaves of A. elata showed potent inhibitory activity against the PTP1B and ${\alpha}$-glucosidase with $IC_{50}$ value of $96.29{\pm}0.3$ and $264.71{\pm}14.87{\mu}g/mL$, respectively. Three known triterpenoids, oleanolic acid, oleanolic acid-28-O-${\beta}$-D-glucopyranoside and oleanolic acid-3-O-${\beta}$-D-glucopyranoside were isolated from the most active EtOAc fraction. We determined the chemical structure of these triterpenoids through comparisons of published nuclear magnetic resonance (NMR) spectroscopic data. Furthermore, we screened these triterpenoids for their ability to inhibit PTP1B and ${\alpha}$-glucosidase over a range of concentrations ($12.5-50{\mu}M$). All three terpenoids significantly inhibited PTP1B in a concentration dependent manner and oleanolic acid effectively inhibited ${\alpha}$-glucosidase. In addition, these compounds revealed potent inhibitory activity with negative binding energies toward PTP1B, showing high affinity and tight binding capacity in the molecular docking studies. Therefore, the results of the present study clearly demonstrate that A. elata leaves and its triterpenoid constituents might be beneficial in the prevention or treatment of diabetic disease.

• Toxicological Research
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• 제31권2호
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• pp.191-201
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• 2015

We evaluated the inhibitory effect of Pueraria lobata root ethanol extract (PLREE) on lipid accumulation during 3T3-L1 differentiation to adipocytes by measuring the intracellular expression of adipogenic, lipogenic, and lipolytic markers and lipid accumulation. The total polyphenol and flavonoid content of PLREE were 47 and 29 mg/g, respectively. The electron donating capacity of PLREE at $1,000{\mu}g/mL$ was 48.8%. Treatment of 3T3-L1 preadipocytes with 100, 250, or $500{\mu}g/mL$ PLREE for 8 days dose-dependently promoted the differentiation of 3T3-L1 cells. In contrast, the lipid content of PLREE-treated cells was significantly reduced by 7.8% (p < 0.05), 35.6% (p < 0.001), and 42.2% (p < 0.001) following treatment with 100, 250, and $500{\mu}g/mL$ PLREE, respectively, as compared to differentiated control cells. PLREE upregulated peroxisome proliferator-activated receptor ${\gamma}$ mRNA and protein, and sterol regulator element-binding protein-1c mRNA levels, but did not affect CCAAT/enhancer binding-protein ${\beta}$ and ${\alpha}$ mRNA levels. PLREE also downregulated acetyl-CoA carboxylase mRNA and protein, fatty acid synthase (FAS) protein, and leptin mRNA levels, but did not affect FAS mRNA expression. PLREE upregulated adipose triglyceride lipase mRNA and protein expression, and hormone-sensitive lipase (HSL) protein expression, but did not affect HSL mRNA expression. In conclusion, we found that PLREE enhanced adipogenesis, but reduced lipogenesis, resulting in decreased lipid accumulation in 3T3-L1 cells.

• 미생물학회지
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• 제38권1호
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• pp.38-44
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• 2002

국내 분리 침습성 균주중선별된 S. pneumoniae KNIH1156 (type 19F)으로부터 페렴구균의 병원성 인자이며 항원학적으로 다양한 표면단백항원인 pneumococcal surface protein A (PspA)를 분리${\cdot}$정제하였다. 폐렴구균을 CDM-ET배지에서 배양하게 되면 배지내로 PspA가 방출된다는 점과 PspA가 인간의 lactoferrin에 특이적으로 결합한다는 사실을 이용하여 CDM-ET 배지에서 S. pneumoniae KNIH1156 을 배양한 후 배양액을 농축하여 lactoferrinaffinity chromatography에 통과시켜 PspA를 분리, 정제하였다. 정제 후 anti-PspA antiserum으로 PspA를 확인하여 순수분리, 정제되었음을 확인하였으며 또한 인간의 lactoferrin과의 결합능력을 유지하고 있음을 확인하였다. 순수하게 분리하여 정제된 PspA의 면역원성을 확인하기 위하여 ICR mice에 욕강주사하였을 때 $LD_{50}$ 가 $1{\times}10^{7.5}$ CFU/ml께서 $1{\times}10^{10}$ CFU/ml로 약 500배 중가함을 관찰하였다. 따라서 본 실험에서S. pneumoniae KNIH1156 으로부터 분리${\cdot}$ 정제한 PspA가 면역원성과 방어능을 가지고 있음을 확인할 수 있었다.