• Biomolecules & Therapeutics
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• 제3권4호
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• pp.266-272
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• 1995

The analysis of membrane receptors for hormones and neurotransmitters has progressed considerably by pharmacological and biochemical means and more recently through the use of specific antibodies. To generate and characterize a moloclonal antibody against $\beta$-adrenergic receptor, a synthetic $\beta$2-adrenergic receptor peptide (Phe-Gly-Asn-Phe-Trp-Cys-Phe-Trp-Thr-Ser-lle-Asp-Val-Leu) which may comprise part of $\beta$-adrenergic receptor ligand binding pocket was coupled to Keyhole Limpet Hemocyanin (KLH) and used as an immunogen. Male BALB/C mice were immunized with this antigen and the immunized spleen was fused with myeloma SP2/0-Ag14 cells to produce monoclonal antibodies. Two clones were obtained but one of monoclonal antibodies, mAb5G09, was used throughout in this study because the other clone, mAb5All showed weak immunoreactivity against KLH as well. The mouse monoclonal antibody mAb5G09 produced in this study showed immunoreactivity to peptide-KLH conjugates and also to human A43l cells and guinea pig lung $\beta$2-adrenergic receptor as revealed by ELISA and western blot. In the course of determination of the effects of mAb5G09 on $\beta$-receptor ligand binding, it was observed that mAb5G09 specifically bound $\beta$-adrenergic radioligand [$^3$H]dihydroalprenolol (DHA) with a dissociation constant (Kd) of 60 nM. The [$^3$H]DHA binding activity of mAb5G09 had characteristics of immunoglobulins and the binding activity was not observed in the control anti-KLH monoclonal antibody. The monoclonal antibody, mAb5G09 produced in this study may provide useful models for the study of the structure of receptor binding sites.

• 대한약리학회지
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• 제29권1호
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• pp.85-96
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• 1993

화학물질에 의한 간암 유발과정에서 transferrin receptor (TfR)의 변동을 밝히기 위해 간을 부분절제한 정상백서의 재생간과 발암물질로 3-Me-DAB를 8주간 투여한 백서 또는 약물 투여 후 부분 간절제 수술을 행하여 세포분열을 유도시킨 백서 간조직으로부터 parenchymal cell (PC)과 nonparenchymal cell (NPC)를 분리하고 각각의 세포막을 제조하여 $^{125}I-transferrin$ 결합실험을 실시한 바 다음과 같은 성적을 얻었다. 1. 3-Me-DAB 투여에 의하여 간조직에서 oval cell의 증식, 재생성 변화, 결절형성, 담관의 증식 및 담관세포암 등의 현저한 조직학적 변화가 동반되었다. 그러나 간세포증식을 더욱 촉진시키기 위하여 부분간절제 수술을 하였을 때 수술 후 경과에 따른 형태학적 변동은 큰 차이가 없었다. 2. 정상 재생간의 PC 및 NPC homogenate에서 transferrin 결합량은 부분간 절제 수술 후 1일 및 3일에 증가되었으며 수술 후 7일에 정상으로 회복되었다. 3-Me-DAB 투여에 의해 두세포군에서 모두 정상세포보다 높게 나타났으며 재생기간에 따라 계속 증가되었다. 3. 정상간의 NPC 세포막에서 transferrin 최대 결합량 (Bmax)은 PC 세포막에서 보다 많이 분포되어 있었으며, Kd는 양세포막에서 5.05 또는 6.3nM로 비슷하였다. 4. 재생간의 NPC 및 PC 세포막에서 transferrin 결합량은 부분 간절제 수술 후 1일 및 3일에 $40{\sim}50%$ 증가되었고 수술 후 7일에 정상치로 회복되었다. 5. 3-Me-DAB 처치에 의하여 NPC 및 PC 세포막의 transferrin 결합량은 정상 간세포막에서 보다 약 3배 증가되었고, 3-Me-DAB 투여후 재생간의 NPC 세포막에서는 부분 간절제 수술 후 3일까지 증가된 후 감소되는 양상인데 반해 PC 세포막에서는 수술 후 7일까지 계속 증가되었다. 6. 3-Me-DAB 투여 후 NPC 및 PC 세포막 transferrin binding site에서 Kd치가 $3.1{\sim}4.1\;nM$과 $25.4{\sim}54.1\;nM$인 두 종류가 존재하는 것으로 나타났다. 이상의 실험성적으로 TfR는 1) 간조직의 PC 및 NPC 세포에 모두 분포되어 있으며, 2) 정상 재생간 및 3-Me-DAB의 처리 후 간세포에서의 세포막 TfR의 증가는 세포내 합성량의 증가에 의하여 일어나며, 3) 정상 재생간의 세포막 TfR는 한 종류의 high affinity site $(Kd,\;<3.1{\sim}7.5\;nM)$에 의하여 증가되나, 3-Me-DAB 처리 후 간세포막에서는 정상에서와 같은 high affinity형 이외에 affinty가 낮은 다른 형태의 TfR $(Kd,\;25.4{\sim}54.1\;nM)$가 세포막으로 출현됨으로써 크게 증가되는 것으로 사료된다.

• 대한의생명과학회지
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• 제16권4호
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• pp.221-227
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• 2010

To elucidate the invasion mechanism of pathogenic Naegleria fowleri, especially a receptor-ligand recognition, we investigated the in vitro cytotoxicity of pathogenic N. fowleri and nonpathogenic N. gruberi to murine macrophages, RAW 264.7, by adding four kinds of saccharides, ${\alpha}$-fucose, ${\beta}$-galactose, ${\alpha}$-D-mannopyranoside (${\alpha}$-mannose) and xylose. There was not enough of a difference in the cytotoxicity of N. fowleri treated with 10 mM of each saccharide. In particular, the cytotoxicity of N. fowleri was highly inhibited by 100 mM ${\alpha}$-mannose, which was 62.3% inhibition calculated by the analysis of lactate dehydrogenase (LDH) release assay. Although murine macrophages were not significantly destroyed by nonpathogenic N. gruberi under hematoxylin staining, the cytotoxicity of N. gruberi was inhibited from 31.5% to 14.5% (P<0.01) by 100 mM ${\alpha}$-mannose treatment. The binding of N. fowleri to macrophages was inhibited from 33% to 50% by 100 mM ${\alpha}$-mannose. Furthermore, as results of the adhesion assays which were performed to determine whether binding of Naegleria is mediated by saccharides-binding protein, the binding ability of N. fowleri as well as N. gruberi was inhibited by 100 mM ${\alpha}$-mannose.

• International Journal of Oral Biology
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• 제34권4호
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• pp.191-197
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• 2009

Somatostatin (SST) is a known neuromodulator of the central nervous system. The substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc) receives many thinmyelinated $A{\delta}$-fiber and unmyelinated C primary afferent fibers and is involved in nociceptive processing. Many studies have demonstrated that SST plays a pivotal role in pain modulation in the spinal cord. However, little is yet known about the direct effects of SST on the SG neurons of the Vc in adult mice. In our present study, we investigated the direct membrane effects of SST and a type 2 SST receptor agonist, seglitide (SEG), on the SG neurons of the Vc using a gramicidin-perforated current clamp in adult mice. The majority (53%, n = 27/51) of the adult SG neurons were hyperpolarized by SST (300 nM) but no differences were found in the hyperpolarization response rate between males and females. When SST was applied successively, the second response was smaller ($76{\pm}9.5%$, n=19), suggesting that SST receptors are desensitized by repeated application. SST-induced hyperpolarization was also maintained under conditions where presynaptic events were blocked ($75{\pm}1.0%$, n=5), suggesting that this neuromodulator exerts direct effects upon postsynaptic SG neurons. SEG was further found to induce membrane hyperpolarization of the SG neurons of the Vc. These results collectively demonstrate that SST inhibits the SG neuronal activities of the Vc in adult mice with no gender bias, and that these effects are mediated via a type 2 SST receptor, suggesting that this is a potential target for orofacial pain modulation.

• Asian-Australasian Journal of Animal Sciences
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• 제28권8호
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• pp.1187-1193
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• 2015

This study determined the effects of dietary restriction on growth and the expression of lipid metabolism and growth hormone signaling genes in the longissimus dorsi muscle (LM) of Korean cattle. Thirty-one Korean cattle steers (average age 10.5 months) were allocated to normal (N; n = 16) or dietary restriction (DR; n = 15) groups. The feeding trial consisted of two stages: for the 8-month growing period, the DR group was fed 80% of the food intake of the normal diet, and for the 6-month growth-finishing period, the DR group was fed a DR total mixed ration with 78.4% of the crude protein and 64% of the net energy for gain of the normal diet. The LM was biopsied 5 months (period 1 [P1] at 15.5 months of age) and 14 months (period 2 [P2] at 24.5 months of age) after the start of feeding. The mRNA levels were determined using real-time polymerase chain reaction. Body weight, daily feed intake, average daily gain, and feed efficiency were lower in the DR group compared with the normal group at both P1 and P2. At P1, the lipogenic fatty acid synthase (FASN) mRNA levels were lower (p<0.05) in the DR group compared with the normal group. The DR group tended (p = 0.06) to have higher of levels of growth hormone receptor (GHR) mRNA than the normal group. At P2, the DR group tended to have lower (p = 0.06) androgen receptor (AR) mRNA levels than the normal group. In conclusion, our results demonstrate that dietary restriction partially decreases the transcription of lipogenic FASN and growth hormone signaling AR genes, but increases transcription of the GHR gene. These changes in gene transcription might affect body fat accumulation and the growth of the animals.

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2003년도 정기총회 및 학술발표회
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• pp.51-51
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• 2003

The medicinal plant Cimicifuga Racemosa (Black cohosh) has been used to treat many kinds of neuronal and menopausal symptoms, such as arthritis, menopausal depression, nerve pain, etc. Here, we examined the effect of Cimicifugoside (CF), one of triterpene glycosides which have been known as pharmacologically active ingredients of C. Racemosa, on nicotinic acetylcholine receptor (nAChR)-mediated catecholamine (CA) secretion in bovine adrenal chromaffin cell. Cimicifugoside inhibited calcium increase induced by 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), a nAChR agonist with a half maximal inhibitory concentration (IC50) of 18${\pm}$2${\mu}$M. In contrast, cimicifugoside did not affect the calcium increases evoked by high K$\^$+/, veratridine, and bradykinin. The DMPP-induced sodium increase was also inhibited by cimicifugoside with IC50 of 2${\pm}$0.3${\mu}$M, suggesting that the activity of nAChRs is inhibited by cimicifugoside. Cimicifugoside did not effect on the KCl-induced secretion but markedly inhibited the DMPP-induced catecholamine secretion which was monitored by carbon-fiber amperometry in real time, and by high performance liquid chromatography (HPLC) through electrochemical detection. The results suggest that cimicifugoside selectively inhibits nAChR-mediated response in bovine chromaffin cells.

• BMB Reports
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• 제47권10호
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• pp.552-557
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• 2014

The main purpose of this study was to investigate whether type 3 muscarinic acetylcholine receptor (M3R) dysfunction induced vascular hyperpermeability. Transwell system analysis showed that M3R inhibition by selective antagonist 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) and small interfering RNA both increased endothelial permeability. Using coimmunoprecipitation and Western blot assay, we found that M3R inhibition increased VE-cadherin and ${\beta}$-catenin tyrosine phosphorylation without affecting their expression. Using PTP1B siRNA, we found that PTP1B was required for maintaining VE-cadherin and ${\beta}$-catenin protein dephosphorylation. In addition, 4-DAMP suppressed PTP1B activity by reducing cyclic adenosine monophosphate (cAMP), but not protein kinase $C{\alpha}$ ($PKC{\alpha}$). These data indicate that M3R preserves the endothelial barrier function through a mechanism potentially maintaining PTP1B activity, keeping the adherens junction proteins (AJPs) dephosphorylation.

• 통합자연과학논문집
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• 제4권4호
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• pp.273-277
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• 2011

It's a threat for the public health that H1N1 (Influenza virus A) causes disease and transmits among humans. WHO (world health organization) declared that the infections caused by the new strain had reached pandemic proportions. The approved neuraminidase inhibitors (Zanamivir and Oseltamivir) and related investigative drug (BCX-1812) are potent, specific inhibitors of influenza A and B viruses. These drugs are highly effective to prevent influenza A and B infections. Early therapeutic use reduces illness duration and respiratory complications. Recently, we found one of the potent inhibitor of erystagallin A ($IC_{50}$ of 2.04 ${\mu}M$) for neuraminidase target, this inhibitor shows most similar structure to its natural substrate, sialic acid. Therefore, we chose 1l7f to get the receptor structure for docking study among many crystal structures. A docking study has been performed in Surflex-Dock module in SYBYL 8.1. In the present study, we attempt to compare the docking studies of pterocarpin and erystagallin A with neuraminidase receptor structure. In the previous report, the methoxy group of pterocarpin had H-bonding with Arg residues. The present docking results for erystagallin A showed the backbone of hydroxyl group shows significant H-bonding interactions with Arg152 and Arg292. The results showed that erystagallin A interacts more favorably with distinctive binding site rather than original active site. Therefore, we tried to reveal plausible binding mode and important amino acid for this inhibitor using docking and site id search calculations of Sybyl. The results obtained from this work may be utilized to design novel inhibitors for neuraminidase.

• Asian Pacific Journal of Cancer Prevention
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• 제15권18호
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• pp.7959-7964
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• 2014

The significance of the single hormone receptor positive phenotype of breast cancer is still poorly understood. The use of hormone therapy has been found to be less effective for this type, which has a survival outcome midway between double positive and double negative phenotypes. The aim of this study was to investigate differences in patient and tumor characteristics and survival between double-receptor positive (ER+PR+), double receptor negative (ER-PR-) and single receptor positive (ER+PR- and ER-PR+) breast cancer in an Asian setting. A total of 1,992 patients with newly diagnosed stage I to IV breast cancer between 2003 and 2008, and where information on ER and PR were available, were included in this study. The majority of patients had ER+PR+ tumors (n=903: 45.3%), followed by 741 (37.2%) ER-PR-, 247 (12.4%) ER+PR-, and 101 (5.1%) ER-PR+ tumors. Using multivariate analysis, ER+PR- tumors were 2.4 times more likely to be grade 3 compared to ER+PR+ tumors. ER+PR- and ER-PR+ tumors were 82% and 86% respectively less likely to be grade 3 compared with ER-PR- tumors. ER-PR+ tumours were associated with younger age. There were no survival differences between patients with ER+PR+ and ER-PR+ tumors. However, ER+PR- tumors have poorer survival compared with ER+PR+ tumours. ER-PR- tumours had the worst survival. Adjuvant hormonal therapy with tamoxifen was found to have identical survival advantage in patients with ER+PR+ and ER-PR+ tumors whereas impact was slightly lower in patients with ER+PR- tumors. In conclusion, we found ER+PR- tumors to be more aggressive and have poorer survival when compared to ER+PR+ tumors, while patients with ER-PR+ tumours were younger, but had a similar survival to their counterparts with ER+PR+ tumours.

• 한국동물위생학회지
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• 제30권4호
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• pp.557-562
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• 2007

This study was carried out to discriminate Hanwoo from the milking and hybrid cattle by detection of MC1R gene related to bovine hair color. One hundred sixty six samples were collected from the abattoir (n = 106) and local market (n = 60). The beef from abattoir were originated from Hanwoo (n=27), Holstein (n=29), Hybrid (n=45) and imported cattle (n=5), respectively. The beef from market consisted of Hanwoo (n=36), Holstein (n=7) and imported ones (n=17). Commercialized screening kit (Kogenebiotec, Korea) was used for MC1R gene analysis. As a result, Hanwoo was discriminated from Holstein. However, 9 of 45 hybrid and 11 of 22 imported beef samples were indistinguishable from Hanwoo. It could be explained by second generation of crossing of Hanwoo with Holstein or the cattle with silver or yellow hair. This results suggest that additional tests as well as MC1R gene detection be needed to confirm Hanwoo beef among cattle beef.