• 약학회지
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• 제55권3호
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• pp.227-233
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• 2011

We previously reported the protein isolated from Coptidis Rhizoma (CRP), which has antifungal activity against a fungal pathogen, Candida albicans. In the current study, we investigated what portion in the CRP is responsible for the antifungal activity. For the investigation, the CRP was fractionated on a Shepadex G-50 column. Data resulting from the fractionation, seven fractions were obtained. Fractions (Fr.) I, II, and III eluted initially from the column showed no inhibitory effect on the growth of C. albicans, whereas Fr. IV, V, and VI eluted later revealed inhibition of the growth, and Fr. IV and VI showed potent antifungal activity by broth susceptibility analysis. However, Fr. VI was contained in the CRP more than Fr. IV, which led us to select the VI for the following experiments. In a murine model of a subcutaneous candidiasis caused by C. albicans, the Fr. VI displayed a therapeutic effect on nude mice pretreated with anti-neutrophil monoclonal antibody (RB68C5) and then infected subcutaneously with live C. albicans. At day 16, these mice were healed almost up to 78% of the infected area when compared to infected area of control nude mice that received diluent (Dulbecco's Phosphate-Buffered Saline; DPBS), instead of the Fr. VI (P<0.01). The Fr. VI blocked hyphal formation from blastoconidial form of C. albicans (P<0.01), which might prevent penetration of hyphae to the deeper site of skin and thus helps the healing. In the ionic strength test, the effect of Fr. was influenced by $Ca^{2+}$ ion just like other known antimicrobial peptides, but the influence was affected at an extremely high concentration such as 500 mM. Thus, such ion-concentration is considered to be meaningless in the clinical situation. Considering all data together, Coptidis Rhizoma is appeared to produce an antimicrobial peptide that has therapeutic effect on subcutaneous infection caused by C. albicans.

• The Korean Journal of Physiology
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• 제23권1호
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• pp.89-98
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• 1989

New Zealand white 계의 토끼에게 EDC를 사용하여 hemocyanin에 접합시킨 human G-17-I을 면역한 결과 gastrin의 방사면역측정에 적합한 항 gastrin 혈청을 얻는데 성공하였다. 본 실험에서 얻은 항 gastrin 혈청의 역가는 1:25,000이며 human G-17-I과 human big gastrin에 매우 친화력이 높은 반면 다른 위장관 peptide 들과는 교차반응성이 아주 낮았다. 본 항 gastrin 혈청의 effective affinity constant는 $1.77{\times}10^{11}/M$이었다. 또한 총 결합부위(total binding sites)는 255 nM, 이질성지수(heterogeneity index)는 0.84 그리고 average affinity constant는 $0.79{\times}10^{11}/M$이었다. 본 실험에서 얻은 항 gastrin 혈청을 사용하여 사람과 흰쥐의 혈장 gastrin 농도를 방사면역측정법으로 측정한 결과 이미 잘 알려진 결과와 동일한 성적을 얻을 수 있었다. 이러한 결과들을 종합하여 볼때 본 실험에서 얻은 항 gastrin 혈청은 생리적농도로 존재하는 gastrin의 방사면역측정에 적합한 것으로 생각된다.

• BMB Reports
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• 제29권1호
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• pp.27-31
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• 1996

Using topoisomerase II (topo II) isozyme-specific antibodies, we investigated the phosphorylation of topo $II{\alpha}$ in mitotic HeLa cells. Topo $II{\alpha}$ was specifically modified in the mitotic cells, resulting in slow migration on SDS-polyacrylamide gel electrophoresis. To characterize the nature of this modification, we treated the nuclear extracts prepared from the mitotic cells with alkaline phosphatase. After the treatment with alkaline phosphatase, the slowly migrated band disappeared and instead a normal (170 kDa) topo $II{\alpha}$ band appeared. These results indicate that human topo $II{\alpha}$ is modified at a specific site(s) in M phase by phosphorylation, supporting the possibility that M phase-specific phosphorylation of topo II is critical for mitotic chromosome condensation and segregation.

• Journal of Yeungnam Medical Science
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• 제32권2호
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• pp.127-131
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• 2015

Churg-Strauss syndrome (CSS) is a necrotizing vasculitis with extra-, peri-vascular eosinophilic infiltration. Chronic symmetric polyarthritis with the presence of rheumatoid factor (RF) and anti-cyclic citrullinated peptide antibody are the mainstay of rheumatoid arthritis (RA) diagnosis. Mononeuritis multiplex is a peripheral neuropathy involving more than 2 separate nerve areas. A 62-year-old male patient was referred for left foot drop and polyarthritis of both hands and feet for 4 months. During evaluation, mononeuritis multiplex was detected on nerve conduction study and electromyography tests: vasculitis with neutrophil, eosinophil, and lymphocyte infiltration on peroneal nerve biopsy. A positive response to methacholin and bronchodilator was observed on the pulmonary function test. Radiologic tests showed peri-articular soft tissue swelling and osteopenia on both hand and foot. Marked peripheral eosinophilia, high RF, and positive perinuclear anti-neutrophil cytoplasmic antibody were detected on blood tests. Here, we report on a patient with overlap syndrome of CSS and RA with review of the relevant literature, from which a few references to overlap syndrome of CSS and RA were available.

• 한국물환경학회지
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• 제23권5호
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• pp.705-711
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• 2007

Green quantum-dot nanocrystal (QD525) with anti-microcystin monoclonal antibody was applied for detection of microcystin, a monocyclic peptide hepatotoxin, extracted from the culture of Microcystis aeruginosa. The presence of microcystin in the cell lysate was verified by HPLC analysis with UV absorbance at 238 nm. Microcystis cell extract exhibited fluorescence emission spectra, which peak was around 460 nm because of their complex organic substances. When a spherical QD525 antibody conjugates (10~20 nm in diameter) were bound to the microcystins in the Microcystis cell lysate, the fluorescence intensity of the primary peak at 525 nm diminished while the secondary emission peak at 460 nm slightly increased intensities. It is due to energy transfer from the primary (major) to the secondary (minor) peak, resulting from physical deformation of QD525 and different environmental factors. On the other hand, other cell extracts did not show any fluorescence emission change. This study is very available for detecting and monitoring the microcystin because it is one step assay without washing step and portable spectrophotometer makes on-site measurement possible. For health risk assessment of the microcystin, the reliable and rapid system to detect and quantify microcystin is seriously required.

• Journal of Pharmaceutical Investigation
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• 제29권2호
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• pp.87-92
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• 1999

Drug delivery to the brain may be achieved by producing chimeric peptide, attaching the drug to protein 'vectors' which are transported into the brain from the blood by a receptor-mediated transcytosis through the blood-brain barrier (BBB). Since the BBB expresses high concentrations of transferrin receptor, and it was reported that anti-transferrin receptor mouse monoclonal antibody (OX26) undergoes transcytosis through the BBB, it is logical to assume that a drug delivery system via transferrin receptor-mediated transcytosis is a promising strategy. In the present study, therefore, we tested feasibility of several OX26 based vectors for the brain delivery of a model drug. Avidin-based delivery vectors such as OX26-streptavidin (OX26-SA), OX26-neutralite avidin (OX26-NLA) were chemically synthesized vectors and OX26 immunoglobulin G 3 type $C_{H}3$ fusion avidin $(OX26\;IgG3C_H3-AV)$ was genetically engineered. To improve the efficiency of producing chimeric peptide, we used avidin-biotin technology. Pharmacokinetics of $[^3H]biotin$ bound to OX26-SA, OX26-NLA and $OX26\;IgG3C_H3-AV$ was determined by intravenous injection technique, and their stabilities in plasma were analyzed using HPLC. The brain delivery of $[^3H]biotin$ bound to OX26-SA, OX26-NLA and OX26\;$IgG3C_{H}3-AV$ (expressed as %ID/g brain) was $0.22{\pm}0.01$, $0.18{\pm}0.01$ and $0.25{\pm}0.09$, respectively. The areas under the plasma concentration versus time curve (AUC) for OX26-SA, OX26-NLA, $OX26\;IgG3C_H3-AV$ from time zero to 60 min were $209{\pm}10$, $195{\pm}9$, $134{\pm}29\;%ID\;min/ml$ respectively and their total clearances $(CL_{tot})$ were $1.00{\pm}0.09$, $1.08{\pm}0.07$ and $1.54{\pm}0.29\;ml/min/kg$, espectively. These results showed that these vectors possess preferable pharmaceutical (e.g., resonable stability) and pharmacokinetics (e.g., significant brain uptake and enhanced AUC) for brain delivery. Therefore, these vectors may be broadly useful in the brain delivery of drugs that are not transported into the brain to a significant extent.

• Restorative Dentistry and Endodontics
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• 제24권2호
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• pp.372-380
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• 1999

The purpose of this study was to investigate the distribution of calcitonin gene-related peptide(CGRP) containing nerve fivers after pulp exposure in rats. The Spague-Dawley rats weighing about 250 - 300g were used. The animals were devided into normal control group and experimental groups. Experimental animals were sacrified on 2, 4, 7, 10 days after pulp exposure. The maxillary teeth and alveolar bone were removed and immersed in the 4% paraformaldehyde plus 0.1M phosphate buffer (pH 7.4). Serial frozen $50{\mu}m$ thick sections were cut with a cryostat. In the immunohistochemical staining procedure, the rabbit CGRP antibody was used as a primary antibody. The sections were incubated for 48 hours at $4^{\circ}C$, and placed into biotinylated anti-rabbit IgG as a secondary antibody and incubated in ABC (avidin-biotin complex), The sections were visualized by 0.05% 3.3 diaminobenzidine tetrahydrochloride. The results of this study were as follows: 1. In control group, CGRP containing nerve fibers ran parallel to the long axis of root and reached the coronal pulp. They were distributed on Raschkow plexus under the odontoblastic layer. 2. In 2 day group after pulp exposure, tissue necrosis and acute inflammation occurred and CGRP containing nerve fibers increased. In 4 day group, the necrotic tissue extended to the pulp and CGRP containing nerve fibers were distributed around the inflammation zone. 3. In 7 day group after pulp exposure, pulp necrosis occurred, and in 10 day group, the abscess under the necrotic pulp extended to the root apex area and CGRP containing nerve fibers were not observed in root canals. 4.The sprouting of CGRP nerve fibers was most remarkable at the pulp chamber under injury in 4 day group, and it was found at inflammation zone under the necrotic tissue in 7 day group and the remaining root pulp tissue in 10 day group. As mentioned above, CGRP nerve fibers had a tendency to increase around the inflammatory zone, especially around the acute inflammation tissue, when compared with control group. It is suggested that CGRP nerve fibers maybe related to the control of inflammatory response of pulp tissue.

• Journal of Plant Biotechnology
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• 제40권3호
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• pp.169-177
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• 2013

최근 급속도로 당뇨병 환자가 증가하면서 인슐린 시장이 크게 성장하고 있다. 또한 최근 식물체를 이용하여 의약용 단백질 생산이 경제적인 측면과 안정성 측면에서 매우 효과적임이 보고되고 있어 이를 이용한 분자농업이 주목을 받고 있다. 본 연구에서는 인슐린 단백질을 식물체에서 생산하기 위한 유전자 발현 construct를 설계하기 위한 실험으로서 식물발현용 preprominiinsulin construct를 제조하기 위한 단계적 실험을 수행하였다. 우선 proinsulin이 무세포 식물 전사/번역시스템에서 성공적으로 발현됨을 확인하였다. Prominiinsulin construct를 제조하여 대장균에서 발현시키는데 성공하였으며, 이를 트립신으로 절단한 후 인간 항인슐린 항체를 이용한 western 분석을 통하여 효과적으로 A-펩타이드와 B-펩타이드가 형성되며 이후 적절하게 접힘이 일어나고 hexamer로 조립됨을 확인하였다. 이후 식물체에서 재조합 인슐린 유전자가 발현되는지를 확인하기 위하여 RFP 결합 construct를 제조하여 담배의 현탁배양세포인 BY-2 세포에 형질전환시켜 RFP 결합 preprominiinsulin이 성공적으로 발현됨을 확인 하였다. 이러한 성공적인 연구 결과를 토대로 향후 이 construct는 RFP 단백질을 제거하여 35S 프로모터에 직접 유도되는 [N 말단 ${\rightarrow}$ tobacco E2 시그널 펩타이드 ${\rightarrow}$ B-펩타이드(1-29 AA) ${\rightarrow}$ AAK ${\rightarrow}$ A-펩타이드(1-21 AA) ${\rightarrow}$ RR ${\rightarrow}$ His6 ${\rightarrow}$ KDEL ${\rightarrow}$ C 말단] construct를 제조하여 담배 식물체에 형질전환시켜 분자농업을 통해 인간 인슐린 단백질을 생산하는데 활용하고자 한다.

• 한국가축번식학회지
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• 제25권2호
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• pp.119-124
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• 2001

본 연구는 한우 난포발달에 있어서 난포액 및 inhibin의 생리적 역할을 검토하기 위해 수행되었다. Anti-inhibin serum(AI)생산을 위해 사용된 항원은 porcine inhibin-$\alpha$-subunit 19~32의 peptide를 사용하여 adjuvant 용액을 혼합, 앙고라종 토끼 5두(체중 2.5kg)에게 주 2회 간격으로 면역 실시 후 52일째의 토끼로부터 항혈청을 생산하였다. 과립막 세포의 체외배양을 위해 D-MEM(10% FCS와 antibiotics를 첨가)을 배양액으로 하여 1$\times$$10^{6}$ cells/$m\ell$로 조절하였으며, 호르몬은 RIA 및 ELISA법으로 분석하였다. Western blotting법에 의해 과립막 세포 및 황체조직의 각 세포질을 SDS-PAGE로 분리하여 nitro cellulose membrane에 transfer하여 검토한 결과, 직경 1.0 cm의 성숙 난포의 granulosa cell의 세포질에서 특이하게 Inhibin이 존재하고 있음이 확인되었으나, 황체조직 및 성숙 난포에서는 검출되지 않았다. 난포 크기별 난포액의 progesterone 및 estradiol-17$\beta$을 농도를 분석한 결과, estradiol-17$\beta$농도는 난포 크기가 직경 2.0 cm부터 유의적으로 높았으나 난포 크기가 적을수록 감소되었다. 이에 반해, Progesterone 농도는 직경 2.0 cm 난포에서 가장 높았으며 난포 크기가 적을수록 낮았다. 과립막 세포의 48시간 체외배양에서 bFF 5% 처리구와 bFF 5%+AI 5% 처리구에서는 progesterone은 대조구보다 유의적으로 억제되었으나, AI 5% 단독 처리구에서는 대조구와 큰 차이가 없었다. 또한, estradiol-17$\beta$농도는 5% AI구와 5% AI+5% bFF 처리구에서는 대조구에 비해 증가하였다. 그러나, 5% bFF 단독처리구에서는 대조구와 큰 차이가 없었다. 이상의 결과로, 한우에 있어서 성숙난포에 존재하는 Inhibin은 AI처리에 의해 내인성 Inhibin의 기능이 약하되어 FSH분비를 조절하는 역할을 함으로써 난포발달 및 난포세포의 스테로이드호르몬합성에 중요하게 관여하고 있는 것으로 사료된다.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 2001년도 추계학술대회 및 정기총회
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• pp.102-102
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• 2001

Taurine (2-ethaneaminosulfonic acid) is one of the major intracellular ${\beta}$ -amino acids in mammals and is required for a number of biological processes including membrane stabilization, osmoregulation, antioxidation, detoxification, modulation of calcium flux and neurornodulation. The taurine transporter (TAUT) which contains 12 hydrophobic membrane-spanning domains has been cloned from dog kidney, rat brain, mouse brain, human thyroid, placenta and retina. In this study, The TAUT cDNA from the human intestinal epithelial cell, HT-29 was cloned and sequenced. Reverse-transcription polymerase chain reaction (RT-PCR) was performed to amplify partial cDNA encoding human intestinal TAUT. The coding region of the PCR product was 732 bp long. The primers were designed to encode highly conserved amino acid sequences near the transmembrane domains III (IPYFIFLF) and Ⅵ (KYKYNSYR) both in human and mouse. The TAUT cDNA amplified was ligated into the pGEX 4T-1 expression vector. The resulting sequence of human intestinal TAUT cDNA (Accession number of NCBI Genebank is AF346763) was identical to the sequences of the TAUTs previously determined in the human placenta and retina except 3 base pairs from that of the reported human thyroid. TAUT specific antibodies were generated to use them as biological tools in the studies of the biological role of TAUT. Peptides of 149-162 amino acid residue (14 amino acids) of the TAUT were synthesized. The synthetic peptide used in this study was LFQSFQKELPWAHC. This region was chosen not only to avoid putative glycosylation sites but also to exclude regions of known homology with GABA transporters in the extracellular hydrophilic domains. The synthetic peptide, TAUT-1 was conjugated with carrier protein, kehole lympet hemocyanin (KLH) to use as an antigen. When used for immunization on a rabbit to produce polyclonal antiserum, the conjugates elicited high -titered specific anti-TAUT-1 antibodies, which reacted well with the ovalbumin (OVA) conjugated peptides in ELISA. The KLH-conjugated peptide was also used as immunizing antigen in BALB/c mice to produce TAUT specific monoclonal antibodies. From the culture supernatant of the hybridoma, the specificity of anti-TAUT-1 monoclonal antibodies was confirmed by ELISA. Further applications of more tools in TAUT expression analysis will be performed such as western blotting and flow cytometry.