• 대한화장품학회지
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• 제35권1호
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• pp.19-25
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• 2009

피부주름 개선에는 콜라겐(collagen)뿐만 아니라 탄력섬유인 엘라스틴(elastin) 등도 기여하는 것으로 보고되고 있다. 더 나아가 자외선에 의하여 사람의 피부에서 광노화 현상이 나타나며 자외선 조사 후 엘라스타제(elastase)의 활성이 증가하기 때문에 엘라스타제의 활성증가는 자외선에 의한 피부 탄성도의 감소 및 주름 생성의 주요원인으로 생각된다. 따라서 본 연구에서는 피부주름 생성에 영향을 미치는 엘라스타제의 활성을 측정하기 위한 모델을 마련하기 위하여 시판되는 두 가지 엘라스타제, 돼지 췌장 엘라스타제(porcine pancreatic elastase)와 사람 호중구 엘라스타제(human neutrophil elastase)를 사용하였으며 다음 세 가지는 정상 사람 섬유아세포(normal primary human fibroblasts), 쥐의 3T3 섬유아세포주(3T3 mouse fibroblasts), 사람의 CCD-25Sk 섬유아세포주(CCD-25Sk human fibroblasts)로부터 elastase를 준비하여 사용하였다. 준비된 5가지 효소의 농도에 기질의 농도 및 배양시간에 따르는 효소의 활성을 비교 평가하였다. 양성대조군으로 사용한 phosphoramidon이 normal human primary fibroblast elastase와 CCD-25Sk fibroblast elastase의 활성을 유의성 있게 억제한 반면에 porcine pancreatic elastase에는 별다른 영향을 미치지 못하였다. 따라서 주름 개선 후보물질 탐색을 위한 엘라스타제의 선정에는 신중한 고려가 필요할 것으로 사료된다.

• Journal of Microbiology and Biotechnology
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• 제16권8호
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• pp.1320-1324
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• 2006

The pancreatic and neutrophil elastases are associated with several illnesses including lung and vascular diseases, various cancers, and pancreatitis. The development of a potent and specific inhibitor to the elastases could lead to new therapies. In this study, an agar diffusion method was modified to include a substrate-dye conjugate (Elastin-Congo red) as a substrate of elastase and an indicator of elastase inhibitory activity. The Elastin-Congo red agar plates consisted of 0.1 % Elastin-Congo red and 2.5% agar. The elastase and elastase inhibitors were simultaneously loaded into wells, ultimately resulting in halo formations in which the halo diameter decreased as the concentration of elastase inhibitor increased. The concentration of elastase inhibitor in the samples, therefore, was inversely proportional to the halo diameters. This simplified method provided an excellent correlation with the standard microplate technique, which uses a chromogenic substrate. The concentration of elastase inhibitor obtained from the culture supernatant of a recombinant elastase inhibitor produced by the yeast Pichia pastoris was easily determined. This study has established a simple modified and inexpensive agar diffusion method that is potentially useful for the identification, quantification, and screening of new elastase inhibitors.

• Natural Product Sciences
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• 제9권4호
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• pp.278-285
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• 2003

The serine proteases such as human leukocyte elastase (HLE) and porcine pancreatic elastase (PPE) are classified in the chymotrypsin family, and possibly the most destructive enzymes having the ability to degrade virtually all of the connective components in the body. In the present study, the extracts of water and methanol of Korean mistletoe (Viscum album var. coloratum) inhibited significantly the PPE activity. The fractions eluated on Amberlite XAD-2 from methanol extract were further purified on the repeated $SiO_2$ column chromatography and the fractions A, B and C were eluated. The fractions A, B and C at 3 mg/ml inhibited significantly the PPE activity up to 66%, 95% and 85%, respectively. In conclusion, the fraction A assumed as lignans or phenylpropanes, and fraction B and C assumed as triterpenoids showed the PPE inhibitory effects on the PPE and that these compounds in mistletoe may be used for treatment of pathological processes such as age-dependent tissue loss or inflammation.

• 대한약리학회지
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• 제16권2호
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• pp.15-24
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• 1980

[${\alpha}$]-Amylase catalyses the hydrolysis of starch, glycogen, and related poly- and oligosac-charide by random cleavage of ${\alpha}$-D-(l-4) glucan linkage. In man large amounts of amylase are secreted into the digestive tract by the salivary and exocrine pancreatic gland, minimal amount being produced also in other tissues. It has been known that ${\alpha}$-amylase exists in multiple molecular forms, isoenzyme which can be separated from each other because of difference in their physicochemical properties. By using various methods, several groups of investigator have separated the many isoenzyme in serum, saliva and pancreatic juice. Furthermore, changes of the normal serum isoenzyme pattern is diagnostically useful even when the total serum enzyme activity is noninformative, such as the clinical use of isoenzyme of serum lactate dehydrogenase. Procarboxypeptidase-A which is one of the pancreatic enzymes is also present as isoenzymes. Four forms of procarboxypeptidase-A haye been found in the bovine enzyme and three forms of the porcine enzyme. In human pancreatic juice four forms of procarboxypeptidase-A isoenzyme were found by isoelectric focusing method. Recently, the so-called isoamylase analysis was developed for the diagnostic use of amylase in pancreatic diseases. In alcohotic patients, the serum concentration of pancreatic isoamylase is subnormal and this lowered activity provides strong evidence for pancreatic exocrine insufficiency. The purpose of this study was to elucidate the variations of the isoenzyme of amylase and procarboxypeptidase-A in serum, saliva and pancreatic juice of the experimental animals. The results are as follow. 1) Three main forms of isoenzyme of amylase by isoelectric focusing were found in pancreatic juice of normal rabbit. However, many new bands were appeared in the pancreatic juice of cholic acid administered animal intravenously while the infusion of cholic acid or elastase into pancreatic duct produced the decrease of number of the fractions on the isoelectric focusing. In the case of serum isoenzyme from normal animal, two major and a few minor isoamylases were observed. By injecting alcohol intravenousely the fractions of serum isoamylase were significantly decreased and in contrary to the pattern in the pancreatic juice the infusion of cholic acid or elastase into pancreatic duct exhitited a significant decrease of the isoenzyme of amylase fractions. In saliva from normal animal three main isoamylase were produced of the administration of alcohol. 2) In the case of procarboxypeptidase-A isoenzyme, two major fractions which have isoelectric point at 6.2 and 6.4 and other two minor bands were observed in the pancreatic juice of normal rabbit. By the treatment of the juice with trypsin, only one band was produced on the isoelectric focusing. No procarboxypeptidase was appeared on the electrofocusing by the infusion of cholic acid or phospholipase A into the pancreatic duct of rabbit. However, a single major fraction of procarboxypeptidase-A was appeared at 3 hr after simple ligation of the pancreatic duct. No significant changes were observed in the juice of the alcohol or cholic acid administered group.

• 대한화장품학회지
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• 제26권1호
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• pp.163-186
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• 2000

Elastase (EC 3. 4. 4. 7)는 구조 단백질인 fibronectin, collagen 그리고 elastin을 포함한 다양한 단백질들을 분해한다. Elastase는 생체 염증 방어의 중요한 성분이며, $\alpha$$_1$-proteinase inhibitor ($\alpha$$_1$PI)와 같은 생체 내부에 있는 단백질 분해 효소 저해제들에 의해 조절된다. 그러나, 만성 염증의 결과, elastase와 $\alpha$$_1$PI 사이의 균형이 elastase 쪽으로 기울어지면 조직 파괴가 일어나게 된다. 본 연구자들은 이미 피부의 진피내에서 백혈구로부터 과다하게 분비되는 elastase를 저해하는 물질을 찾기 위하여, 150 여가지의 한방식물 추출물에 대해 elastase 저해활성을 스크리닝 (screening) 하여, 빈랑 (Areca catecheu) 추출물이 가장 높은 저해활성을 보여줌을 확인하였다. 본 연구에서는 유효성분을 찾기 위하여 빈랑을 90% 에탄올로 추출한 다음, 다양한 용매를 사용하여 분획하였고, 각 분획에 대해 elastase 저해활성을 측정하였다. 이 중 저해 활성이 뛰어난 분획은 silica gel 컬럼 크로마토그래피, Preparative TLC 그리고 역상 HPLC를 실시하였다. 역상 HPLC에서 elastase 저해 활성이 있는 peak은 성분을 확인할 수 있는 여러 가지 정색 반응과 UV, IR 스펙트럼을 통하여 phenol 성 물질임을 확인하였다. 빈랑으로부터 정제한 phenol 성 물질은 돼지 췌장 elastase (Porcine pancreatic elastase : PPE)와 사람 백혈구 elastase (Human neutrophil elastase ; HNE)에 대한 $IC_{50}$/ 값이 각각 26.9 $\mu\textrm{g}$/$m\ell$과 60.8 $\mu\textrm{g}$/$m\ell$로서, 기존에 elastase의 저해제로 알려진 oleanolic acid (각각 76.5 $\mu\textrm{g}$/$m\ell$, 219.2 $\mu\textrm{g}$/$m\ell$)와 ursolic acid (각각 31.0 $\mu\textrm{g}$/$m\ell$, 118.6 $\mu\textrm{g}$/$m\ell$)에 비하여 더 우수한 저해 활성을 나타내었으며, PPE와 HNE에 대하여 기질과 경쟁적으로 반응함을 알 수 있었다. 또한 빈랑으로부터 정제한 phenol 성 물질이 염증 과정 중에 발생하는 free radical을 소거할 수 있는지를 조사한 결과, free radical을 50% 소거시키는 농도값이 6 $\mu\textrm{g}$/$m\ell$로 표준물질인 비타민 C (19 $\mu\textrm{g}$/$m\ell$)나 butylated hydroxyl toluene (18.5 $\mu\textrm{g}$/$m\ell$)보다 우수한 효과를 나타내었다. 그리고 mast cell내에서 활성화되며 hyaluronic acid를 분해하는 hyaluronidase에 대한 저해를 조사한 결과, $IC_{50}$/ 값이 210 $\mu\textrm{g}$/$m\ell$로서 효과적인 저해 활성을 나타내었다 따라서, 본 연구에서 빈랑으로부터 분리한 phenol 성 물질은 피부 결합 조직을 구성하는 단백질들을 보호함으로써 피부 노화를 억제하는 효과를 나타낼 것으로 생각된다.

• 한방안이비인후피부과학회지
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• 제23권1호
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• pp.135-148
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• 2010

Recently, the demands for the effective and safe depigmentating and anti-aging agents of the skin have increased due to the medical, pharmaceutical and cosmetic reasons. The aim of this study is to search Galla Rhois (GR) inhibitory activity against aging process of skin in vitro. GR was screened for their inhibitory activity against elastase. GR (final concentration 1 mg/ml) appeared over 40% of inhibition of elastase activity. So we investigated anti wrinkle effects of GR look through MMP-1 inhibition activity. Also Extracts of GR showed higher anti-tyrosinase activity than arbutin (final concentration 1 mg/ml). These results suggest that GR could be potential sources of anti-aging and whitening effects for the skin. Objective : Galla Rhois extracts, Elastase (pancreatic from porcine pancreas), N-Succinyl-(Ala)3-p-nitroanilide, Ursolic acid, mushroom tyrosinase, L-tyrosine, arbutin, Amicon Ultra, Hand Hold UV lamp, MMP-1 primary antibody, MMP-1 secondary antibody Result : 1. GR appeared over 40% of elastase inhibitory activity. 2. GR had MMP-1 inhibition activity. 3. GR showed higher anti-tyrosinase activity than arbutin. Conclusion : According to above results, it is supposed that Galla Rhois could be strong potential sources of anti-aging and whitening effects for the skin.

• BMB Reports
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• 제33권2호
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• pp.112-119
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• 2000

Three kinds of serine protease inhibitors, members of the Bowman-Birk trypsin inhibitor, were purified from Dolichos lablab seeds and named Dolichos protease inhibitor 1, 2 and 3 (DI-1, DI-2 and DI-3), respectively. Each inhibitor showed a single band with gel mobility at around 15.9, 12.1 and 14.6 kDa on 20% SDS-PAGE under reducing conditions. To characterize inhibitory specificity, the inhibition constant (Ki) for these inhibitors was measured against several known serine proteases. All three Dolichos protease inhibitors (DI-1, DI-2 and DI-3) inhibited the activity of trypsin and plasmin, but had no effect on thrombin and kallikrein (either for human plasma kallikrein or for porcine pancreas kallikrein). DI-1 inhibited chymotrypsin most effectively (Ki = $3.6{\times}10^{-9}\;M$), while DI-2 displayed inhibitory activity for porcine pancreatic elastase (Ki = $6.2{\times}10^{-8}\;M$). Pre-treatment of the 33 mg/kg of DI-mixture (active fractions from $C_{18}$ open column chromatography that included DI-1, DI-2 and DI-3) inhibited the induction of pseudomonal elastase-induced septic hypotension and prevented an increase in bradykinin generation in pseudomonal elastase-treated guinea pig plasma. Also, the increase of kallikrein activity, by injection of pseudomonal elastase, was inhibited by the pretreatment of the DI-mixture in a guinea pig. Since the DI-mixture had no inhibitory effect on kallikrein activity when Z-Phe-Arg-MCA was used as a substrate in vitro, its inhibitory activity in the pseudomonal elastase-induced septic hypotension model might not be due to a direct inhibition of plasma kallikrein in the activation cascade of the Hageman factor and prekallikrein system. These results suggest that the Dolichos DI-mixture might be used as an inhibitor in pathogenic bacterial protease-induced septic shock.

• International Journal of Stem Cells
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• 제15권2호
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• pp.136-143
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• 2022

Background and Objectives: Circulating endothelial progenitor cells (EPCs) participate in vascular repair and predict cardiovascular outcomes. The aim of this study was to investigate the correlation between EPCs and abdominal aortic aneurysms (AAAs). Methods and Results: Patients (age 67±9.41 years) suffering from AAAs (aortic diameters 58.09±11.24 mm) were prospectively enrolled in this study. All patients received endovascular aneurysm repair (EVAR). Blood samples were taken preoperatively and 14 days after surgery from patients with aortic aneurysms. Samples were also obtained from age-matched control subjects. Circulating EPCs were defined as those cells that were double positive for CD34 and CD309. Rat models of AAA formation were generated by the peri-adventitial elastase application of either saline solution (control; n=10), or porcine pancreatic elastase (PPE; n=14). The aortas were analyzed using an ultrasonic video system and immunohistochemistry. The levels of CD34⁺/CD309⁺ cells in the peripheral blood mononuclear cell populations were measured by flow cytometry. The baseline numbers of circulating EPCs (CD34⁺/CD309⁺) in the peripheral blood were significantly smaller in AAA patients compared with control subjects. The number of EPCs doubled by the 14th day after EVAR. A total of 78.57% of rats in the PPE group (11/14) formed AAAs (dilation ratio >150%). The numbers of EPCs from defined AAA rats were significantly decreased compared with the control group. Conclusions: EPC levels may be useful for monitoring abdominal aorta aneurysms and rise after EVAR in patients with aortic aneurysms, and might contribute to the rapid endothelialization of vessels.

• 대한화장품학회지
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• 제24권3호
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• pp.24-30
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• 1998

Inhibitory effects of the new material obtained from Areca catechu seed (CC-516) according to a special process, and its applicability to the skin as a cosmetic raw material in terms of its efficacy were presented. Areca catechu extract out of 150 medicinal plants, exhibited high inhibitory effect on the porcine pancreatic elastase ($IC_{50}$ : $40.8{\mu}$g/ml). It also had an inhibitory effect on the human leukocyte elastase ($IC_{50}$ : 48.1$\mu$g/ml), hyaluronidase ($IC_{50}$ : $416{\mu}$g/ml), antioxidative activity ($IC_{50}$ : $45.4\mu$g/ml) and free radical scavenging activity ($SC_{50}$ : $10.2{\mu}$g/ml). The cream contained 3％ of CC-516 improved skin hydration above 16.5％. Especially, the skin elasticity increases more than 35% and skin wrinkles decreased more than 23％. The CC-516 was designed to be utilized in cosmetology. The cream containing 3％ of this product has not only protecting effect on the skin mechanical properties provided by the collagen and the elastin in the derm but also restructuring effect of scarring or aging tissue.

• BMB Reports
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• 제56권8호
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• pp.439-444
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• 2023

Emphysema is a chronic obstructive lung disease characterized by inflammation and enlargement of the air spaces. Regorafenib, a potential senomorphic drug, exhibited a therapeutic effect in porcine pancreatic elastase (PPE)-induced emphysema in mice. In the current study we examined the preventive role of regorafenib in development of emphysema. Lung function tests and morphometry showed that oral administration of regorafenib (5 mg/kg/day) for seven days after instillation of PPE resulted in attenuation of emphysema. Mechanistically, regorafenib reduced the recruitment of inflammatory cells, particularly macrophages and neutrophils, in bronchoalveolar lavage fluid. In agreement with these findings, measurements using a cytokine array and ELISA showed that expression of inflammatory mediators including interleukin (IL)-1β, IL-6, and CXCL1/KC, and tissue inhibitor of matrix metalloprotease-1 (TIMP-1), was downregulated. The results of immunohistochemical analysis confirmed that expression of IL-6, CXCL1/KC, and TIMP-1 was reduced in the lung parenchyma. Collectively, the results support the preventive role of regorafenib in development of emphysema in mice and provide mechanistic insights into prevention strategies.