• 대한약침학회지
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• 제19권1호
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• pp.37-44
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• 2016

Objectives: This study examined the relative efficacies of a derivative of betulinic acid (dBA) and its poly (lactide-co-glycolide) (PLGA) nano-encapsulated form in A549 lung cancer cells in vivo and in co-mutagen [sodium arsenite (SA) + benzo[a]pyrene (BaP)]-induced lung cancer in mice in vivo. Methods: dBA was loaded with PLGA nanoparticles by using the standard solvent displacement method. The sizes and morphologies of nano-dBA (NdBA) were determined by using transmission electron microscopy (TEM), and their intracellular localization was verified by using confocal microscopy. The binding and interaction of NdBA with calf thymus deoxyribonucleic acid (CT-DNA) as a target were analyzed by using conventional circular dichroism (CD) and melting temperature (Tm) profile data. Apoptotic signalling cascades in vitro and in vivo were studied by using an enzyme-linked immunosorbent assay (ELISA); the ability of NdBA to cross the blood-brain barrier (BBB) was also examined. The stage of cell cycle arrest was confirmed by using a fluorescence-activated cell-sorting (FACS) data analysis. Results: The average size of the nanoparticles was ~ 110 nm. Confocal microscopy images confirmed the presence of NdBA in the cellular cytoplasm. The bio-physical properties of dBA and NdBA ascertained from the CD and the Tm profiles revealed that NdBA had greater interaction with the target DNA than dBA did. Both dBA and NdBA arrested cell proliferation at G0/G1, NdBA showing the greater effect. NdBA also induced a greater degree of cytotoxicity in A549 cells, but it had an insignificant cytotoxic effect in normal L6 cells. The results of flow cytometric, cytogenetial and histopathological studies in mice revealed that NdBA caused less nuclear condensation and DNA damage than dBA did. TEM images showed the presence of NdBA in brain samples of NdBA fed mice, indicating its ability to cross the BBB. Conclusion: Thus, compared to dBA, NdBA appears to have greater chemoprotective potential against lung cancer.

• BMB Reports
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• 제41권4호
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• pp.287-293
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• 2008

In a yeast two-hybrid screen, we identified the microtubule-destabilizing protein SCG10 as a potential effector protein of $BRI_3$. The association was verified using GST pull-down, Co-IP, and their perinuclear co-localization. The analysis of in vitro microtubule polymerization/depolymerization showed that the binding of $BRI_3$ to SCG10 effectively blocked the ability of SCG10 to induce microtubule disassembly, as determined by turbidimetric assays. In intact PC12 cells, $BRI_3$ exhibited the ability to stabilize the microtubule network and attenuate the microtubule-destabilizing activity of SCG10. Furthermore, co-expression of $BRI_3$ with SCG10 attenuated SCG10-mediated PC12 cell neurite outgrowth induced by NGF. These results identify a novel connection between a neuron-specific BRI protein and the cytoskeletal network, suggesting possible roles of BRI3 in the process of neuronal differentiation.

• BMB Reports
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• 제43권6호
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• pp.432-437
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• 2010

LBH is a transcription factor as a candidate gene for CHD associated with partial trisomy 2p syndrome. To identify potential LBH-interacting partners, a yeast two-hybrid screen using LBH as a bait was performed with a human heart cDNA library. One of the clones identified encodes ${\alpha}B$-crystallin. Co-immunoprecipitation and GST pull-down assays showed that LBH interacts with ${\alpha}B$-crystallin, which is further confirmed by mammalian two-hybrid assays. Co-localization analysis showed that in COS-7 cells, ${\alpha}B$-crystallin that is cytoplasmic alone, accumulates partialy in the nucleus when co-transfected with LBH. Transient transfection assays indicated that overexpression of LBH or ${\alpha}B$-crystallin reduced the transcriptional activities of p53 and p21, respectively, Overexpression of both ${\alpha}B$-crystallin and LBH together resulted in a stronger repression of the transcriptional activities of p21 and p53. These results showed that the interaction of LBH and ${\alpha}B$-crystallin may inhibit synergistically the transcriptional regulation of p53 and p21.

• 생명과학회지
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• 제22권9호
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• pp.1159-1165
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• 2012

${\gamma}$-Aminobutyric acid (GABA)는 신경세포 밖으로 분비된 후 GABA 수송체들(GATs)에 의하여 다시 신경세포 안으로 재흡수 된다. 그러나, GABA 수송체들이 어떻게 연접전막의 위치에 안정적으로 존재하는지 또한 어떤 단백질과 결합하여 조절을 받는지는 알려져 있지 않다. 본 연구에서 효모 two-hybrid system을 이용하여 betaine-${\gamma}$-aminobutyric acid transporter 1 (BGT-1/mGAT2)의 C-말단과 특이적으로 결합하는 Munc-18-interacting (Mint) 단백질을 분리하였다. BGT-1/mGAT2의 C-말단에 존재하는 "T-H-L" 아미노산배열은 Mint2와의 결합에 필수적으로 관여하였다. Mint2은 BGT-1/mGAT2와는 결합하지만, 다른 종류의 GAT와는 결합하지 않았다. 또한 HEK-293T 세포에 Mint2와 BGT-1/mGAT2을 동시에 발현시켜 면역침강한 결과 두 단백질은 같이 면역침강하였으며, 두 단백질은 세포 내에서 세포막 부위에 같이 존재함도 확인하였다. 이러한 결과들은 Mint2가 BGT-1/mGAT2와 결합하여 BGT-1/mGAT2을 조절하는 역할을 함을 시사한다.

• Journal of Ginseng Research
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• 제46권3호
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• pp.348-356
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• 2022

Background: Gintonin is a ginseng-derived exogenous G-protein-coupled lysophosphatidic acid (LPA) receptor ligand. Gintonin exerts its neuronal and non-neuronal in vitro and in vivo effects through LPA receptor subtypes. However, it is unknown whether gintonin can bind to the plasma membrane of cells and can transactivate the epidermal growth factor (EGF) receptor. In the present study, we examined whether gintonin-biotin conjugates directly bound to LPA receptors and transactivated the EGF receptor. Methods: We designed gintonin-biotin conjugates through gintonin biotinylation and examined whether gintonin-biotin conjugate binding sites co-localized with the LPA receptor subtype binding sites. We further examined whether gintonin-biotin transactivated the EGF receptor via LPA receptor regulation via phosphor-EGF and cell migration assays. Results: Gintonin-biotin conjugates elicit [Ca²⁺]_i transient similar to that observed with unbiotinylated gintonin in cultured PC3 cells, suggesting that biotinylation does not affect physiological activity of gintonin. We proved that gintonin-biotin conjugate binding sites co-localized with the LPA1/6 receptor binding sites. Gintonin-biotin binding to the LPA1 receptor transactivates the epidermal growth factor (EGF) receptor through phosphorylation, while the LPA1/3 receptor antagonist, Ki16425, blocked phosphorylation of the EGF receptor. Additionally, an EGF receptor inhibitor AG1478 blocked gintonin-biotin conjugate-mediated cell migration. Conclusions: We observed the binding between ginseng-derived gintonin and the plasma membrane target proteins corresponding to the LPA1/6 receptor subtypes. Moreover, gintonin transactivated EGF receptors via LPA receptor regulation. Our results suggest that gintonin directly binds to the LPA receptor subtypes and transactivates the EGF receptor. It may explain the molecular basis of ginseng physiology/pharmacology in biological systems.

• 한국의학물리학회지:의학물리
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• 제14권1호
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• pp.34-42
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• 2003

World Wide Web (WWW)에서 Virtual Reality Modeling Language (VRML)를 이용하는 3차원 (3D) 디스플레이는 사용자에게 직관적인 정보를 더 효과적으로 제공해 준다. 웹을 기반으로 하는 해부학적 영상과 융합되는 기능적 영상의 3D 가시화는 아직까지 체계적인 방식으로 연구가 활발히 진행되지 않았다. 이 연구의 목적은 2D 영상들과 함께 웹에서 VRML을 이용하여 구현되는 3D 해부학적 표면 영상들과 기능적 표면 영상들을 동시적으로 관찰할 수 있게 하고 VRML을 통해 만들어진 거리 측정 도구를 가지고 관심영역의 공간적인 위치 정보를 제공하는 것이다. 본 연구에서는 한 명의 간질 환자로부터 Magnetic Resonance (MR) 축면 영상과 발작기 및 발작간기 Single Photon Emission Computed Tomography (SPECT) 축면 영상들을 각각 획득하였다. 발작 진원지의 확인을 향상시키기 위해서 subtractionictal SPECT coregistered to MRI (SISCOM)을 수행하였다 SISCOM 결과로 나타난 각 2D 영상들은 모든 voxel들의 평균값 위로 1-표준편차와 2-표준편차에 해당하는 문턱 이상의 영상 값을 갖도록 하였다. SISCOM으로 나타나는 간질 발작 진원지들과 MRI 영상에서 회색질, 백색질 및 뇌척수액의 경계들을 각각 분할하고 marching cube 알고리즘에 의해 VRML 표면 영상들로 나타내었다. 축면 영상에서 실제 거리를 나타내는 x, y축의 길이를 획득하고 z축선의 길이를 계산하였다. VRML을 이용한 거리 측정도구를 만들어 이전의 VRML 표면 영상들과 융합하였다. MRI 영상을 이용하여 3D 표면 영상들의 단면을 나타내고 3D 표면 영상들의 투명도를 설정하기 위해 Java Script 루틴을 사용자 인터페이스 도구로서 삽입하였다 웹 페이지에서 구현되는 3D 표면 영상들의 투명도와 관찰 위치를 조절함에 따라 모델들 사이의 공간적인 정보를 직관적으로 알 수 있었다. 간질 발작 진원지에 대응하는 해부학적 구조를 3D 표면 영상들을 가로지르는 MRI 평면 영상들을 통해서 확인하였다 간질 발작 진원지는 뇌의 오른쪽 측두엽에서 나타났고 공간적으로 발작 진원지의 실제 위치를 VRML 거리 측정 도구에 의해 알 수 있었다. 결론적으로 본 연구에서 제시하는 웹에 근거한 3D 융합 영상의 가시화와 위치 측정은 진단 및 치료 방사선학과 외과학 등의 분야에서 온라인 방식의 연구와 교육에 있어 많은 도움을 줄 것이다.

• Parasites, Hosts and Diseases
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• 제42권4호
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• pp.185-193
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• 2004

Toxoplasma gondii is an obligate intracellular protozoan parasite, which invades a wide range of hosts including humans. The exact mechanisms involved in its invasion are not fully understood. This study focused on the roles of $Ca^{2+}$ in host cell invasion and in T. gondii replication. We examined the invasion and replication of T. gondii pretreated with several calcium modulators, the conoid extrusion of tachyzoites. Calmodulin localization in T. gondii were observed using the immunogold method, and $Ca^{2+}$ levels in tachyzoites by confocal microscopy. In light microscopic observation, tachyzoites co-treated with A23187 and EGTA showed that host cell invasion and intracellular replication were decreased. The invasion of tachyzoites was slightly inhibited by the $Ca^{2+}$ channel blockers, bepridil and verapamil, and by the calmodulin antagonist, calmidazolium. We observed that calcium saline containing A23187 induced the extrusion of tachyzoite conoid. By immunoelectron microscopy, gold particles bound to anti-calmodulin or anti-actin mAb, were found to be localized on the anterior portion of tachyzoites. Remarkably reduced intracellular $Ca^{2+}$ was observed in tachyzoites treated with BAPTA/AM by confocal microscopy. These results suggest that host cell invasion and the intracellular replication of T. gondii tachyzoites are inhibited by the calcium ionophore, A23187, and by the extracellular calcium chelator, EGTA.

• Molecules and Cells
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• 제26권2호
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• pp.206-211
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• 2008

HI0381 of Haemophilus influenzae was investigated by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy. HI0381 is a 153-residue peptidoglycan-associated outer membrane lipoprotein, and a part of the larger Tol/Pal network. Here, we report its backbone $^1H$, $^{15}N$, and $^{13}C$ resonance assignments, and secondary structure predictions. About 97% of all of the $^1HN$, $^{15}N$, $^{13}CO$, $^{13}C{\alpha}$, and $^{13}C{\beta}$ resonances covering 131 non-proline residues of the 134 residue, mature protein, were clarified by sequential and specific assignments. CSI and TALOS analyses revealed that HI0381 contains five ${\alpha}$-helices and five ${\beta}$-strands. To characterize the structure of HI0381, the effects of pH and salt concentration were investigated by CD. In addition, the structural changes occurring when HI0381 was in a membranous environment were investigated by comparing its HSQC spectra and CD data in buffer and in DPC micelles; the results showed that helix ${\alpha}4$ and strand ${\beta}4$ became aligned with the membrane. We conclude that the conformation of HI0381 is affected by the membrane environment, implying that its folded state is directly related to its function.

• ALGAE
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• 제18권2호
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• pp.121-127
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• 2003

The pyrenoid ultrastructure and distribution of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and Rubisco activase in the chloroplasts of Chlamydomonas reinhardtii was studied using the immunogold localization technology with electron microscopy. There were several tubular thylakoids invading in the pyrenoid matrix to form several spokewise channels. The connections between pyrenoid matrix and stroma of chloroplast were the partial of channels. The starch sheath surrounding the pyrenoid was separated into several parts by the connections in transection. Some thylakoids were packed together near the connections in one side of the pyrenoid. Those special structures might be used to transport substance between pyrenoid and stroma of chloroplasts. With the antibody raised against the large subunits of Rubsico from C. protothecoides, the result of the gold immunolocalization of Rubisco in Chlamydomonas reinhardtii showed most of the gold particles heavily labeled the pyrenoid matrix, as well as the starch sheath matrix, and very few in the stroma of chloroplasts. The gold particle density was 880.00 $\pm$ 164.32, 190.00 $\pm$ 152.39 and 9.60 $\pm$ 5.37 ${\mu}m^{-2}$ in pyrenoid matrix, starch sheath and stroma region of chloroplast respectively (background: 5.67 $\pm$ 1.53 ${\mu}m^{-2}$). 99.59% of the total Rubiscos was calculated to be concentrated in the pyrenoid matrix and starch sheath by spatial densities. The gold immunolocalization of Rubisco activase also showed that Rubisco activase was mainly concentrated in the periphery of the pyrenoid and the starch sheath (the density was as high as 229.69 $\pm$ 96.96 ${\mu}m^{-2}$). There were very few gold particles located in the stroma of chloroplasts. These results indicated that pyrenoid surface and starch sheath was the site for Rubisco activation and $CO_2$ fixation, which supported the suggestion that pyrenoids perform photosynthesis function.

• 대한전자공학회논문지SP
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• 제42권1호
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• pp.71-77
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• 2005

본 논문에서는 5.1채널 입체음향 오디오 신호를 2채널의 헤드폰으로 재생하기 위한 HRTF (Head Related Transfer Function) 기반의 입체음향 생성 시스템에 대하여 다룬다. 각 채널의 모노 입력신호는 HRTF를 이용한 바이노럴(binaural) 필터링을 통해 가상적으로 음상정위되며, 입체감과 공간감을 증가시키기 위해 잔향효과가 추가된다. 연산량 감소를 위해 음상정위 성능을 저하시키지 않는 범위에서 HRTF의 임펄스 응답 탭 수를 줄였으며, 잔향효과를 위한 음장제어부에서는 초기반사열중 주요한 성분만을 지연기로 모델링하였다. 또한 비개인화된 HRTF DB에 의란 앞/뒤 혼돈 문제를 줄이기 위하여 앞/뒤 스펙트럼의 차를 가중치로 하여 HRTF 스펙트럼을 강조하는 방법을 적용하였다. 구현한 시스템의 성능 평가 결과, 단순한 스테레오 방법이나 2채널 Down Mixing 방식에 비해 현실감 있고 방향성 있는 입체음향을 느낄 수가 있었다.