• 한국식품과학회지
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• 제44권5호
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• pp.600-606
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• 2012

콩을 소재로한 전통 발효식품으로부터 혈전용해능이 뛰어난 균주를 분리하였으며, B. subtilis로 동정되었다. 따라서 이를 B. subtilis IDCC 9204(특허균주기탁: KCTC-11471 BP), 그 혈전용해 효소는 NK-IL9204로 명명하였다. B. subtilis IDCC 9204가 생산하는 고역가의 NK-IL9204를 단백질 분석법에 기초하여 분석한 결과, 분자량은 27.7 kDa의 homogenous enzyme으로 확인되었다. 또한 기존에 알려진 일본의 발효식품인 낫도 유래의 B. subtilis var. natto가 생산하는 nattokinase 와의 sequence 분석을 진행한 결과, 99.5% homology가 일치하는 serine protease계열의 nattokinase로 확인되었다. 그러나 NK-IL9204는 물리 화학적인 조건에서 B. subtilis var. natto가 생산하는 nattokinase와 다소 차이를 나타내었으며 본 실험에서는 B. subtilis var. natto가 생산하는 nattokinase보다 상대적으로 높은 열 안정성과 pH 안정성을 나타내었다. In vitro 실험에서 NK-IL9204는 최적 반응온도 $40^{\circ}C$, 열 안정성은 $90^{\circ}C$까지 효소활성을 유지하였으며, 최적 반응 pH는 pH 8로 알칼리-혈전용해효소의 특성을 나타내었으며, 약산성에서 강알칼리 영역까지 넓은 pH 구간 안정성을 갖는 것이 특징이다. NKIL9204의 in vivo에서의 효능과 생체 내 안정성을 동물실험을 통해 확인한 결과, 생체 내에서도 혈전용해효소의 활성이 소실되지 않고 유지되며, 혈전분해와 관련된 생체 내 인자들을 활성화시키는 역할을 하는 특징을 갖는다. NK-IL9204는 30,000 FU/g 이상의 고역가를 달성하여 산업적 측면에서 생산성도 확보함으로써 수입의존적 원료를 국산화할 수 있을 것으로 예상된다.

• Tuberculosis and Respiratory Diseases
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• 제70권2호
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• pp.132-138
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• 2011

Background: Maspin (mammary serine protease inhibitor) is a member of the serpin superfamily. A few studies have examined the role of maspin in tumor suppression of non-small cell lung cancer (NSCLC); however, its role in the development and progression of NSCLC still remains controversial. We evaluated the immunohistochemical expression of maspin in order to elucidate its clinical significance in NSCLC. Methods: We analyzed 145 patients with pathologically confirmed NSCLC, including 66 cases of squamous cell carcinomas (SCCs) and 79 cases of adenocarcinomas (ADCs). We performed a immuno-histochemical stain with maspin and PCNA (proliferating cell nuclear antigen) using tissue microarray blocks. Results: There were 108 men and 37 women in the study population. The mean age of patients in the study was 63.7 years (range, 40.0~82.0; median, 65.0). The proportion of maspin expression was significantly higher in SCCs (52/66, 78.8%; p<0.01) than in ADCs (17/79, 21.5%; p<0.01). Maspin expression was not associated with PCNA (p=0.828), lymph node involvement (p=0.483), or tumor stage (p=0.216), but showed correlation with well-to-moderate tumor differentiation (p=0.012). There was no observed correlation between maspin expression and survival with NSCLC (p=0.218). Conclusion: The present study suggests that maspin expression was significantly higher in SCCs than in ADCs and was associated with low histological grade. However, maspin expression was not an independent factor to predict a prognosis in NSCLC.

• 생명과학회지
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• 제18권10호
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• pp.1348-1354
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• 2008

mi transcription factor (MITF)는 비만세포의 분화를 조절하는 중요한 전사인자이다. 특히 MITF는 결합조직형 비만세포에서 일반적으로 발현하는 비만세포 특이적 세린 단백분해효소의 일종인 mMCP-6 유전자의 전사를 조절한다. 본 연구는 마우스 골수유래 배양비만세포에서 mMCP-6 유전자의 전사를 조절하는 MITF이형체를 규명하였다. MITF 이형체들의 발현은 RT-PCR로 확인하였다. IL-3존재 하에서 배양한 점막형 비만세포들은 MITF-A,-E, -H, -Mc 등이 발현하였다. 반면에 SCF존재 하에서 배양한 결합조직형 비만세포들은 MITF-A가 발현하였다. MITF이형체를 과발현시키면 NIH-3T3 세포에서 mMCP-6 promoter를 통한 luciferase 활성을 증가시키고, MC/9 비만세포주에서는 증가된 mMCP-6발현을 유도하였다. 더불어 비만세포에서의 mMCP-6 발현은 MITF-A 고갈로 인하여 유의적으로 억제되었다. MITF-A의 전사활성과 DNA결합은 MITF-E, -H, -Mc 등의 타 이형체들의 결과와 유사하였다. 따라서 본 연구의 결과들은 MITF-A가 마우스 결합조직형 비만세포에서 발현하여 mMCP-6 전사를 조절하는 중요한 이형체임을 제시한다.

• 대한수의학회지
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• 제43권3호
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• pp.383-392
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• 2003

Insulin-like growth factors (IGFs) and their binding proteins (IGFBPs) are important regulators on the development of maternal tissues during pregnancy. This study was performed to examine the relationship between maternal IGFs/IGFBPs system (i.e: IGF-I, II, their receptors, and IGFBPs) in pre- and post-partum rats. The liver and kidney are important organs for the synthesis of IGFs and IGFBPs in adults. The levels of materanal IGFs and IGFBPs in serum, liver, and kidney were examined at 14 and 21 days of gestation and at 3, 7, 11, and 14 days after birth. The expression of IGFs and their receptors mRNA was also examined in fetal and maternal rat liver, kidney. IGF-I concentrations in maternal serum and liver were decreased during pregnancy. However, IGF-I concentration in maternal kidney was increased, having maximal effect at 14 days of gestation. IGF-I concentrations were decreased in serum, liver, and kidney of postpartum rat, compared to control (p < 0.05). On the other hand, IGF-II concentrations in serum, liver, and kidney were increased during pregnancy (p<0.05) and gradually decreased to control level in postpartum period. The levels of IGFBP-3 and IGFBP-2 are expressed in serum, liver, and kidney. However, IGFBP-3 is mainly expressed in serum and liver, and IGFBP-2 in kidney. The levels of IGFBP-3 and IGFBP-2 in maternal serum were markedly decreased during pregnancy and gradually recovered to control level during postpartum period by western ligand blotting. However, there was no change of IGFBP-3 and IGFBP-2 levels by western immunoblotting. The levels of IGFBP-3 and IGFBP-2 in maternal liver and kidney also showed the same pattern of serum, although the main IGFBP is different. In normal rat serum, IGF-I 150 kDa and 50 kDa carrier proteins were detected. The level of IGF-I 150 kDa carrier proteins in pregnant rat was decreased compared to normal rat, but that of 50 kDa carrier proteins was increased. IGFBP-3 protease activity was identified in pregnant rat serum and maternal placenta, and it was inhibited by EDTA ($Ca^{2+}$ chelating agent) and aprotinin (serine proteinase inhibitor). Taken together, these results suggest that the changes of IGFs and IGFBPs in maternal rats are regulated by liver and kidney IGFs and their receptors mRNA during the pregnancy.

• 한국식품영양과학회지
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• 제22권4호
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• pp.458-464
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• 1993

전보에 이어, 정제한 혈합육어 trypsin에 대하여 효소적 성질 및 열 역학적 성질에 관하여 비교 검토한 내용을 요약하면 다음과 같다. 이들 혈합육어 trypsin은 BA-p-NA와 SP-p-NA 같은 amide기질 중 BA-p-NA기질만에 대하여, 그리고 ATEE, BAEE, BTEE 및 TAME 등의 ester 기질 중에서는 BAEE와 TAME에 대하여만 현저한 활성을 보였다. 이들 효소는 antipain, leupeptin, DFP, TLCK, SBTI 등 화학약제와 금속이온 Cu$^{2＋}$ 및 Hg$^{2＋}$에 의하여서는 그 활성이 현저히 저해를 받았으나, $Mg^{2＋}$에 의하여서는 부활하였다. 이 효소들은 모두 5$0^{\circ}C$ 이상의 온도에서는 불안정하였으며, 날개다랭이의 trypsin이 온도 변화에 대하여 가장 안정하였다. 그리고, 활성화에너지는 멸치 trypsin이 13.91㎉/mo1e, 고등어 trypsin A는 11.61㎉/ mole, 고등어 trypsin B는 8.43㎉/mo1e, 황다랭이 trypsin은 4.35㎉/mole, 그리고 날개다랭이 trypsin은 3.76㎉/mole 이었다.

• Biomolecules & Therapeutics
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• 제15권1호
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• pp.16-26
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• 2007

Tissue plasminogen activator (tPA) is a serine protease catalyzing the proteolytic conversion of plasminogen into plasmin, which is involved in thrombolysis. During last two decades, the role of tPA in brain physiology and pathology has been extensively investigated. tPA is expressed in brain regions such as cortex, hippocampus, amygdala and cerebellum, and major neural cell types such as neuron, astrocyte, microglia and endothelial cells express tPA in basal status. After strong neural stimulation such as seizure, tPA behaves as an immediate early gene increasing the expression level within an hour. Neural activity and/or postsynaptic stimulation increased the release of tPA from axonal terminal and presumably from dendritic compartment. Neuronal tPA regulates plastic changes in neuronal function and structure mediating key neurologic processes such as visual cortex plasticity, seizure spreading, cerebellar motor learning, long term potentiation and addictive or withdrawal behavior after morphine discontinuance. In addition to these physiological roles, tPA mediates excitotoxicity leading to the neurodegeneration in several pathological conditions including ischemic stroke. Increasing amount of evidence also suggest the role of tPA in neurodegenerative diseases such as Alzheimer's disease and multiple sclerosis even though beneficial effects was also reported in case of Alzheimer's disease based on the observation of tPA-induced degradation of $A{\beta}$ aggregates. Target proteins of tPA action include extracellular matrix protein laminin, proteoglycans and NMDA receptor. In addition, several receptors (or binding partners) for tPA has been reported such as low-density lipoprotein receptor-related protein (LRP) and annexin II, even though intracellular signaling mechanism underlying tPA action is not clear yet. Interestingly, the action of tPA comprises both proteolytic and non-proteolytic mechanism. In case of microglial activation, tPA showed non-proteolytic cytokine-like function. The search for exact target proteins and receptor molecules for tPA along with the identification of the mechanism regulating tPA expression and release in the nervous system will enable us to better understand several key neurological processes like teaming and memory as well as to obtain therapeutic tools against neurodegenerative diseases.

• Journal of Ginseng Research
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• 제44권6호
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• pp.799-807
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• 2020

Background: The skin acts as a barrier to protect organisms against harmful exogenous agents. Compound K (CK) is an active metabolite of ginsenoside Rb1, Rb2 and Rc, and researchers have focused on its skin protective efficacy. In this study, we hypothesized that increased expression of the serine protease inhibitor Kazal type-5 (SPINK5) may improve skin barrier function. Methods: We screened several ginsenosides to increase SPINK5 gene promoter activity using a transactivation assay and found that CK can increase SPINK5 expression. To investigate the protective effect of CK on the skin barrier, RT-PCR and Western blotting were performed to investigate the expression levels of SPINK5, kallikrein 5 (KLK5), KLK7 and PAR2 in UVB-irradiated HaCaT cells. Measurement of transepidermal water loss (TEWL) and histological changes associated with the skin barrier were performed in a UVB-irradiated mouse model and a 1-chloro-2,4-dinitrobenzene (DNCB)-induced atopic dermatitis-like model. Results: CK treatment increased the expression of SPINK5 and decreased the expression of its downstream genes, such as KLKs and PAR2. In the UVB-irradiated mouse model and the DNCB-induced atopic dermatitis model, CK restored increased TEWL and decreased hydration and epidermal hyperplasia. In addition, CK normalized the reduced SPINK5 expression caused by UVB or DNCB, thereby restoring the expression of the proteins involved in desquamation to a level similar to normal. Conclusions: Our data showed that CK contributes to improving skin-barrier function in UVB-irradiated and DNCB-induced atopic dermatitis-like models through SPINK5. These results suggest that therapeutic attempts with CK might be useful in treating barrier-disrupted diseases.

• Journal of Acupuncture Research
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• 제25권1호
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• pp.139-154
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• 2008

목적 : 홍화(紅花)는 활혈거어(活血祛瘀), 해독지통(解毒止痛)의 효능이 있어 관절염, 동맥경화(動脈硬化), 종양(腫瘍), 월경부조(月經不調), 뇌혈전(腦血栓)에 사용되어 왔다. 이에 홍화약침액(紅花藥鍼液)의 분자생물학적 효능 분석을 하고자 Lipopolysaccharide(LPS)로 염증을 유발한 RAW 264.7 cell의 유전자(遺傳子) 발현(發顯)에 미치는 영향을 Microarray를 통하여 관찰하였다. 방법 : RAW cell을 배양하고 홍화약침액(紅花藥鍼液)의 세포 독성을 확인한 후 (1) LPS, (2) 홍화약침액(紅花藥鍼液), (3) 홍화약침액(紅花藥鍼液)과 LPS를 처치했을 때의 유전자 발현양상을 microarray를 이용하여 관찰하였다. 대조군에 비해 2배 이상 발현의 차이가 있는 경우를 유의한 것으로 보았다. 결과 : 8,170개의 유전자 중 (1) LPS를 처치하였을 경우 35개의 유전자에서 발현이 상승되었고, (2) 홍화약침액(紅花藥鍼液)을 처치하였을 경우 11개의 유전자에서 발현이 상승되고 53개의 유전자에서 발현이 억제되었으며, (3) 홍화약침액(紅花藥鍼液)과 LPS를 동시에 처치하였을 경우에는 47개의 유전자에서 발현이 상승되었고 11개의 유전자에서 발현이 억제되었다. LPS 자극으로 발현이 상승되었지만 홍화약침액(紅花藥鍼液)을 처치할 때 발현이 억제되는 유전자는 SUMO1/sentrin specific protease 7(SENP7), Serine(or cysteine) proteinase inhibitor, clade B(ovalbumin), member 7(SERPINB7), M-phase phosphoprotein, mpp8(HSMPP8), Glycogenin 2(GYG2), InaD-like(Drosophila)(INADL), Copine III(CPNE3), Loss of heterozygosity, 11, chromosomal region 2, gene A(LOH11CR2A), Chromosome 9 open reading frame 33(SHC3), NADH dehydrogenase(ubiquinone) 1 beta subcomplex, 2, 8kDa(NDUFB2)로 9개가 있었다. 요약 : 홍화약침액(紅花藥鍼液)이 LPS로 염증을 유발시킨 RAW 264.7 cell의 유전자 발현에 미치는 영향을 Microarray를 통해 분석하였다. 홍화약침액(紅花藥鍼液)이 LPS로 발현을 항진시킨 35개의 유전자 중 9개를 효과적으로 억제하는 것을 확인하여 염증 치료 기전을 시사하는 유용한 자료를 얻을 수 있었으며 홍화약침액(紅花藥鍼液)이 발현을 항진시킨 유전자들을 통해 혈관생성과 종양억제 등 보다 넓은 범위에 대한 연구가 가능할 것으로 사료된다.

• 생명과학회지
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• 제17권8호통권88호
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• pp.1115-1120
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• 2007

본 연구에서는 다섯 종류의 phytochemical (resveratrol, genistein, epicatechin gallate, diaIly disulfide, caffeic acid phenetyl ester)과 sulindac sulfide가 암 억제 단백질 p53을 유도할 수 있는지에 대해 연구하였다. 처리한 모든 phytochemical에 의해 p53 단백질의 발현이 강하게 유도된 반면, sulindac sulfide에 의해서는 p53 단백질이 유도되지 않았다. 처리한 phytochemical 중 포도껍질이나 와인에 많이 들어있는 resveratrol에 의해 p53 단백질이 농도의존적 혹은 처리시간 의존적으로 증가 발현되는 것을 확인하였다. 암 억제 단백질인 p53 하위 단계의 유전자들만 집적되어 있는 membrane microarray를 이용하여 실험을 수행한 결과, 25개의 유전자가 up-regulation 된 반면, 2개의 유전자가 down-regulation 되는 것을 확인하였다. Up-regulation 되는 유전자중 4개를 선택하여, RT-PCR을 수행한 결과 모두 membrane microarray 실험의 결과와 일치하였다. 게다가 p53 null인 HCT116 세포주를 이용한 RT-PCR을 통하여 TSP-1 유전자의 발현은 p53 의존적이지 않은 반면, MASPIN 유전자는 p53 의존적임을 확인하였다. 이러한 연구 결과는 resveratrol에 의한 화학적 암 예방법의 분자생물학적 기전을 이해하는데 도움을 줄 것으로 기대된다.

• Archives of Pharmacal Research
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• 제28권9호
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• pp.1002-1012
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• 2005

5-Substituted 4-oxo-2-thioxo-1,,2,3,4-tetrahydropyrimidine were synthesized by interaction of 4­oxo-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-sulfonylhydrazide with some aldehydes to give the corresponding Schiff-bases, which after cyclization gave corresponding thiazolidinones. For some of the thiazolidinones, Mannich bases reaction was carried out. All the derivatives were tested for their possible inhibitory effect on Schistosoma mansoni cercarial elastase (CE). Only, N'-(4-methylbenzyledine)-4-oxo-2-thioxo-1,2 ,3,4-tetrahydropyrimidine-5-sulfonylhydrazide was found to have potent inhibitory effect on the CE activity with $IC_{50} = 264{\mu}M.$ Upon its use as a paint for mice tails before infection with S. mansoni cercariae, the compound formulated in jojoba oil caused a significant reduction ($93\%$; P-value = 0.0002) in the worm burden. IgG & IgM in mice sera were measured by using several S. mansoni antigens by ELISA. Sera from treated infected mice (TIM) 2, 4, and 6 weeks (W) post infection (PI) showed 1.2 folds lower, 1.2 folds higher, 1.7 folds lower IgM reactivity against soluble cercarial antigenic preparation (CAP), respectively, when compared with sera collected from infected untreated mice (IUM). Sera from TIM 2, 4, and 6WPI showed 1.3, 1.6, and 1.7 folds higher IgG reactivity, respectively against CAP than the IgG reactivity from IUM. Sera from TIM 2, 4 and 6WPI showed 1.5, 1.2 folds lower and 1.4 folds higher IgM reactivity, respectively against soluble worm antigenic preparation (SWAP) when compared with sera collected from IUM. Sera from TIM 2, 4, and 6WPI showed 1.4, 1 folds lower and 1 fold higher IgG reactivity, respectivley to SWAP when compared with sera from IUM. Sera from TIM 2, 4, and 6WPI had generaly lower IgM and IgG reactivities against soluble egg antigen (SEA) when compared with sera from IUM.