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( Yee-seul So ),( Dong-hoon Yang ),( Kwang-woo Jung ),( Won-ki Huh ),( Yong-sun Bahn ) 한국미생물 · 생명공학회 2017 Journal of microbiology and biotechnology Vol.27 No.2
In this study, we aimed to generate a series of versatile tagging plasmids that can be used in diverse molecular biological studies of the fungal pathogen Cryptococcus neoformans. We constructed 12 plasmids that can be used to tag a protein of interest with a GFP, mCherry, 4×FLAG, or 6×HA, along with nourseothricin-, neomycin-, or hygromycin-resistant selection markers. Using this tagging plasmid set, we explored the adenylyl cyclase complex (ACC), consisting of adenylyl cyclase (Cac1) and its associated protein Aca1, in the cAMP-signaling pathway, which is critical for the pathogenicity of C. neoformans. We found that Cac1-mCherry and Aca1-GFP were mainly colocalized as punctate forms in the cell membrane and nonnuclear cellular organelles. We also demonstrated that Cac1 and Aca1 interacted in vivo by coimmunoprecipitation, using Cac1-6×HA and Aca1-4×FLAG tagging strains. Bimolecular fluorescence complementation further confirmed the in vivo interaction of Cac1 and Aca1 in live cells. Finally, protein pull-down experiments using aca1Δ::ACA1-GFP and aca1Δ::ACA1- GFP cac1Δ strains and comparative mass spectrometry analysis identified Cac1 and a number of other novel ACC-interacting proteins. Thus, this versatile tagging plasmid system will facilitate diverse mechanistic studies in C. neoformans and further our understanding of its biology.
Ha-Won Jo,Yee-Rin Jung,Hyung-Min Park,Byoung-Sun Chu 대한시과학회 2020 대한시과학회지 Vol.22 No.4
목적 : 본 연구에서는 다양한 방법들을 이용하여 토릭렌즈 마킹 회전량을 평가할때 일치도를 평가측정하고자 하였다. 방법 : 모형안과 콘택트렌즈 착용이 가능한 57안(평균 24.79±1.29세)을 대상으로 토릭소프트렌즈를 제조사의 가이드라인에 따라 피팅한 후, 사진 촬영, 각도기 어플리케이션, 휴대용 돋보기와 세극등 현미경을 사용하여 회전 평가를 진행했을 때의 서로 다른 검사자 간의 검사 결과값을 비교 분석하였고, 서로 다른 회전평가 방법 간의 일 치도는 블랜드 알트만 분석을 사용하였다. 결과 : 서로 다른 검사자 간 같은 회전평가 방법으로 토릭소프트렌즈의 회전량을 측정하였을 때, 측정값의 평 균 차이가 사진 촬영, 각도기 어플리케이션, 휴대용 돋보기, 세극등 현미경 평가 방법 각각 –0.50, -0.70, -0.30, -0.13°로 유사한 결과를 보였다. 회전평가 방법 간 비교에 대해서는 사진 촬영과 각도기 어플리케이션을 사용했을 때, 허용 오차 범위(5°) 내의 측정값이 각각 74.56%, 75.44%로 비교적 낮은 신뢰도를 보였고 휴대용 돋 보기를 사용한 방법에서는 95.61%로 높은 신뢰도를 보였다. 결론 : 토릭소프트렌즈 처방에서 세극등 현미경을 사용할 수 없어도 사진 촬영, 핸드폰 각도기 어플리케이션, 휴대용 돋보기를 통해 토릭소프트렌즈 회전평가에 대한 임상적으로 유용한 결과를 얻을 수 있다. Purpose : The purpose of this study was to evaluate the agreement of different methods on assessing toric lens rotation. Methods : After fitting toric soft lens according to the manufacturer's guidelines at model eye and 57 eyes (average 24.79±1.29 years old), rotational evaluation was performed using a mobile phone photography, protractor application, portable magnifier and slit-lamp. Agreements among different methods and two testers were analyzed using Bland-Altman analysed. Results : When the rotation amount of the toric soft lens was measured by the same rotation evaluation method with different testers, the average difference of the measured values was –0.50, -0.70, -0.30 and –0.13° showing similar results. Regarding the comparison of orientationevaluate methods, when using the mobile phone photography and protractor App, the measured values within the tolerance range were relatively low with 74.56% and 75.44%, respectively, and with the portable magnifier, the reliability was high with 95.61%. Conclusion : Even if a slit-lamp can not be used in the prescription of a toric soft lens, clinically useful results for toric soft lens rotation evaluation can be obtained through photography, mobile phone protractor application and portable magnifier.
Interaction of microglia and amyloid-&bgr; through &bgr;2-integrin is regulated by RhoA
Jeon, Yee-Jin,Won, Ha-Young,Moon, Mi-Young,Choi, Won-Ho,Chang, Chun-Ho,Lee, Jae-Yong,Kim, Jaebong,Kim, Sung-Chan,Kim, Yong-Sun,Park, Jae-Bong Lippincott Williams Wilkins, Inc. 2008 NEUROREPORT - Vol.19 No.17
Amyloid-&bgr; (A&bgr;) is one of the main factors to cause Alzheimer’s disease. Although fibrillar A&bgr; (fA&bgr;) activates microglial cells that release toxic compounds to induce partial neuronal death, the mechanism of interaction between A&bgr; and microglia remains unclear. Therefore, we examined the interaction of microglial cells (BV2) and fA&bgr; on a gelatin-precoated plate. The binding was markedly enhanced by RhoA inactivation using Tat-C3, dominant negative RhoA, and si-RhoA. To identify the receptor for fA&bgr;, we tested various antibodies to mask receptors. Among them, anti-&bgr;2-integrin antibody mostly suppressed cell binding to fA&bgr;. The incremental binding of cells induced by RhoA inhibition was also blocked by addition of anti-&bgr;2-integrin antibody. These results suggest that RhoA inhibition stimulates &bgr;2-integrin-mediated cell interaction to fA&bgr;.
Choi, Jae-Sun,Ryu, Jaewook,Bae, Woom-Yee,Park, Aron,Nam, Seungyoon,Kim, Ja-Eun,Jeong, Joo-Won MDPI 2018 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.19 No.9
<P>Cancer cells undergo uncontrolled proliferation resulting from aberrant activity of various cell-cycle proteins. Therefore, despite recent advances in intensive chemotherapy, it is difficult to cure cancer completely. Recently, cell-cycle regulators became attractive targets in cancer therapy. Zingerone, a phenolic compound isolated from ginger, is a nontoxic and inexpensive compound with varied pharmacological activities. In this study, the therapeutic effect of zingerone as an anti-mitotic agent in human neuroblastoma cells was investigated. Following treatment of BE(2)-M17 cells with zingerone, we performed a 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay and colony-formation assay to evaluate cellular proliferation, in addition to immunofluorescence cytochemistry and flow cytometry to examine the mitotic cells. The association of gene expression with tumor stage and survival was analyzed. Furthermore, to examine the anti-cancer effect of zingerone, we applied a BALB/c mouse-tumor model using a BALB/c-derived adenocarcinoma cell line. In human neuroblastoma cells, zingerone inhibited cellular viability and survival. Moreover, the number of mitotic cells, particularly those in prometaphase, increased in zingerone-treated neuroblastoma cells. Regarding specific molecular mechanisms, zingerone decreased cyclin D1 expression and induced the cleavage of caspase-3 and poly (ADP-ribose) polymerase 1 (PARP-1). The decrease in cyclin D1 and increase in histone H3 phosphorylated (p)-Ser10 were confirmed by immunohistochemistry in tumor tissues administered with zingerone. These results suggest that zingerone induces mitotic arrest followed by inhibition of growth of neuroblastoma cells. Collectively, zingerone may be a potential therapeutic drug for human cancers, including neuroblastoma.</P>