경옥고가미방 추출물이 생쥐의 양모 및 발모 관련 단백 발현에 미치는 영향

도은주 ( Eun Ju Do ),황미열 ( Mee Yul Hwang ),김승연 ( Seung Yeon Kim ),이진상 ( Jin Sang Lee ),양대석 ( Dae Seok Yang ),양재하 ( Chae Ha Yang ),김미려 ( Mi Ryeo Kim ) 대한본초학회 2011 大韓本草學會誌 Vol.26 No.4

Objective: Present study was carried out to investigate the effect of Gyungohkgo-gamibang extract on hair growth and protein expression in an alopecia model of C57BL/6 mice. Methods: Mice were divided into 3 experimental groups including normal (vehicle), Gyungohkgo-gamibang extract (YNS-10) and 5% minoxidil-treated group. The test materials were daily applied with 0.1 ml per mouse on shaved dorsal skin for 3 weeks. The hair growth was monitored by photograph at 5, 10, 15, 21 days after topical application. Then the changes of hair density and hair thickness in the hair-removed area were evaluated by phototrichogram using folliscope. Also the expression level of growth factors related to hair growth was measured by western blotting. Results: Application of minoxidil or YNS-10 stimulated the hair growth compared to vehicle treatment. Therefore hair density of minoxidil or YNS-10 application was increased about 200% and 210% more than in vehicle application on 14 day, respectively. And hair thickness of both minoxidil group and YNS-10 group was increased about 220% and 210 % more than in vehicle spreading on 14 day, respectively. Futhermore the protein expression of IGF-1 and VEGF were significantly up-regulated on 7 day in YNS-10 and minoxidil-spreaded group compared to vehicle-applied group. Conclusion: These data suggest that YNS-10 has potent stimulating activity on hair growth in C57BL/6 mice and potential usefulness as ingredients of hair tonic and hairrestore.

한약재 추출물 함유 앰플의 주름개선 효능에 관한 기초 임상연구

도은주,이진상,박현진,하일도,김영철,김미려,Do, Eun-Ju,Lee, Jin-Sang,Park, Hyun-Jin,Ha, Il-Do,Kim, Young-Chul,Kim, Mi-Ryeo 대한한의학방제학회 2009 大韓韓醫學方劑學會誌 Vol.17 No.2

Objectives : This study was performed to evaluate the efficacy of ampule containing herbal extract(Ulmus davidiana, Ginkgo biloba, Perilla ocymoides, Morus alba, Glycyrrhiza uralensis (licorice), and Angelica gigas) on skin wrinkles in adult women with facial wrinkles. Methods : A total of 12 women, 30 to 46 years of age, with wrinkles in the corner of eyes, applied ampule containing herbal extract twice daily for up to 12 weeks. Silicon replicas of right crow's feet area and antecubital fossa were taken before use of test product and at 4, 8 and 12 weeks. The replicas were analyzed by optical profilometry with Skin Visiometer SV600. The wrinkle and roughness parameters, R1, R2, R3, R4 and R5 were calculated and statistically analyzed. In addition, a subjective evaluation of product efficacy was conducted by patient's assessment. Results : The mean values of all the skin roughness parameters were decreased at 4-week, and decreased significantly at 8 and 12-week except R4 at 12-week after test product use. The subjective evaluation of wrinkle by patient's assessment was also improved. However, one patient noted stiffness of face after use of test product. Conclusions : These study suggested that the application of ampule containing herbal extract twice daily for 12 weeks may effectively improve the facial wrinkles without severe side effect.

TAR cloning 법에 의한 인간 및 마우스의 상동성 HPRT 유전자의 분리

도은주,김재우,정정남,박인호,임선희,Do, Eun-Ju,Kim, Jae-Woo,Chung, Chung-Nam,Park, In-Ho,Leem, Sun-Hee 한국생명과학회 2006 생명과학회지 Vol.16 No.6

TAR (Transformation-Associated Recombination) cloning법은 복잡한 고등생물의 게놈으로부터 유전자나 특정 염색체 부위를 선별적 분리를 가능하게 한다. 이 방법은 목적으로 하는 염색체 부위의 주변에 존재하는 비교적 짧은 게놈 염기서열에 대한 정보를 필요로 한다. 이 기술은 출아효모의 spheroplasts 형질전환 동안 목적 유전자를 포함한 게놈 DNA와 그 유전자의 5' 또는 3' 말단 서열 (hook)을 포함하고 있는 TAR vector 사이에 일어나는 상동성 재조합에 의해 이루어진다. 본 연구에서는 TAR cloning 법을 상동성 유전자의 분리에 사용할 수 있는가를 조사하기 위해, 연간과 마우스 게놈의 HPRT 유전자를 선택하였다. 그 결과, 인간과 마우스의 게놈으로부터의 HPRT 유전자의 분리 빈도는 TAR vector로서 hHPRT hook 혹은 mHPRT hook을 사용한 경우에 거의 동일하게 나타났다. 또한 mHPRT 유전자의 gap 부분의 염기서열을 결정하여, 이 부분에 염기서열의 불안정의 요인이 되는 비정상적 특성을 발견하였다. 결론적으로 TAR cloning법을 이용하여 다른 이종 간의 게놈으로부터 상동성 유전자 즉 orthologue의 분리가 가능하였다. 더욱이 TAR cloning 시스템을 이용하여 고등동물 게놈 상에 남아있는 gap 부분을 메움으로서 고등동물의 모든 유전자들의 확인이 가속화될 수 있을 것으로 사료된다. The transformation-associated recombination (TAR) cloning technique allows selective isolation of chromosome regions or genes from complex genome. The procedure requires knowledge of relatively small genomic sequences that reside adjacent to the chromosome region of interest. This method involves homologous recombination during spheroplast transformation between genomic DNA and a TAR vector that has 5' and 3' gene targeting sequences (hooks). To examine whether TAR cloning can be applied to the isolation of gene homologues, we chose the HPRT genes from human and mouse genome. As results, the yield of positive clones for HPRT gene from human and mouse genome when using a TAR vector containing mHPRT hook or hHPRT hook was almost same level. Analysis of the gap regions in mHPRT revealed that they contain abnormalities that could result in instability of the sequences. In conclusion, we were able to use the TAR cloning technology to isolate gene homologue (orthologue) from nonidentical genome. Moreover, the use of the TAR cloning system may accelerate work on closing the remaining gaps in mammalian genome to achieve the goal of annotation of all mammalian genes.

블랙초크베리 분획물로부터의 주름억제 효과에 대한 작용기전

최은영(Eun-Young Choi),김은희(Eun-Hee Kim),이재봉(Jae-Bong Lee),도은주(Eun-Ju Do),김상진(Sang-Jin Kim),김세현(Se-Hyeon Kim),박정열(Jeong-Yeol Park),이진태(Jin-Tae Lee) 한국생명과학회 2016 생명과학회지 Vol.26 No.1

학명이 Aronia melanocarpa인 블랙초크베리는 항산화, 항염증, 항암 효능이 뛰어난 것으로 보고되고 있다. 본 연구에서는 블랙초크베리에 대한 collagenase inhibition effects와 산화적 스트레스에 유도된 matrix metalloproteinase(MMP), MAPkinase 그리고 AP-1의 발현 그리고/또는 인산화와 같은 분자생물학적 메카니즘을 조사하였다. Collagenase inhibition 효과는 블랙초크베리 에틸아세테이트 분획물(AE)이 500 μg/ml의 농도에서 77.2% 이상의 저해효능을 나타내었고 이는 대조군인 Epigallocatechin gallate의 결과(500 μg/ml에서 83.9%)와 비교해서 유의할 만한 결과였다. Reactive oxygen species (ROS) assay는 AE에서 가장 농도의존적으로 ROS 생성이 감소되었고, 75 μg/ml의 농도에서 약 70%로 가장 낮은 활성산소가 생성되었다. MTT assay 결과, H2O2에 유도된 CCRF 세포에 AE를 처치하였을 때 농도의존적으로 세포 생존율이 증가하였다. 그리고 특히, AE는 H2O2에 유도된 CCRF 세포에서의 MMPs (MMP-1, -3 그리고 -9), MAPK (ERK, JNK 그리고 p38) 그리고 AP-1 (c-Fos와 c-Jun)의 발현과 인산화를 억제하였고, pro-collagen type I의 발현은 증가시켰다. 따라서 블랙초크베리 에틸아세테이트 분획물은 주름억제 및 콜라겐 생성의 효능이 있으며 기능성 식품 및 화장품 소재 개발 산업에서의 응용이 가능할 것으로 기대된다. Black chokeberries (scientific name Aronia melanocarpa) have been reported to have major effects due to anti-oxidant, anti-inflammatory, and anti-cancer capabilities. In this study, we investigated the anti-wrinkle effects of A. melanocarpa, including collagenase inhibition effects and their molecular biological mechanisms, such as oxidative stress-induced matrix metalloproteinase (MMP), mitogen-activated protein (MAP) kinase, and activator protein (AP)-1 expression and/or phosphorylation. In collagenase inhibition activity, the ethyl acetate fraction of black chokeberry (AE) was 77.2% at a concentration of 500 μg/ml, which was a significant result compared to that of Epigallocatechin gallate (positive control, 83.9% in 500 μg/ml). In the reactive oxygen species (ROS) assay, the AE produced 78% of ROS in 10 μg/ml and 70% of ROS in 75 μg/ml, which was a much lower percentage than the ROS production of H2O2-induced CCRF S-180II cells. In the MTT assay, cell viability was increased dose-dependently with AE in H₂O₂-induced cells. In protein expression by western blot assay, the AE suppressed the expression and phosphorylation of MMPs (MMP-1, -3, -9), MAPK (ERK, JNK, and p38), and AP-1 (c-Fos and c-Jun), and expressed the pro-collagen type I in H₂O₂-induced cells. These results suggest that black chokeberries have anti-wrinkle and collagen-production effects, and they may be used in applications for material development in the functional food and cosmetic industries.

Target DNA 염기서열 내에 존재하는 비상동성 간격이 상동성재조합을 이용한 클로닝 빈도에 미치는 영향

김재우,도은주,윤세련,정윤희,윤영호,임선희,선우양일,박인호,Kim Jae-Woo,Do Eun-Ju,Yoon Se-Lyun,Jeong Yun-Hee,Yoon Young-Ho,Leem Sun-Hee,Sunwoo Yangil,Park In-Ho 한국미생물학회 2005 미생물학회지 Vol.41 No.4

Transformation-associated recombination (TAR) 클로닝 법은 복잡한 게놈으로부터 염색체 내의 특정부위나 유전자를 선택적으로 분리할 수 있다. 이 방법은 목적 유전자에 근접한 작은 게놈DNA 염기서열 정보를 필요로 한다. 이 기술은 효모의 spheroplast transformation을 시키는 동안 목적으로 하는 유전자의 5' 또는 3' 서열을 포함하고 있는 TAR vector와 게놈DNA사이에서 일어나는 상동성재조합에 의해 이루어진다. 본 연구에서는 plasmid 모델시스템을 이용하여 target hooks 내에 존재하는 비상동성 염기서 열이 상동성재조합에 미치는 영향을 조사하였다. plasmid에 존재하는HIS3유전자와 변형시킨 his3-TRP1-his3 단편 사이의 상동성재조합의 효율은 $Ura^+$ 형질전환체의 형질분석에 의해 이루어졌다. $Ura^+$ 형질전환체의 수는 7종류의 서로 달리 변형된 his3-TRP1-his3 단편들을 사용하였을 매 거의 동일하게 나타났다. 그러나 $Trp^+His^+$ positive recombinants의 빈도는 변형된 his3-TRP1-his3 단편 내에 비상동성 영역에 부정확한 간격을 지닐 때 현저한 감소를 나타내었다. 이러한 결과로서, 부정확한 간격이 target hook과 substrate DNA 사이에 일어나는 상동성재조합을 방해하는 것으로 사료된다. 그러므로 이종간의 상동유전자를 클로닝 할 때에는 target hook내의 비상동성 염기서열이 존재한다면 이것이 정확한 간격을 지니는지 여부를 중요란 요인으로 고려해야 한다. Transformation-Associated Recombination (TAR) cloning technique allows selective isolation of chromosomal regions and genes from complex genomes. The procedure requires knowledge of relatively small genomic sequences that reside adjacent to the chromosomal region of interest. This technique involves homologous recombination during yeast spheroplast transformation between genomic DNA and a TAR vector that has 5' and 3' gene targeting sequences. In this study, we examined the effect of non-homologous spacing sequence in target hooks on homologous recombination using a plasmid model system. The efficiency of homologous recombination between the modified his3-TRP1-his3 fragments and HlS3 gene on plasmid were analyzed by the characterization of $Ura^+$ transformants. The numbers of $Ura^+$ transformant showed same level when seven different modified his3-TRP1-his3 fragments were used. But the percentage of positive recombinants. $Trp^+His^-$, dramatically decreased when used the modified his3-TRP1-his3 fragments contained incorrect spacing of nonhomologous region. As a result, we suggest that incorrect spacing inhibits the homologous recombination between target hook and substrate DNA. Therefore, we should consider the correct spacing in target hook when the target hook are used for cloning of orthologue gene.

여주 추출물을 함유한 한방화장품의 주름 및 미백 개선에 관한 임상적 연구

정호준,도은주,이진상,박현진,오규석,김미려,하일도,정용하,지선영,Jung, Ho-Jun,Do, Eun-Ju,Lee, Jin-Sang,Park, Hyun-Jin,Oh, Gyue-Seok,Kim, Mi-Ryeo,Ha, Il-Do,Jeong, Yong-Ha,Jee, Seon-Young 대한한방안이비인후피부과학회 2011 한방안이비인후피부과학회지 Vol.24 No.2

Objective : The purpose of this clinical research was to investigate the effects of herbal cosmetics containing Momordica charntia L. extracts on the anti-wrinkle and whitening effects. Methods : A total of 25 subjects who visited Daegu Hanny Oriental Medical Center from November 1st, 2010 to December 31th, 2010 were included in this study. In this study, we observed change of R1, R2, R3, R4, R5 for using products, change of average melanin content and pigmental area, analysis results of subjective wrinkle improvement, Analysis results of subjective whitening improvement. Statistical analysis was performed by using paired t-test and wilcoxon signed ranks test. Statistical significance was achieved if the probability was less than 5%(p<0.05) Results : Statistically, herbal cosmetics containing Momordica charntia L. extracts showed siginificant effect on change of R1, R2, R3, R4, R5 for using products, change of average melanin content and pigmental area, analysis results of subjective wrinkle improvement, Analysis results of subjective whitening improvement(p<0.05). And satisfaction after using herbal cosmetics was near good. Conclusions : Considering the above results, we have concluded that herbal cosmetics containing Momordica charntia L. extracts have the remarkable effects on the anti-wrinkle and whitening effects.

아토피 피부염 환자에서 상황버섯 추출물을 함유한 한방화장품의 임상적 연구

정호준,황보민,도은주,석장미,김미려,김영훈,도국배,이창언,지선영,Jung, Ho-Jun,HwangBo, Min,Do, Eun-Ju,Suk, Jang-Mi,Kim, Mi-Ryeo,Kim, Young-Hun,Do, Kook-Bae,Lee, Chang-Eon,Jee, Seon-Young 대한한방안이비인후피부과학회 2010 한방안이비인후피부과학회지 Vol.23 No.3

Objective : The purpose of this clinical research was to investigate the effects of herbal cosmetics containing Phellinus linteus extracts on patients with atopic dermatitis. Methods : A total of 35 patients who visited Daegu Hanny Oriental Medical Center from November 4th, 2008 to December 28th, 2008 were included in this study. In this study, we observed transepidermal water loss, skin moisture content, modified scorad index of atopic dermatitis and pruritic degree. Statistical analysis was performed by using paired t-test and wilcoxon signed ranks test. Statistical significance was achieved if the probability was less than 5%(p<0.05). Results : Statistically, herbal cosmetics containing Phellinus linteus extracts showed siginificant effect on transepidermal water loss, skin moisture content, modified scorad index of atopic dermatitis and pruritic degree(p<0.05). And satisfaction after using herbal cosmetics was near good. Conclusions : Considering the above results, we have concluded that herbal cosmetics containing Phellinus linteus extracts have the remarkable effects on atopic dermatitis.

한약재 혼합추출물이 생쥐 탈모 모델의 모발성장 촉진에 미치는 영향

황미열 ( Mee Yul Hwang ),도은주 ( Eun Ju Do ),정금희 ( Geum Hee Jung ),김미려 ( Mi Ryeo Kim ) 한약응용학회 2014 한약응용학회지 Vol.14 No.2

Objective:This study was performed to examine the effect of mixed medicinal herbal extract including Lespedeza bicolor Turcz. and 18 others (CS-10-H) on hair growth in an alopecia model of C57BL/6 mice. Methods:Male mice were divided into 3 experimental groups for treatment of CS-10-H or 5% minoxidil or vehicle solution, respectively. Dorsal skin of animals were shaved and topically applied with an amount of 200 μL of test materials once a day for 3 weeks. The apparent hair growth was monitored at 5, 10, 15 and 21 days after topical application. The changes of hair density and hair thickness were evaluated by phototrichogram using folliscope. Protein expression levels of hair growth-related factors were measured by western blotting. Results:Application of minoxidil or CS-10-H stimulated the hair growth compared to vehicle treatment. Hair density of minoxidil and CS-10-H group was increased about 206% and 208% on 14 day compared to that of vehicle group, respectively. Hair thickness of minoxidil and CS-10-H group was also increased about 220 % and 200% on 14 day compared to that of vehicle group. The protein expression of hair growth factors, IGF-1 and VEGF was significantly up-regulated in CS-10-H and minoxidil group than in vehicle treated group. Conclusion:These data suggest that CS-10-H shows potential usefulness as ingredients of hair tonic and hair-restorer for development of cosmetics and quasi-drugs, as result of its potent stimulating activity on hair growth in C57BL/6 alopecia model.

혈중지질에 대한 진피추출물의 유효성 평가를 위한 인체적용시험: 무작위배정, 이중맹검

이진상 ( Jin Sang Lee ),도은주 ( Eun Ju Do ),곽민아 ( Min A Kwak ),박현진 ( Hyun Jin Park ),하일도 ( Il Do Ha ),성기준 ( Ki Jun Sung ),김미려 ( Mi Ryeo Kim ) 대한본초학회 2011 大韓本草學會誌 Vol.26 No.1

Objectives:This study was performed to evaluate the efficacy of extract from Citri Pericarpium (CP) on lipid-lowering effect in semi-healthy subjects with increased serum levels of total cholesterol and triglyceride. Methods: A experimental group (n=45) and placebo group (n=46) were randomly established based on the plasma lipid profiles (triglyceride, total cholesterol etc.) in the semi-healthy subjects, then all subjects randomly took experimental or placebo capsules, which contain 1,200 mg of CP or the same dose of soluble starch per day with regular meals for 8 weeks by double-blind method. Results: One-hundred forty (140) subjects were screened, then 91 subjects enrolled in the study. No serious adverse events were reported for trial period. The CP treatment showed statistically significant decrease in serum levels of triglyceride both at 4-weeks (p<0.01) and at 8-weeks (p=0.0001). Futhermore, there was significant difference in the triglyceride levels between CP-treated and placebo group. The CP-treated but not placebo group, experienced decrease in serum levels of total cholesterol at 8-weeks. Then there was no significant difference in the total cholesterol levels between the two groups. In addition, statistically significant difference of AST, ALT, γ-GT serum levels wasn``t shown before and after trial in two groups. Conclusion: These results demonstrate that CP can significantly suppress the elevated triglyceride but not total cholesterol level. Therefore, CP may play a role in the improvement of blood lipid levels, as an effective functional food.

효모의 재조합 변이주를 이용한 인간 Centromeric Alphoid DNA Repeat의 안정성에 관한 연구

김광섭,신영선,이상엽,안은경,도은주,박인호,임선희,선우양일,Kim, Kwang-Sup,Shin, Young-Sun,Lee, Sang-Yeop,Ahn, Eun-Kyung,Do, Eun-Ju,Park, In-Ho,Leem, Sun-Hee,SunWoo, Yang-Il 한국미생물학회 2007 미생물학회지 Vol.43 No.4

Centromere는 채세포분열과 생식세포분열 등 맡은 주요 기능을 담당하는 고도로 분화된 구조이다. Alphoid DNA (${\alpha}$-satellite)는 인간뿐 아니라 모든 영장류의 염색체 내 centromere에서 발견되는 반복서열의 대부분을 차지한다. 인간 인공염색체(Human Artificial Chromosome, HAC)의 개발에서 가장 핵심적인 부분은 centromere의 분리 및 안정적인 유지에 있다. 이 영역은 출아효모에서 alphoid DNA 반복서열을 hook으로 이용하여 Transformation-associated recombination (TAR) cloning법을 사용하여 선택적으로 분리할 수 있다. 이러한 실험방법으로 먼저 repeat array를 rolling-circle amplication (RCA)를 통하여 약 5 kb까지 길이를 연장시킨 후, 효모내에서 상동성재 조합을 이용한 TAR cloning법을 사용하여 분리할 수 있다. 이렇게 분리된 35 kb-50 kb 길이의 4종류의 centromeric DNA repeat arrays (2,4,5,6 mer)를 사용하여, 반복서열의 안정성 유지를 조사하기 위해 상동성재조 합 변이주인 rad51, rad52, rad54를 사용하여 비교 분석하였다. 야생주, rad51과 rad54 변이주를 이용하여 형질전환을 수행한 결과, 반복서열의 크기에 있어서 많은 변화를 나타내었다. 반면, rad52 변이주는 야생주와 다르게 형질전환빈도가 매우 낮은 비율로 나타났으나, centromeric DNA repeat array의 안정성은 3배 이상으로 높게 나타냈다. 이러한 결과들을 미루어, rad52 변이주를 사용하여 centromeric DNA repeat arrays의 형질전환실험에서 발생하는 맡은 변이를 줄일 수 있을 것으로 보인다. 이러한 유전적 방법은 HAC 제작에서 반복서열의 유지에 훨씬 효율적으로 사용할 수 있을 것으로 사료된다. The centromere is a highly differentiated structure of the chromosome that fulfills a multitude of essential mitotic and meiotic functions. Alphoid DNA (${\alpha}$-satellite) is the most abundant family of repeated DNA found at the centromere of all human chromosomes, and chromosomes of primates in general. The most important parts in the development of Human Artificial Chromosomes (HACs), are the isolation and maintenance of stability of centromeric region. For isolation of this region, we could use the targeting hook with alphoid DNA repeat and cloned by Transformation-Associated Recombination (TAR) cloning technique in yeast Saccharomyces cerevisiae. The method includes rolling-circle amplification (RCA) of repeats in vitro to 5 kb-length and elongation of the RCA products by homologous recombination in yeast. Four types of $35\;kb{\sim}50\;kb$ of centromeric DNA repeat arrays (2, 4, 5, 6 mer) are used to examine the stability of repeats in homologous recombination mutant strains (rad51, rad52, and rad54). Following the transformation into wild type, rad51 and rad54 mutant strains, there were frequent changes in inserted size. A rad52 mutant strain showed extremely low transformation frequency, but increased stability of centromeric DNA repeat arrays at least 3 times higher than other strains. Based on these results, the incidence of large mutations could be reduced using a rad52 mutant strain in maintenance of centromeric DNA repeat arrays. This genetic method may use more general application in the maintenance of tandem repeats in construction of HAC.