90~93년도 서울 지역에 출하된 돼지의 Aujeszky`s병 감염 실태 조사

최준식 ( Joon Sig Choi ),육동현 ( Dong Hyeun Yuk ),김성삼 ( Sung Sham Kim ),문현칠 ( Hyeun Chil Mon ) 한국가축위생학회 1994 韓國家畜衛生學會誌 Vol.17 No.1

The porcine Aujeszky`s disease was surveyed by Enzyme Immunodiffusion method and serologic neutralization test to the slaughtered pigs at slaughtehouse only in Seoul from March, 1990 to October, 1993. After detecting the positive by enzyme immunodiffusion method primary, we decided finally the positive by serologic neutralization test secondary. Results obtained through the experiments were summarized as followed; 1. The positive of Aujeszky`s disease in March, 1990 was 2 of 1,000 sera. 2 positive were decided as the consigned pigs in Kalsan, HongSung, Chungnam. 2. The positive of Aujezsky`s disease in May, 1991 was decided 1 serum at Pogok, Yongin, Kyeonggi. 3. All of the positive detected by Enzyme Immunodiffusion Method in 1993 were decided finally in the negative sera. 4. The positive sera detected in 1992 were decided 32 sera at Gyeonggi, 6 at Chungnam, and 1 at Gangwon. Especially, the positive sera percentage detected by Kit Latex Aujeszky Test appeared 78.04% at Gyonggi and by enzyme immunodiffusion Method appeared 11.11% at Chungnam and Gangwon.

염기성 올리고펩티드 유도체를 가진 고분자 리피드의 합성 및 유전자 전달 효과 연구

최준식(Joon Sig Choi),배선주(Seon Joo Bae),최혜(Hye Choi) 한국고분자학회 2013 폴리머 Vol.37 No.1

폴리디아세틸렌(polydiacetylene, PDA)은 자기조립된 디아세틸렌(diacetylene) 단량체의 광중합에 의해 만들어 진다. 디아세틸렌 단량체들이 조직적으로 배열되면 254 nm의 자외선 노광에 의해 1,4-첨가 중합이 일어나 고분자 주사슬에 이중결합과 삼중결합이 교대로 존재하는 폴리디아세틸렌이 만들어진다. 폴리디아세틸렌 수용액은 일반적으로 약 640 nm에서 최대흡수파장을 지니는 청색을 띠게 되며 여기에 온도나 pH의 변화, 다른 물질의 결합 등 외 부 자극에 의해 약 550 nm의 최대 흡수 파장을 띠는 적색으로 색 전이가 일어나게 된다. 본 연구에서, 우리는 고체 상 펩티드 합성을 이용하여 PCDA(10,12-pentacosadyinoic acid) 리포좀의 표면에 양이온성 올리고펩티드를 도입하였다. 또한 다양한 몰 비율로 리포좀 수용액을 제조하여 동물 세포에 트랜스펙션한 결과, 향상된 유전자 전달 효율 과 낮은 독성을 보이는 것을 확인하였고, PCDA의 특성을 이용하여 세포에 처리 후 세포 관련 비표지 형광을 관찰 하였다. Polydiacetylene (PDA) is made by photopolymerization of self-assembled diacetylene monomers. If diacetylene monomers are arranged systematically and close enough with distance of atoms, 1,4-addition polymerization will occur by the irradiation of 254 nm ultraviolet rays and then PDA will have alternated ene-yne polymer chains at the main structure. Aqueous solutions of diffused PDA is tinged with blue which shows λmax 640 nm. Visible color changes from blue to red occurs in response to a variety of environmental perturbations, such as temperature, pH, and ligand-receptor interactions. In this study, we synthesized cationic peptides - PCDA(10,12-pentacosadyinoic acid) liposome using a solid phase peptide synthesis (SPPS) method and prepared liposome solutions at various molar ratios using MPEG-PCDA. When mammalian cells were treated with the liposomes, high transfection efficiency and low toxicity were observed.

pH 기능성 안정화된 플라스미드-리피드 고분자 나노입자의 낮은 pH 처리 효과

최준식 ( Joon Sig Choi ) 충남대학교 기초과학연구원 2023 충남과학연구지 Vol.40 No.1

The pH-sensitive stabilized plasmid-lipid polymeric nanoparticles were preincubated at low pH prior to in vitro transfection. The nanoparticles were characterized by measuring time-dependent difference in mean diameter at pH 8.2 and the size change profile when preincubated at low pH was also measured. The particles were applied to the transfection of CV-1 cells and showed better gene transfection efficiency when it was preincubated at low pH prior to transfection experiments. From the confocal microscopy image, the nanoparticles were localized inside cells after 45 hr of incubation. This study suggests that the pre-treatment of pH-sensitive stabilized plasmid-lipid nanoparticles at low pH provides a more functional synthetic vector potentially useful for nonviral gene therapy.

ADV 국내 분리주를 접종한 실험 동물의 조직병리 소견과 항체가 연구

최준식 ( Joon Sig Choi ),육동현 ( Dong Hyeun Yuk ),김성삼 ( Sung Sham Kim ),박석기 ( Seok Gee Park ) 한국가축위생학회 1991 韓國家畜衛生學會誌 Vol.14 No.2

We surveyed the antibody titers in the slaughtered pigs by enzyme immunodiffusion method for the epidemiological distribution of Aujeszky`s disease in Korea. And also we observed the clinical symptoms, histopathological findings and antibody titers by serologic neutralization test in experimental animals infected with Aujeszky`s disease virus(ADV) isolated from Korea. The results of the experiments were summarized as follows. 1. We detected 2 pigs infected with ADV of 1000 in 1990 and 1 of 600 in 1991 by enzyme immunodiffusion method. 2. In histopathological findings of experimental animals inoculated ADV, the typical characteristics of Aujeszky`s disease were not observed in pig, but edema and degeneration in the epidermis were observed in rats, vasculoendotheriosis, perivascular cuffing and cellular infiltration in the cerebrum were appeared in rabbits and perivascular cuffing and forcal infiltration of glial cells in the cerebrum were appeared in mice. 3. The increasing antibody titers (1:64) in the serological neutralization test were ascertained in 2 carrier pigs inoculated ADV. 4. Rabbits, mice and rats died all about 5 days after inoculation, but pigs and chickens didn`t die. 5. In 1 cat and 2 rabbits inoculated with the Aujeszky`s disease virus(ADV), the typical clinical findings of Aujeszky`s disease were observed in rabbit, but not in cat and so we slaughtered without any signs of Aujeszky`s disease.

다양한 금속 이온을 이용한 카세인 단백질 나노입자 형성 및 약물 전달체 특성 연구

김민주,이슬기,최준식,Minju Kim,Seulgi Lee,Joon Sig Choi 한국공업화학회 2023 공업화학 Vol.34 No.2

Casein is a milk protein and one of the most important nutrients in milk. The composition is over 80% in cow's milk and about 20~45% in human's milk. Casein is highly biocompatible and biodegradable, so it has been studied for various biomedical materials applications as well as drug delivery systems. It is widely known that casein can be prepared as nanoparticles in the presence of the Ca²⁺ metal ion. Because casein is amphiphilic, hydrophobic drugs could be loaded inside to form a protein-based drug delivery system. In this study, we studied the optimum conditions for casein nanoparticle formation using natural metal ions present in the body, such as calcium, magnesium, zinc, and iron. It was confirmed that nanoparticles have a uniform size of around 150 nm and negative zeta potential values. In addition, it was demonstrated that casein nanoparticles have a cell viability of more than 80% and efficient intracellular uptake properties using confocal microscopy. From the results, it was also shown that the casein nanoparticles prepared using various metal ions have the potential to be biocompatible drug delivery carriers.

리포좀 개질화된 로즈마린산의 특성 분석과 항암 · 항산화 효과

이슬기(Seulgi Lee),최준식(Joon Sig Choi),노주예(Ju-Ye Ro) 한국고분자학회 2021 폴리머 Vol.45 No.4

애플 민트(Mentha suaveolens)는 전통적으로 사용된 허브의 한 종류로써 저혈압, 진통, 항염 효과를 갖고 있다. 로즈마린 산(rosmarinic acid, RA)은 그 활성 성분으로 알려져 있다. RA는 항산화, 항세포사멸, 항암, 항염증과 같은 약리학적 효과를 나타내는 것으로 알려져 있다. 본 연구에서는 RA를 폴리에틸렌 글리콜과 리피드를 이용하여 리포좀화시켜 기존의 RA 보다 더 높은 약리학적 효과를 나타내는 약물전달체를 개발하였다. 광적동산란(dynamic light scattering), 제트 전위(zeta potential), 전계 방사형 주사전자현미경(field emission scanning electron microscopy) 등을 이용한 입자의 물리화학적 물성을 분석하고, 마우스 해마 유래 세포주인 HT22에서 항산화 효과 및 사람 유방암 유래 세포주인 MDA-MB-231에서 확인하였다. Apple mint (Mentha suaveolens) has been widely used in traditional herbal medicine since it shows hypotensive, analgesic, and anti-inflammatory effects. The active ingredient of apple mint is rosmarinic acid (RA), an ester of caffeic acid and 3,4-dihydroxyphenyl lactic acid. RA has many pharmacological effects, such as anti-oxidative, anti-apoptotic, anti-tumorigenic, and anti-inflammatory properties. In this study, RA is incorporated in PEGylated liposomes to enhance its pharmacological effects. The physical characteristics of RA liposomes were demonstrated by dynamic light scattering, zeta potential, and field emission scanning electron microscopy. The anti-oxidation and anti-cancer effects of RA liposomes were evaluated in HT22 and MDA-MB-231 cell lines, respectively.

생체 적합성을 지니는 글리콜 키토산을 기반으로 하는 나노입자 제조를 통한 유전자 전달체 연구

이영화(Young Hwa Lee),최준식(Joon Sig Choi) 한국고분자학회 2018 폴리머 Vol.42 No.5

최근 많은 연구를 통해 유전자 전달체로써 생체 적합하며 생분해성이 뛰어난 키토산에 대한 연구가 활발히 진행되고 있다. 본 연구에서는 키토산의 유도체인 글리콜 키토산을 사용하여 메틸 아크릴레이트와 폴리에틸렌이민을 도입하여 유전자 전달체를 개발 및 연구하였다. 최종 합성물인 GMP 1300과 GMP 2000은 핵자기 공명 분광기를 이용해 합성 정도를 평가하였고, 플라스미드 DNA와의 결합력 확인을 위해 전기영동을 수행하였다. 그리고 나노입자 분석기를 이용하여 플라스미드 DNA와 결합시킨 GMP 1300과 GMP 2000의 입자 크기와 표면 전하를 측정하였다. In vitro 실험을 위하여, HeLa와 HEK 293 세포주를 이용하여 세포 독성 평가를 진행하였고, 세포 유입은 공초점 현미경을 통해 확인하였다. 또한 유전자 발현율을 평가하기 위해 녹색 형광 단백질을 발현하여 이미지로 확인하였고, 루시페라아제와 단백질 정량을 통해 유전자 발현 효율을 측정하였다. 그 결과, 본 연구에서 제조된 나노 입자들은 독성이 현저히 낮으며, 유전자와의 결합력이 강하고, 세포 내 유입에 용이하다는 것을 통해 생체적합한 유전자 전달체로써 응용될 수 있을 것으로 기대된다. Since chitosan is biocompatible and biodegradable, it has been studied as a good material as a gene delivery carrier. In this study, we have developed and studied a gene delivery carrier by introducing methyl acrylate and polyethylenimine using glycol chitosan, a derivative of chitosan. GMP1300 and GMP2000 were identified by 1H NMR and electrophoresis was performed to confirm the binding ability with pDNA. The particle size and surface charge of GMP1300 and GMP2000 with pDNA were measured using a zetasizer. Cytotoxicity was assessed using HeLa and HEK293 cell lines, and cellular uptake was confirmed by confocal microscopy. In addition, the gene expression rate in HeLa cell line was confirmed by confocal image through green fluorescent protein (GFP), and transfection efficiency was measured by protein quantification. As a result, the prepared nanoparticles were expected to be used as biocompatible gene delivery carriers because of their low toxicity and strong binding force with genes.

2세대의 PAMAM 덴드리머와 KKRK 펩티드를 접합한 고분자 소재의 합성 및 유전자 전달체 특성 연구

김연진(Younjin Kim),최준식(Joon Sig Choi) 한국고분자학회 2022 폴리머 Vol.46 No.1

본 연구에서, 2세대의 폴리도아민 덴드리머(PAMAM G2)와 nuclear localization sequence(NLS) 서열을 최적화한 KKRK, KKRKH 펩티드를 이용하여 새로운 하이브리드 고분자를 합성하였고 이들의 유전자 전달체로서의 특성을 확인하였다. Hep G2와 N I H3T3 세포주를 이용하여 유전자 형질변환 실험으로, 원래의 PAMAM G2 고분자와 비교하여 각 세포에서 20배, 2700배 이상 증가된 유전자 전달 효율을 확인하였다. 다양한 세포 내 이입 저해제들을 이용한 실험을 통해 본 고분자/플라스미드 DNA 복합체의 주된 엔도사이토시스 경로가 거대음작용 메카니즘임을 확인하였다. 공초점 현미경을 통해 플라스미드 DNA가 세포 핵 주변에 분포함을 확인하였다. 또한, FACS 실험으로 고분자/DNA 복합체인 폴리플렉스의 엔도좀의 탈출 효과가 활발함을 확인하였다. 본 연구를 통해 KKRK-PAMAM G2, KKRKH-PAMAM G2가 효율적인 비바이러스성 벡터의 후보군으로 활용될 수 있는 잠재적 가능성을 확인하였다. In this study, new hybrid polymers composed of polyamidoamine dendrimer generation 2 (PAMAM G2) and KKRK, KKRKH peptides derived from a nuclear localization sequence (NLS) were synthesized and characterized as gene carriers. Using Hep G2 and NIH3T3 cell lines, it was confirmed that the gene transfection efficiency was increased about 20- and 2700-fold in each cell compared to that of the native PAMAM G2. The main endocytosis mechanism of the polymer/plasmid DNA complex (polyplex) was proved to be macropinocytosis through endocytosis inhibition experiments using various inhibitors. It was confirmed that the polyplexes existed around the cell nucleus using a confocal microscope. In addition, it was observed that the polyplex showed active escape from endosomes using FACS analysis using acridine orange. As a result, KKRK-PAMAM G2 and KKRKH-PAMAM G2 show their potential to be one of efficient candidates as non-viral vectors.

유전자 전달에서 양친매성 디펩티드들을 결합시킨 폴리아미도아민 덴드리머의 증대된 형질주입 효율

이제일(Jeil Lee),이선화(Sun Hwa Lee),최준식(Joon Sig Choi) 한국고분자학회 2018 폴리머 Vol.42 No.5

폴리아미도아민(PAMAM) 덴드리머는 널리 이용되는 비바이러스성 벡터 중 하나이며 생분해성 잔기를 가지고 있기 때문에 비교적 낮은 세포독성을 가지고 있는 것으로 알려져 있다. 그러나 빈약한 유전자 전달 능력은 폴리아미도아민 덴드리머의 생체 내 연구 및 임상적용에 장벽으로 작용하고 있다. 이러한 문제를 극복하기 위해서 폴리아미도아민 덴드리머 3차와 4차에 양친매성의 디펩티드인 라이신과 페닐알라닌을 도입하고 형질주입 능력을 평가하였다. 표면개질되지 않은 폴리아미도아민 덴드리머 3차, 4차와 비교할 때, 라이신과 페닐알라닌을 도입한 폴리아미도아민 덴드리머 4차는 HeLa, HepG2 세포 주에서 향상된 형질주입 효율을 나타내었고 또한 비교적 낮은 세포독성을 나타내었다. 이러한 결과들은 고분자들에 라이신, 페닐알라닌 디펩티드의 도입은 빈약한 유전자 전달 능력을 향상시킬 수 있음을 암시한다. Polyamidoamine (PAMAM) dendrimer consisting of biodegradable functional groups is one of the non-viral vectors and has showed relatively low cytotoxicity. However, its poor transfection efficiency acts as a barrier of in vivo research and medical application. To overcome this problem, we introduced amphiphilic KF dipeptides to PAMAM generation 3 and 4, and evaluated transfection efficiency. Compared with those of native PAMAM generation 3 and 4, PAMAM generation 4-KF showed enhanced transfection efficiency in HeLa and HepG2 cell lines. These results imply that introduction of KF dipeptide to polymers can improve the transfection efficiency.

시스타민 코어 폴리아미도아민 덴드리머에 펩티드와 감마 아미노부티르산의 도입을 통한 새로운 유전자 전달체 연구

박혜원(Hye Won Park),유광식(Gwang Sig Yu),송수정(Su Jeong Song),최준식(Joon Sig Choi) 한국고분자학회 2015 폴리머 Vol.39 No.5

유전자 전달체 가운데 비바이러스성 전달체인 PAMAM 덴드리머의 일종인 cPAMAM G4를 사용하여 분자의 표면에 GABA와 히스티딘, 아르기닌을 도입해 새로운 유전자 전달체를 합성하였다. cPAMAM G4-GABA-HR 최종합성물은 핵자기 공명 분광기를 이용해 합성률을 확인하였다. 전기영동을 이용하여 폴리머와 pDNA간의 결합력을 확인하였고, 폴리머와 DNA의 복합체에 환원제를 처리하여 폴리머의 pDNA 방출 능력을 확인하였다. 나노 입자 분석기와 원자력 현미경을 이용하여 pDNA와 폴리머 복합체의 크기와 모양을 분석하였다. HeLa 세포주를 이용한 WST-1 assay를 통해 폴리머의 세포 독성을 확인하였고, 동일한 세포주를 이용한 luciferase assay를 통해 유전자 전달 효율성을 확인하였다. Polyamidoamine (PAMAM) dendrimer are well known non-viral vectors for gene delivery application. Especially, PAMAM conjugated with amino acids have enhanced cell uptake capability and low cytotoxicity compared to other non-viral, vectors. Moreover cystamine core PAMAM has an ability to degrade inside low pH of endosome leading to ease release of biomolecules. In present work, we designed γ-amino butyric acid (GABA), histidine, arginine modified cystamine core PAMAM. GABA has four carbon chains that plays a role as flexible linkers, hence easy interaction with cell membrane. cPAMAM-GABA-HR synthesis was confirmed by 1H NMR. We performed gel retardation assay, DNA release phenomenon of cystamine core PAMAM using DTT. We measured size of polyplex by DLS and AFM. Cytotoxicity was examined by WST-1 assay and transfection efficiency was confirmed using reporter plasmid in HeLa cells.