著者は、彼女の貢献のためにクリスティPahrmannに感謝します。縫合材料を提供するためのエチコン、ノルダーシュテット、ハンブルク（ドイツ）に感謝します。 資金調達ソーニャSchrepferは、ドイツ学術振興（DFG）（SCHR992 / 3 1とSCHR992/4-1）から研究助成を受けています。]仕事がISHLTシャムウェイキャリア開発グラント2010ファルク研究資金（スタンフォード大学）によってサポートされていました。

1。内胸動脈（IMA）の準備<ol><li>動脈内皮は2フランスのフォガティ動脈塞栓除去カテーテル（バクスターヘルスケア、ディアフィールド、イリノイ州、米国）の通過によって荒廃しています。カテーテルが血管内皮の損傷を確実にするために二回全体の血管の長さを介してプルアップされています。 </li><li> 8mmの長さと直径が2.5ミリメートル、3ミリメートル（例えばTranslumina、ユーコンステント）のいずれかの人間のステントを使用しています。注意：ステントの直径は、事前と事後ステント狭窄を回避するために、10％以上の血管径を超えないようにしてください。注意：同一の研究内ステントの長さを切り替えないでください。 </li><li>所望の直径を達成するために適切なバルーン圧（ステントパッケージに記載されている）を使用してステントを展開します。 </li><li>ストアでは、移植まで氷上°C RPMI +ヘパリン（500 IE/10 ml）を4でIMAをステント。 </li></ol> 2。動物の準備 RNUヌード（CRL：NIH-Foxn1rnu）ラット（300〜350 g）はHOUです。scantainer、従来の条件の下でsedはキャビネット、飼料標準ラット固形飼料およびオートクレーブ処理水の広告libidumを換気。 <ol><li>誘導チャンバーを用いて（2.5から3パーセント）イソフルランでラットを麻酔。 </li><li>腹部の毛を剃ると、その後ろにラットを置いて、その鼻と麻酔を維持するために口にフェイスマスクを配置します。 </li><li>広くプロボ、ヨウ素、次回の使用の80％エタノールを用いて腹部を消毒する - が二回、この手順を繰り返します。ラットが麻酔で十分であることを確認するために、後肢の足をつまんで反射を確認してください。 </li><li>微視的なビューでは、大動脈腹部大動脈を露出させる上部の中央値は開腹術を実行します。 </li><li>生理食塩水保湿手袋で腸を配置します。水分の損失を防ぐために、腸の周りにグローブを折る。 </li><li>大動脈領域から分岐する大動脈を解剖、慎重に血管の枝に損傷を引き起こすことがありません。 </li><li>大動脈の血流を停止するには、マイクロクランプを使用しています。配置近位クランプは、まず遠位クランプ続く。 </li><li>アプリを削除します。 0.5〜0.7ミリメートルの大動脈セグメントとヘパリン（200単位）で、残りの大動脈をフラッシュします。 </li><li>ステントIMAを取り、十分な長さに短縮し、ギャップに配置します。 </li><li> 8から0プロレン縫合糸（エチコン、ノルダーシュテット、ドイツ）を用いて縫合を実行することによって、受信者の大動脈へのIMA接続します。 </li><li>慎重に第一脳神経クランプして、尾クランプを開きます。 </li><li>移植されたIMAおよび大動脈の遠位端で目に見えるパルスがあるはずです。 </li><li>腹部に戻し腸を配置します。 </li><li>予め温めておいた滅菌生理食塩水で腹部をフラッシュします。 </li><li> 6から0プロレン実行している縫合糸（エチコン、ノルダーシュテット、ドイツ）を使用して、腹壁の筋層を閉じます。 </li><li>ラットが麻酔に入れたまま、皮下に4から5 mg / kgのカルプロフェンを注入します。 </li><li>ポストsurg 3日間必要に応じて、外科手術後の鎮痛（例えば、カルプロフェンまたはメロキシカム）の管理ery。 </li></ol> 3。代表的な結果 1％乾燥した過酸化ベンゾイルとMMA I、1日のフタル酸ジブチル組織のために、検体はアルコールの段階的なシリーズで脱水し、4％ホルマリンで固定した、と80％のメチルメタクリレートの混合物（MMA I）に浸潤し、20％ 4で1〜2日で3％乾燥過酸化ベンゾイル（MMA III）で1日、MMA I℃のその後、標本は、プレ重合ベースで水浴中でガラスバイアルに新鮮なMMA IIIで重合された。ゆっくりと重合が26でバイアルを維持することによって達成された℃で一晩、重合が発生するまで°C翌朝、その後徐々に0.5°C 12時間かけて温度を増加させる28の温度を上昇させる。重合ブロックは、炭化タングステンのナイフを装備マイクロモルHM 360ミクロトームを使用して、5μmの厚さで切片。 増殖細胞の起源（ 図1）、sを識別するlidesのいずれか緑色蛍光タンパク質（GFP）またはラットMHC-Iとヒト平滑筋細胞を識別する抗体で染色した。これらの研究では、人間のIMAは、一晩IMAの細胞の安定な形質導入のためにレンチウイルス粒子を用いたレポーター遺伝子GFPとインキュベートした。人間の原点から分割する娘細胞は、GFPを発現することによって同定することができた。脱パラフィンした後、熱誘導エピトープ検索は蒸し器を使って抗原賦活液中でスライドを加熱することによって実行されます。画像-IT FXシグナルエンハンサーブロッキングステップに使用することができます。ヒト由来の細胞は、GFP（一次抗体希釈液（Dako社製）​​で希釈した1:100）に対するマウスモノクローナル抗体を用いて同定し、さらにヤギ抗マウスIgG、のAlexa Fluor®488（1:1000二次抗体希釈液で希釈しが付いています。）平滑筋細胞は、ヤギ抗ウサギIgG、Alexaのフルオ続いて、ウサギpolycolonal抗平滑筋α-アクチン（一次抗体希釈液で希釈した1:100）でマークされていたR 555（二次抗体希釈液で希釈した1:1000）。各抗体のインキュベーションステップは三回PBSの間での洗浄で1時間37℃で行われます。核は10分間DAPIで染色されています。 Gold褪色防止試薬を延長使用してスライドをマウントした後、試料を共焦点顕微鏡を用いて分析した。 <img alt="図1" src="/files/ftp_upload/3663/3663fig1.jpg" /> 図1。ヒト平滑筋細胞のような新生内膜細胞：緑=抗GFP、ヒト由来の細胞を標識、赤=抗ヒト平滑筋細胞のアクチン、青= DAPI、細胞核を識別B：グリーン=抗ラットMHC-I、ラベリングラット由来の細胞、赤=抗ヒト平滑筋細胞のアクチン、青= DAPI、細胞核を識別します。増殖細胞は平滑筋細胞として同定され、GFP陽性であるが、ラットのMHC-I分子の負されています。したがって、増殖細胞はヒト由来である。

<ol>
	<li>Deuse, T., Ikeno, F., Robbins, R. C., Schrepfer, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+In-Stent+Restenosis:+An+Inexpensive,+Reliable,+and+Rapid+Preclinical+Model.">Imaging In-Stent Restenosis: An Inexpensive, Reliable, and Rapid Preclinical Model.</a> J. Vis. Exp. (31), e1346 (2009).</li><li>Oyamada, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trans-iliac+rat+aorta+stenting:+a+novel+high+throughput+preclinical+stent+model+for+restenosis+and+thrombosis.">Trans-iliac rat aorta stenting: a novel high throughput preclinical stent model for restenosis and thrombosis.</a> J. Surg. Res. 166, e91-e95 (2011).</li><li>Chamberlain, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+mouse+model+of+in+situ+stenting.">A novel mouse model of in situ stenting.</a> Cardiovascular research. 85, 38-44 (2010).</li><li>Deuse, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Introducing+the+first+polymer-free+leflunomide+eluting+stent.">Introducing the first polymer-free leflunomide eluting stent.</a> Atherosclerosis. 200, 126-134 (2008).</li><li>Finn, A. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+healing+after+sirolimus,+paclitaxel,+and+bare+metal+stent+placement+in+combination+with+peroxisome+proliferator-activator+receptor+gamma+agonists:+requirement+for+mTOR/Akt2+in+PPARgamma+activation.">Differential healing after sirolimus, paclitaxel, and bare metal stent placement in combination with peroxisome proliferator-activator receptor gamma agonists: requirement for mTOR/Akt2 in PPARgamma activation.</a> Circulation research. 105, 1003-1012 (2009).</li><li>Tellez, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coronary+bare+metal+stent+implantation+in+homozygous+LDL+receptor+deficient+swine+induces+a+neointimal+formation+pattern+similar+to+humans.">Coronary bare metal stent implantation in homozygous LDL receptor deficient swine induces a neointimal formation pattern similar to humans.</a> Atherosclerosis. 213, 518-524 (2010).</li><li>Suzuki, Y., Yeung, A. C., Ikeno, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+pre-clinical+animal+model+in+the+translational+research+of+interventional+cardiology.">The pre-clinical animal model in the translational research of interventional cardiology.</a> JACC Cardiovasc. Interv. 2, 373-383 (2009).</li><li>Holt, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intimal+proliferation+in+an+organ+culture+of+human+internal+mammary+artery.">Intimal proliferation in an organ culture of human internal mammary artery.</a> Cardiovascular research. 26, 1189-1194 (1992).</li><li>Swanson, N., Javed, Q., Hogrefe, K., Gershlick, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+internal+mammary+artery+organ+culture+model+of+coronary+stenting:+a+novel+investigation+of+smooth+muscle+cell+response+to+drug-eluting+stents.">Human internal mammary artery organ culture model of coronary stenting: a novel investigation of smooth muscle cell response to drug-eluting stents.</a> Clin. Sci. (Lond). 103, 347-353 (2002).</li></ol>

利害の衝突が宣言されません。

in vivoでの研究のモデルで異なるが、ステント留置後の内膜肥厚の開発を調査するために既存していますが、これらのモデルは、まだ克服するために、トランスレーショナルハードルに直面している。さらに、大型の動物モデルは、高価で特殊な住宅事情であるだけでなく、手術の機器はすべての研究所では使用できません。 人間の内膜増殖およびステント内再狭窄の開発を研究するために人間のIMAを使用することをex situで器官培養8,9の前に検討した。新しいヒトIMAモデルでは、免疫不全ラットの腹部大動脈の位置に人間のIMAを注入することにより、in vivoで再狭窄ステントでの問題に対処するためのトランスレーショナルアプローチを構成します。人間のIMAのラットの腹部大動脈の大きさに完璧にマッチしています。したがって、我々に提示されたモデルは、実行するために、再現可能な簡単かつ高速ですが、特別な手術の訓練を必要とせず、です。安価な。 ヒト平滑筋細胞のような新生内膜細胞の同定は、臨床設定に翻訳ギャップを閉じて、in vivoでヒトの病態生理学的プロセスを調査する機会を示しています。

<table border="1"><tbody><tr><td> 試薬の名前 </td><td> 会社 </td><td> カタログ番号 </td><td> コメント </td></tr><tr><td> 2フランスのフォガティ·カテーテル</td><td>バクスターヘルスケア、ディアフィールド、イリノイ州、アメリカ合衆国</td><td> 120602F </td><td></td></tr><tr><td>ユーコンステント</td><td> Translumina GmbHは、ヘッヒンゲン、ドイツ</td><td></td><td>あなたの研究プロトコルに従って、お好みのステントを使用します。 </td></tr><tr><td> RPMI培地</td><td> Biochrom </td><td> Nr.F1275 </td><td></td></tr><tr><td>ヘパリン</td><td>バクスター</td><td> 2B0953 </td><td></td></tr><tr><td>イソフルレン</td><td>アボット</td><td> B506 </td><td></td></tr><tr><td>プロボ - ヨウ素</td><td> Betadine Puredueファーマ</td><td> EAN：5995327165830 </td><td></td></tr><tr><td> 80パーセントエタノリットル</td><td>ガイヤー</td><td> ETV 80/0500 </td><td></td></tr><tr><td>マイクロクランプ</td><td>ハーバード装置</td><td> PY2-61から0186 </td><td></td></tr><tr><td>縫合8から0 </td><td>ジョンソン·エンド·ジョンソン、 </td><td> 2808G </td><td></td></tr><tr><td>縫合6から0 </td><td>ジョンソン·エンド·ジョンソン、 </td><td> 1698年H </td><td></td></tr><tr><td>カルプロフェン</td><td> Feizerの獣医</td><td> PZN：0110208 </td><td></td></tr><tr><td> Metamizol </td><td> Ratiopharm </td><td></td><td></td></tr><tr><td>ターゲット検索ソリューションは、pH9の</td><td> Dako社</td><td> S2368 </td><td></td></tr><tr><td>画像-IT FXシグナルエンハンサー</td><td>インビトロジェン</td><td> I36933 </td><td></td></tr><tr><td>マウスモノクローナル抗GFP抗体</td><td> BDの生活の色</td><td> 632381 </td><td></td></tr><tr><td>一次抗体希釈液</td><td> Dako社</td><td> S3022 </td><td></td></tr><tr><td>ヤギ抗マウスIgG、のAlexa Fluor®488 </td><td>インビトロジェン</td><td> A11017 </td><td></td></tr><tr><td>二次抗体希釈液</td><td> Dako社</td><td> S0809 </td><td></td></tr><tr><td>ウサギpolycolonal抗平滑筋α-アクチン</td><td>アブカム、 </td><td> ab5694 </td><td></td></tr><tr><td>ヤギ抗ウサギIgG、のAlexa Fluor®555 </td><td>インビトロジェン</td><td> A21430 </td><td></td></tr><tr><td> Gold褪色防止試薬を延長</td><td>インビトロジェン</td><td> P36930 </td><td></td></tr></tbody></table>

human internal mammary artery (ima) transplantation and stenting: a human model to study the development of in-stent restenosis

血管ステント留置後の内膜肥厚の開発にpathobiologicalと病態生理学的プロセスを調べることがin vivoでの研究モデルの前臨床試験では、トランスレーショナルアプローチ1,2のために重要である。 一般的に使用される動物モデルはマウス、ラット、ウサギ、ブタ3-5含まれています。それらの生物学的プロセスが既に動物の血管で研究が、ヒトの研究モデル6,7の前に実行されることはありませんしかし、臨床現場へのこれらのモデルの変換は、困難である。このビデオでは、この翻訳のギャップを克服するための新たなヒト化モデルを示しています。に示す手順は、実行するために、再現性が容易で、高速であり、内膜肥厚と多様なステントの適用の開発を勉強するのに適しています。 このビデオでは、免疫不全ラットへの移植に続いて、人間の血管内ステント技術を実行する方法を示し、人間として増殖する細胞の起源を識別します。

Watch this Scientific Journal Video about Human Internal Mammary Artery IMA Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis at JoVE.com

Human Internal Mammary Artery IMA Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis

このビデオでは、免疫不全ラットモデルにおいてヒト血管（IMA）を使用して、ステント展開後に内膜肥厚の開発を研究するためのモデルを示しています。

人間の内胸動脈（IMA）移植およびステント留置術：ステント内再狭窄の開発を研究するための人体モデル

Preclinical in vivo research models to investigate pathobiological and pathophysiological processes in the development of intimal hyperplasia after vessel ...

human-internal-mammary-artery-ima-transplantation-stenting-human

Nanyang Technological University

Research

JoVE Journal

Medicine

Human Internal Mammary Artery (IMA) Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis

11.2K ビュー.  TSI-Lab, Germany. このビデオでは、免疫不全ラットモデルにおいてヒト血管（IMA）を使用して、ステント展開後に内膜肥厚の開発を研究するためのモデルを示しています。

ビデオ: Human Internal Mammary Artery IMA Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis

Orthotopic Aortic Transplantation: A Rat Model to Study the Development of Chronic Vasculopathy

Research models of chronic rejection are essential to investigate pathobiological and pathophysiological processes during the development of transplant vasculopathy (TVP).
The commonly used animal model for cardiovascular chronic rejection studies is the heterotopic heart transplant model performed in laboratory rodents. This model is used widely in experiments since Ono and Lindsey (3) published their technique. To analyze the findings in the blood vessels, the heart has to be sectioned and all vessels have to be measured.
Another method to investigate chronic rejection in cardiovascular questionings is the aortic transplant model (1, 2). In the orthotopic aortic transplant model, the aorta can easily be histologically evaluated (2). The PVG-to-ACI model is especially useful for CAV studies, since acute vascular rejection is not a major confounding factor and Cyclosporin A (CsA) treatment does not prevent the development of CAV, similar to what we find in the clinical setting (4). A7-day period of CsA is required in this model to prevent acute rejection and to achieve long-term survival with the development of TVP.
This model can also be used to investigate acute cellular rejection and media necrosis in xenogeneic models (5).

This video demonstrates the orthotopic aortic transplant model as a simple model to study the development of transplant vasculopathy (TVP) in rats.

同所性大動脈移植：慢性血管障害の発生研究のラットモデル

Research models of chronic rejection are essential to investigate pathobiological and pathophysiological processes during the development of transplant ...

医学

Utilizing Transcranial Magnetic Stimulation to Study the Human Neuromuscular System

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade. While the use of TMS has expanded to the study of many systems and processes during this time, the original application and perhaps one of the most common uses of TMS involves studying the physiology, plasticity and function of the human neuromuscular system. Single pulse TMS applied to the motor cortex excites pyramidal neurons transsynaptically 2 (Figure 1) and results in a measurable electromyographic response that can be used to study and evaluate the integrity and excitability of the corticospinal tract in humans 3. Additionally, recent advances in magnetic stimulation now allows for partitioning of cortical versus spinal excitability 4,5. For example, paired-pulse TMS can be used to assess intracortical facilitatory and inhibitory properties by combining a conditioning stimulus and a test stimulus at different interstimulus intervals 3,4,6-8. In this video article we will demonstrate the methodological and technical aspects of these techniques. Specifically, we will demonstrate single-pulse and paired-pulse TMS techniques as applied to the flexor carpi radialis (FCR) muscle as well as the erector spinae (ES) musculature. Our laboratory studies the FCR muscle as it is of interest to our research on the effects of wrist-hand cast immobilization on reduced muscle performance6,9, and we study the ES muscles due to these muscles clinical relevance as it relates to low back pain8. With this stated, we should note that TMS has been used to study many muscles of the hand, arm and legs, and should iterate that our demonstrations in the FCR and ES muscle groups are only selected examples of TMS being used to study the human neuromuscular system.

Transcranial magnetic stimulation (TMS) is a non-invasive tool to gain insight on the physiology and function of the human nervous system. Here, we present our TMS techniques to study cortical excitability of the upper limb and lumbar musculature.

人間の神経筋システムを研究するために経頭蓋磁気刺激の活用

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade.  While ...

Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model

Stem cell transplantation protocols are finding their way into clinical practice1,2,3. Getting better results, making the protocols more robust, and finding new sources for implantable cells are the focus of recent research4,5. Investigating the effectiveness of cell therapies is not an easy task and new tools are needed to investigate the mechanisms involved in the treatment process6. We designed an experimental protocol of ischemia/reperfusion in order to allow the observation of cellular connections and even subcellular mechanisms during ischemia/reperfusion injury and after stem cell transplantation and to evaluate the efficacy of cell therapy. H9c2 cardiomyoblast cells were placed onto cell culture plates7,8. Ischemia was simulated with 150 minutes in a glucose free medium with oxygen level below 0.5%. Then, normal media and oxygen levels were reintroduced to simulate reperfusion. After oxygen glucose deprivation, the damaged cells were treated with transplantation of labeled human bone marrow derived mesenchymal stem cells by adding them to the culture. Mesenchymal stem cells are preferred in clinical trials because they are easily accessible with minimal invasive surgery, easily expandable and autologous. After 24 hours of co-cultivation, cells were stained with calcein and ethidium-homodimer to differentiate between live and dead cells. This setup allowed us to investigate the intercellular connections using confocal fluorescent microscopy and to quantify the survival rate of postischemic cells by flow cytometry. Confocal microscopy showed the interactions of the two cell populations such as cell fusion and formation of intercellular nanotubes. Flow cytometry analysis revealed 3 clusters of damaged cells which can be plotted on a graph and analyzed statistically. These populations can be investigated separately and conclusions can be drawn on these data on the effectiveness of the simulated therapeutical approach.

Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model

We demonstrate how to set up an in vitro ischemia/reperfusion model and how to evaluate the effect of stem cell therapy on postischemic cardiac cells.

における幹細胞移植 in vitroで模擬虚血/再灌流モデル

Stem cell transplantation protocols are finding their way into clinical practice1,2,3. Getting better results, making the protocols more robust, and ...

Human Neuroendocrine Tumor Cell Lines as a Three-Dimensional Model for the Study of Human Neuroendocrine Tumor Therapy

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human malignancies.1 Other than surgery for the minority of patients who present with localized disease, there is little or no survival benefit of systemic therapy. Therefore, there is a great need to better understand the biology of NETs, and in particular define new therapeutic targets for patients with nonresectable or metastatic neuroendocrine tumors. 3D cell culture is becoming a popular method for drug screening due to its relevance in modeling the in vivo tumor tissue organization and microenvironment.2,3 The 3D multicellular spheroids could provide valuable information in a more timely and less expensive manner than directly proceeding from 2D cell culture experiments to animal (murine) models.
To facilitate the discovery of new therapeutics for NET patients, we have developed an in vitro 3D multicellular spheroids model using the human NET cell lines. The NET cells are plated in a non-adhesive agarose-coated 24-well plate and incubated under physiological conditions (5% CO2, 37 &deg;C) with a very slow agitation for 16-24 hr after plating. The cells form multicellular spheroids starting on the 3rd or 4th day. The spheroids become more spherical by the 6th day, at which point the drug treatments are initiated. The efficacy of the drug treatments on the NET spheroids is monitored based on the morphology, shape and size of the spheroids with a phase-contrast light microscope. The size of the spheroids is estimated automatically using a custom-developed MATLAB program based on an active contour algorithm. Further, we demonstrate a simple method to process the HistoGel embedding on these 3D spheroids, allowing the use of standard histological and immunohistochemical techniques. 
This is the first report on generating 3D spheroids using NET cell lines to examine the effect of therapeutic drugs. We have also performed histology on these 3D spheroids, and displayed an example of a single drug's effect on growth and proliferation of the NET spheroids. Our results support that the NET spheroids are valuable for further studies of NET biology and drug development.

We present a simple agarose overlay platform to grow 3D multicellular spheroids using neuroendocrine cancer cell lines. This method provides a very convenient way to examine the effect of therapeutic drugs on the neuroendocrine tumor cells. It could also help us establish human neuroendocrine tumor spheroids for cancer therapy.

人間の神経内分泌腫瘍療法の研究のための三次元モデルとしての人間の神経内分泌腫瘍細胞株

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human ...

A Murine Model of Stent Implantation in the Carotid Artery for the Study of Restenosis

Despite the considerable progress made in the stent development in the last decades, cardiovascular diseases remain the main cause of death in western countries. Beside the benefits offered by the development of different drug-eluting stents, the coronary revascularization bears also the life-threatening risks of in-stent thrombosis and restenosis. Research on new therapeutic strategies is impaired by the lack of appropriate methods to study stent implantation and restenosis processes. Here, we describe a rapid and accessible procedure of stent implantation in mouse carotid artery, which offers the possibility to study in a convenient way the molecular mechanisms of vessel remodeling and the effects of different drug coatings.

A model of stent implantation in mouse carotid artery is described. Compared to other similar methods, this procedure is very rapid, simple and accessible, offering the possibility to study in a convenient way the vascular wall reaction to different drug-eluting stents and the molecular mechanisms of restenosis.

再狭窄の研究のための頸動脈内ステント留置のマウスモデル

Despite  the considerable progress made in the stent development in the last decades,  cardiovascular diseases remain the main cause of death in western ...

A Human Ex Vivo Atherosclerotic Plaque Model to Study Lesion Biology

Atherosclerosis is a chronic inflammatory disease of the vasculature. There are various methods to study the inflammatory compound in atherosclerotic lesions. Mouse models are an important tool to investigate inflammatory processes in atherogenesis, but these models suffer from the phenotypic and functional differences between the murine and human immune system. In vitro cell experiments are used to specifically evaluate cell type-dependent changes caused by a substance of interest, but culture-dependent variations and the inability to analyze the influence of specific molecules in the context of the inflammatory compound in atherosclerotic lesions limit the impact of the results. In addition, measuring levels of a molecule of interest in human blood helps to further investigate its clinical relevance, but this represents systemic and not local inflammation. Therefore, we here describe a plaque culture model to study human atherosclerotic lesion biology ex vivo. In short, fresh plaques are obtained from patients undergoing endarterectomy or coronary artery bypass grafting and stored in RPMI medium on ice until usage. The specimens are cut into small pieces followed by random distribution into a 48-well plate, containing RPMI medium in addition to a substance of interest such as cytokines or chemokines alone or in combination for defined periods of time. After incubation, the plaque pieces can be shock frozen for mRNA isolation, embedded in Paraffin or OCT for immunohistochemistry staining or smashed and lysed for western blotting. Furthermore, cells may be isolated from the plaque for flow cytometry analysis. In addition, supernatants can be collected for protein measurement by ELISA. In conclusion, the presented ex vivo model opens the possibility to further study inflammatory lesional biology, which may result in identification of novel disease mechanisms and therapeutic targets.

A Human Ex Vivo Atherosclerotic Plaque Model to Study Lesion Biology

Atherosclerosis is a chronic inflammatory process. This manuscript illustrates an easy to use ex vivo model to investigate fresh carotid or coronary artery plaques. The ex vivo model allows for the investigation of potential substances on the inflammatory milieu in human atherosclerotic lesions and results can be analyzed by various methods.

ヒューマン<em&gt;生体外</em&gt;動脈硬化性プラークのモデルは、病変生物学を研究する

Atherosclerosis is a chronic inflammatory disease of the vasculature. There are various methods to study the inflammatory compound in atherosclerotic ...

Excitotoxic Stimulation of Brain Microslices as an In vitro Model of Stroke

Examining molecular mechanisms involved in neuropathological conditions, such as ischemic stroke, can be difficult when using whole animal systems. As such, primary or &#39;neuronal-like&#39; cell culture systems are commonly utilized. While&#160;these systems are relatively easy to work with, and are useful model systems in which various functional outcomes (such as cell death) can be readily quantified, the examined outcomes and pathways in cultured immature neurons (such as excitotoxicity-mediated cell death pathways) are not necessarily the same as those observed in mature brain, or in intact tissue. Therefore, there is the need to develop models in which cellular mechanisms in mature neural tissue can be examined. We have developed an in vitro technique that can be used to investigate a variety of molecular pathways in intact nervous tissue. The technique described herein utilizes rat cortical tissue, but this technique can be adapted to use tissue from a variety of species (such as mouse, rabbit, guinea pig, and chicken) or brain regions (for example, hippocampus, striatum, etc.). Additionally, a variety of stimulations/treatments can be used (for example, excitotoxic, administration of inhibitors, etc.). In conclusion, the brain slice model described herein can be used to examine a variety of molecular mechanisms involved in excitotoxicity-mediated brain injury.

Excitotoxic Stimulation of Brain Microslices as an In vitro Model of Stroke

We have developed a brain slice model which can be used to examine molecular mechanisms involved in excitotoxicity-mediated brain injury.&#160;This technique generates viable mature brain tissue and reduces animal numbers required for experimentation, whilst keeping the neuronal circuitry, cellular interactions, and postsynaptic compartments partly intact.

脳Microslicesの興奮毒性刺激<em&gt;インビトロ</em&gt;脳卒中のモデル

Examining molecular mechanisms involved in neuropathological conditions, such as ischemic stroke, can be difficult when using whole animal systems. As ...

Murine Corneal Transplantation: A Model to Study the Most Common Form of Solid Organ Transplantation

Corneal transplantation is the most common form of organ transplantation in the United States with between 45,000 and 55,000 procedures performed each year. While several animal models exist for this procedure and mice are the species that is most commonly used. The reasons for using mice are the relative cost of using this species, the existence of many genetically defined strains that allow for the study of immune responses, and the existence of an extensive array of reagents that can be used to further define responses in this species. This model has been used to define factors in the cornea that are responsible for the relative immune privilege status of this tissue that enables corneal allografts to survive acute rejection in the absence of immunosuppressive therapy. It has also been used to define those factors that are most important in rejection of such allografts. Consequently, much of what we know concerning mechanisms of both corneal allograft acceptance and rejection are due to studies using a murine model of corneal transplantation. In addition to describing a model for acute corneal allograft rejection, we also present for the first time a model of late-term corneal allograft rejection.

Mice have been used as a model for studying many forms of transplantation, including corneal transplantation. We describe in this report a murine model for both acute and late-term corneal transplantation.

ネズミ角膜移植：固体臓器移植の最も一般的な形態を研究するためのモデル

Corneal transplantation is the most common form of organ transplantation in the United States with between 45,000 and 55,000 procedures performed each ...

Establishment of a Human Multiple Myeloma Xenograft Model in the Chicken to Study Tumor Growth, Invasion and Angiogenesis

Multiple myeloma (MM), a malignant plasma cell disease, remains incurable and novel drugs are required to improve the prognosis of patients. Due to the lack of the bone microenvironment and auto/paracrine growth factors human MM cells are difficult to cultivate. Therefore, there is an urgent need to establish proper in vitro and in vivo culture systems to study the action of novel therapeutics on human MM cells. Here we present a model to grow human multiple myeloma cells in a complex 3D environment in vitro and in vivo. MM cell lines OPM-2 and RPMI-8226 were transfected to express the transgene GFP and were cultivated in the presence of human mesenchymal cells and collagen type-I matrix as three-dimensional spheroids. In addition, spheroids were grafted on the chorioallantoic membrane (CAM) of chicken embryos and tumor growth was monitored by stereo fluorescence microscopy. Both models allow the study of novel therapeutic drugs in a complex 3D environment and the quantification of the tumor cell mass after homogenization of grafts in a transgene-specific GFP-ELISA. Moreover, angiogenic responses of the host and invasion of tumor cells into the subjacent host tissue can be monitored daily by a stereo microscope and analyzed by immunohistochemical staining against human tumor cells (Ki-67, CD138, Vimentin) or host mural cells covering blood vessels (desmin/ASMA).
In conclusion, the onplant system allows studying MM cell growth and angiogenesis in a complex 3D environment and enables screening for novel therapeutic compounds targeting survival and proliferation of MM cells.

Human multiple myeloma (MM) cells require the supportive microenvironment of mesenchymal cells and extracellular matrix components for survival and proliferation. We established an in vivo chicken embryo model with engrafted human myeloma and mesenchymal cells to study effects of cancer drugs on tumor growth, invasion and angiogenesis.

腫瘍の増殖、浸潤及び血管新生を研究するために、ニワトリでのヒト多発性骨髄腫異種移植モデルの確立

Multiple myeloma (MM), a malignant plasma cell disease, remains incurable and novel drugs are required to improve the prognosis of patients. Due to the ...

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular weight metabolites in tissues of the treated host to achieve long-term survival. The semipermeable membrane allows engrafted encapsulated cells to avoid rejection by the immune system. The encapsulation procedure was designed to enable a controlled release of bioactive compounds, such as insulin, other hormones, and cytokines. Here we describe a method for encapsulation of catabolic cells, which consume lipids for heat production and energy dissipation (thermogenesis) in the intra-abdominal adipose tissue of obese mice. Encapsulation of thermogenic catabolic cells may be potentially applicable to the prevention and treatment of obesity and type 2 diabetes. Another potential application of catabolic cells may include detoxification from alcohols or other toxic metabolites and environmental pollutants.

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

移植のためのカプセル化サーモ前脂肪細胞の脂肪組織デポへ

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular ...

人間の内胸動脈（IMA）移植およびステント留置術：ステント内再狭窄の開発を研究するための人体モデル

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