지치(Lithospermum erythrorhizon) 추출물의 멜라닌 생합성 억제효과

이황희(Hwanghee Blaise Lee),배석(Suk Bai),진종언(Jong-Eon Chin) 한국식품영양과학회 2005 한국식품영양과학회지 Vol.34 No.9

유전자 발현 조절 수준에서 멜라닌 색소 생합성 억제물질을 탐색하고자 지치 뿌리로부터 유용성 물질을 추출ㆍ분획하여 tyrosinase 프로모터를 지닌 B16 mouse melanoma cell에 처리한 결과 그 메탄올 추출물은 10 ㎍/mL의 농도에서 약 33% 이상의 tyrosinase 프로모터 활성 억제효과를 나타내었으며, 세포 활성능이 100 ㎍/mL의 농도에서 약 108%로 매우 안전하였다. 그리고 methylene chloride, ethyl acetate, butyl alcohol, 물 등의 용매 분획물들도 농도에 따라 차이는 있었지만 100 ㎍/mL 또는 500 ㎍/mL의 고농도에서 tyrosinase 프로모터의 활성을 억제하였다. 또한, 지치 메탄올 추출물의 농도를 달리하여 3일 동안 처리하였을 때 멜라닌 색소의 생성능은 10 ㎍/mL와 100 ㎍/mL의 농도에서 대조군 세포에 비해 각각 약 11%와 24%로 크게 감소하는 결과를 보여주었다. To estimate the inhibitory effect of Lithospermum erythrorhizon root extract on melanin biosynthesis, we tested its inhibitory effects on tyrosinase promoter in B16 mouse melanoma cells. Lithospermum erythrorhizon root extract had inhibitory effect above 33% on tyrosinase promoter at 10 ㎍/mL and exhibited no cytotoxicity under 100 ㎍/mL. Also, melanin biosynthesis decreased approximately 11% and 24% at 10 ㎍/mL and 100 ㎍/ mL, respectively. Therefore, Lithospermum erythrorhizon root extract would be considered very effective regulator of tyrosinase promoter and melanin biosynthesis.

산업용 Saccharomyces cerevisiae에서 Aspergillus awamori Glucoamylase 유전자의 발현

강동명,이수아,전영현,진종언,이황희,배석,Ghang Dong-Myeong,Lee Su-A,Chun Young-Hyun,Chin Jong-Eon,Lee Hwanghee Blaise,Bai Suk 한국미생물학회 2005 미생물학회지 Vol.41 No.2

전분 이용이 가능한 산업용 Saccharomyces cerevisiae균주를 개발하기 위해 alcohol dehydrogenase 유전자 프로모터(ADClp)의 조절하에 발현되는 Aspergillus awamori glucoamylase cDNA 유전자(GA1)를 산업용 S. cerevisiae의 염색체에 도입하였다. 산업적 이용에 적합한 효모균주를 얻기 위해 세균 ampicillin 저항성 유전자가 제거되고 GA1 유전자와 선별 표지유전자로 S. cerevisiae aureobasidin A 저항성 유전자(AUR1-C)와 재조합 부위로 Tyretrotransposon $\delta$-서열이 포함된 integrative cassette를제조하였다. 이 $\delta-integrative$ cassette로 형질전환된 산업용 S. cerevisiae는 배지상에 glucoamylase를 생산 분비하였고 전환을 유일한 탄소원으로 하여 생장하였다. 형질전환체를 비선택배지에서 배양했을 매 삽입된 GA1유전자가 100세대까지 안정되게 유지되었다. To construct an amylolytic industrial strain of Saccharomyces cerevisiae, the glucoamylase cDNA gene (GAl) from Aspergillus awamori was expressed under the control of the alcohol dehydrogenase gene promoter (ADC1p) and integrated into the chromosomes of industrial S. cerevisiae. An integrative cassette lacking bacterial ampicillin resistance gene but containing the GA1 gene, $\delta$ sequences of Ty1 retrotransposon as target sites for homologous recombination and S. cerevisiae aureobasidin A resistance gene (AUR1-C) as the selection marker was constructed to obtain a strain eligible for commercial use. Industrial S. cerevisiae transformed with this 15-integrative cassette efficiently secreted glucoamylase into the medium and grew on starch as the sole carbon source. The transformants were mitotically stable for 100 generations in nonselective medium.

뇌하수체 세포인 GH3세포에서 non-genomic action을 통한 Nonylphenol의 nitric oxide 증진효과

이경진(Kyung-Jin Lee),최철웅(Chul-Yung Choi),손현정(Hyun-Jung Sohn),정백진(Back-Jin Jeong),문소희(So-Hee Moon),이황희(Hwanghee Blaise Lee),이종빈(Jong-Bin Lee) 환경독성보건학회 2003 환경독성보건학회지 Vol.18 No.4

본 연구는 환경호르몬(endocrine disruptors)으로 분류되었으며,에스트로젠 화합물의 특성을 지닌 4-Nonylphenol(NP)이 설치류 pituitary 세포 중 성장호르몬을 분비하는 GH3 세포의 Nitric oxide (NO)을 증가시키는 작용기전을 규명코자 수행되었다. 먼저 GH3세포에 NP처리 농도에 따른 NO의 생성을 측정한결과 NP처리농도 의존적으로 증가시켰다. 이러한 NO의 증가가 genomic action인지를 확인하기 위해GH3 세포의 NO를 증가시키는 효소인 neuronal oxide synthase의 단백질량을 측정한 결과 CH3 세포에서NP에 의한 nNOS의 단백질의 변화는 없었다. 에스트로젠 화합물인 NP가 에스트로젠 리셉터 (ER)와의관계를 조사하기 위해 ER억제제(ICI 168,780)를 처리한 경우 NP에 의해 증가한 NO가 감소하였다. 또한유전자 전사억제제인 actlnomycin D 및 단백질 발현 억제제인 cycloheximide을 처리한 경우는 NP에 의한NO 증가억제효과가 없었다. 이러한 결과를 종합해 볼 때 GH3 세포에서 NP는 ER을 매개한 non-genomicaction에 의해 NO를 증가키는 것으로 사료된다.

피부암 및 피부암 전구증에서의 Acid Stable Trypsin Inhibitor ( ASTI ) 의 발현양상에 관한 연구

이종육 ( Jong Yuk Yi ),이동원 ( Dong Won Lee ),윤두희 ( Dou Hee Yoon ),조백기 ( Baik Kee Cho ),강창석 ( Chang Suk Kang ),이황희 ( Hwanghee Blaise Lee ),장원희 ( Won Hee Jang ),유욱준 ( Ook Joon Yoo ) 대한피부과학회 1996 대한피부과학회지 Vol.34 No.2

Background: Recently, much attention has been focused on the role of protease inhibitors such as acid stable trypsin inhibitor (ASTI) in the invasive growth of malignant tumors. However, there are no report on expression of ASTI from premalignant and/or malignant skin tumors. Objectives : In the present study the expression of ASTI was investigated in the different type of premalignant and/or, malignant skin tumors in attempt to clarify the relation between the expression of the ASTI and malignancy. Methods : For the detection of ASTI in the tumor tissue, the immunoperoxidase techniques that used mouse antibody raised against highly purified ASTI. The degree of ASTI immunoreactivity was semiquantitatively assessed for staining intensity as the percentage of ASTI-positive cells. Results : ASTI immunoreactivity was detected in most of the premalignant and malignant skin tissues. Especially, ASTI expression was present widespread in squamous cell carcinoma(SCC) with strong cytoplasmic membrane, where as in normal epidermis they were primarily present in the horny layer. The strong staining was the SCC, keratoacanthoma, Bowen's disease, actinic keratosis, basal cell epithelioma, Paget's disease in decreasing order. The significant difference in the staining intensity was observed between SCC and other groups. Conclusion : Results of immunohistochemical studies suggest that the tumor cells themselves could produce ASTI. Considering the suggestion that ASTI is a self protector, inhibitor to proteolytic protease as well as growth-stimulating factor. The present findings may indicate that ASTI expressed in malignant cells may play a role possibly closely associated with tumor development. (Kor J Dermatol 1996;34(2): 179-184)

Apicidin, Histone Deacetylase Inhibitor에 의한 Human Promyelocytic U937 세포고사

정은현(Eun-Hynu Jung),박찬희(Channy Park),임창인(Chang-In Lim),이황희(Hwanghee Blaise Lee),송훈섭(Hoon-Seob Song),염성섭(Seong-Seob Yeom),정은배(Eun-Bae Jung),이병곤(Byeong-Gon Lee),김영훈(Young-Hoon Kim),박래길(Raekil Park) 한국독성학회 2003 Toxicological Research Vol.19 No.3

Apicidin, a histone-deacetylase inhibitor, has been successfully used to inhibit the growth of cancer cells. In this study, the apoptotic potential and mechanistic insights of apicidin were<br/> investigated in human myeloid leukemia U937 cells. Treatment of U937 cells with apicidin resulted in a decrease of cell viability with apoptotic characteristics, including chromatin condensation and ladderpattern fragmentation of genomic DNA. Apicidin converted the procaspase-3 protease to catalytically active effector protease, resulting in subsequent cleavage of poly (ADP-ribose) polymerase (PARP) and inhibitor of caspase-activated deoxyribonuclease (ICAD). In addition, apicidin induced the activation of caspase-9 protease and the cytosolic release of mitochondrial cytochrome c with mitochondrial membrane potential transition. Moreover, apicidin transiently increased the expression of Fas and Fas ligand proteins. Taken together, the results suggest that apicidin induces apoptosis of U937 cells through activation of intrinsic caspase cascades and Fas/FasL system with mitochondrial dysfunction.

자외선조사에 의한 TNF-α/IFN-γ 생산능과 Listeria 감염에 대한 면역성 변화

강인철(In-Chol Kang),김천상(Chun-Sang Kim),노경아(Keong-A Rho),임선영(Suhn-Young Im),이황희(Hwanghee Blaise Lee),전순배(Soon-Bai Chun),이현철(Hyun-Chul Lee) 大韓微生物學會 1997 大韓微生物學會誌 Vol.32 No.5

Allomyrina Dichotoma Larva Extracts Protect Streptozotocin-induced Oxidative Cytotoxicity

Deok-Song Kim(김덕송),Jin Huh(허진),Guen-Chang You(유근창),Soo-Chul Chae(채수철),Oh-Sun Lee(이오선),Hwanghee Blaise Lee(이황희),Jong-Bin Lee(이종빈),Jong Sun Kim(김종선) 환경독성보건학회 2007 환경독성보건학회지 Vol.22 No.4

장수풍뎅이 유충(Allomyrina dichotoma larva, ADL)은 중국의 전통 약재로서, 특히 항산화 효과가 우수하여 항당료 제재로 사용되고 있다. 본 연구에서는 이러한 ADL의 추출물을 이용하여 헴스터 췌장의 β­세포(HIT-T15)에서 Streptozotocin에 의해 유발된 산화적 손상에 대한 보호효과 및 그 작용기전을 조사하였다. ADL추출물은 처리농도 의존적으로 Streptozotocin에 의해 유발된 지질과산화 및 세포 내 자유산소종의 양을 억제함으로서 β-세포의 산화적 스트레스에 의한 손상을 보호하였다. 또한 DNA laddering 방법을 사용하여 Streptozotocin에 의해 유발된 DNA손상을 조사한 결과, ADL추출물 처리농도에 비례하여 Streptozotocin에 의해 유발된 DNA 손상이 감소하였다. 이러한 산화적 손상의 억제능 관련 작용 기전을 조사하기 위해 DPPH free radical 소거능을 실시하였다. 그 결과 ADL추출물 자체가 DPPH 자유 레디컬 소거능이 있음을 확인하였으며, 또한 플라스미드를 이용한 Single-strand break 방법을 통한 DNA 손상 보호능을 측정한 결과도 Fe³? 및 H₂O₂에 의해 유발된 DNA 손상이 ADL추출물 처리농도에 비례하여 감소하였다. 이러한 결과들을 종합하여 볼 때, 장수풍뎅이 유충의 추출물들이 자체의 레디컬 소거능 및 산화적 손상에 의한 DNA 손상을 억제함으로써, Streptozotocin에 의해 유발된 산화적 손상을 억제할 수 있을 것이라 사료된다.

합성펩타이드를 이용한 영양배엽세포-특이 가토 다클론 항혈청의 제작

이희섭,오재민,김정중,문형배,김원신,이황희 원광대학교 생명공학연구소 1995 생명공학연구소보 Vol.3 No.1

Within the last few years, a different approach to generating protein-reactive antibodies has been developed that has several advantages over conventional immunization. This involves synthesizing short peptide sequences, coupling them to immunogenic carrier molecules, and immunizing animals with the conjugates. 3βHSD(3β-hydroxy-5-ene steroid dehydrogenase; EC 1.1.1.145) is the enzyme of the plasma membrane of human trophoblast and it's cDNA sequence was identified by Nickon et al(Molecular cloning and expression of human trophoblast antigen FDO161G and its identification as 3β-hydroxy-5-ene steroid dehydrogenase. J Reprod Fert 1991;149;156). For the production of trophoblast-specific antibody, we synthesized three oligopeptides that are epitope sites chosen from cDNA sequence of 3βHSD. Oligopetides were coupled with KLH(keyhole limpet hemocyanin) under 25% glutaraldehyde. The trophoblast-specific rabbit polyclonal antisera was produced by conventional methods. This antisera reacts with a 43kDa protein in human placental lysate by Western blotting analysis and The syncytiotrophoblasts and cytotrophoblasts from villi are stained positively with this antisera by immunohistochemistry. Villous trophoblasts were cultured in methionine-free media for 1 hour and [^(35)S]-Methionine for 24 hours. Media and cell lysate were immunoprecipitated with this antisera and 12% SDS-PAGE was performed. In fluorography, bend was not noted in media and 43kDa band was noted in medis and 43kDa band was noted in lysate. It was concluded that anti-3βHSD antibody produced by synthetic peptide was specific to trophoblasts and 3βHSD was membrane-bound protein of trophoblasts.