私たちは、VEVO 2100超音波システムを提供するための香港理工大学の生命科学における大学研究施設（ULS）を感謝したいです。訓練及び円滑のための医工学、香港理工大学、学際課から博士Chunyiウェンに感謝します。この作品は、香港政府RGCグラントGRF 17127215、764812とビリー・KCチョウにHKU6 / CRF / 11Gによってサポートされていました。

プロトコルは、香港の大学の教育・研究（CULATR）でライブ動物の使用に関する委員会のガイドラインに従います。 1.運用サイトの準備<ol><li> 75％イソプロピルアルコールで手術部位を消毒します。心の変化や対象動物の呼吸速度を避けるために、37℃に加熱パッドの温度を調整します。 </li><li> 30分間、または他の適切な方法により、121℃で重力置換滅菌器で滅菌手術器具を使用してください。 </li><li>ライゲーションおよび組織分離および開創器として使用するためのツールを作るために、 図1に見られるように、4〜30ゲージの針と針ホルダとカーブして2 -を取ります。手術で使用するための綿棒になりまし-湾曲針をマウントします。 </li><li>それは針ホルダーと90°27ゲージの針と曲線を鈍ら。連結工程で0.4 mmのスペーサーを作るために、ハード面を擦ることにより、先端を滑らかに。 </li></ol><ol><li>負のテールピンチ反射で麻酔を確認してください100ミリグラム/キログラム+ 10mg / kgの腹腔：体重との関係で適切なケタミンおよびキシラジンの混合物と2カ月齢、オス、22〜25グラムC57BL / 6Nマウスを麻酔。 </li><li>脱毛クリームの2〜3分のアプリケーションと前頸部と胸の毛を削除してください。暖かい水と綿棒でクリームを削除します。 </li><li>角膜の乾燥を防ぐために、動物の眼に眼軟膏またはゲルを適用します。 </li><li>仰臥位で加熱パッド上に動物を配置し、すべての15分間隔で適切な文書を維持することを確認され、術中モニタリングを行っています。 </li><li>手術中に正しい位置に動物を維持するために加熱パッドに手足を固定するための外科紙テープを使用してください。 </li><li>アルコールとポビドンヨード溶液で交互に3回皮膚を消毒。 </li><li>無菌DRを使用汚染を防ぐために露出され、操作フィールドと猿。各マウスについての新しい滅菌手袋を使用し、必要に応じて。 </li></ol> 3.連結手順<ol><li>メスで首と胸の正中線の位置でマウスの皮膚を開きます。 </li><li>優しく鈍はさみで結合組織を分離することにより、頭部に向かって甲状腺を引き出します。 </li><li>湾曲した針との両側に向かって正中線で気管に筋層を分離します。 </li><li>鈍はさみを使用して、第2のリブ（約5mm）に胸骨を切りました。開創又は湾曲鉗子でカットを開きます。 </li><li>湾曲した針で結合組織を分離することによって、互いと下胸壁からの胸腺葉を分離します。横方向の大動脈弓と2頸動脈は、この時点ではっきりと見えるようになります。 </li><li>アーチの下に湾曲した針を配置し、膿によって反対側の血管壁および結合組織の間に穴を開ける湾曲したヘッドを興。 </li><li>湾曲した針を使用して、大動脈弓の下に6-0モノフィラメント縫合糸を引っ張ります。 </li><li>ループ内で、スペーサを配置し、ダブルノットで所定の位置に縫合糸を固定します。優しくスペーサーを取り外します。 </li><li>結び目の位置に成功したくびれを確認して、縫合糸の端をカット。 </li><li>単純結節縫合パターンで6/0シルク縫合糸を使用して、胸壁を閉じます。連続縫合パターンの6/0モノフィラメント縫合糸で皮膚を閉じます。 </li></ol> 4.術後ケア<ol><li>縫合部位にポビドンヨード溶液を塗布し、回復および術後のモニタリングに予熱したケージ内に動物を配置します。 </li><li>動物が痛みを緩和し、適切なソフトな食品とマウスを提供するために、意識を取り戻した後 - （0.1ミリグラム/ kg皮下12時間ごとに0.05）ブプレノルフィンを注入します。 </li><li>完全な回復後12時間の明/暗サイクルの部屋に動物を返します。 </li></ol> 5。心エコー検査：成功ライゲーションおよび測定の確認<ol><li> 1 L /分の酸素流量で、メンテナンスのために、1.5％ - 誘導のために、5％イソフルラン、および1 - 一週間後、3と以前バンドマウスを麻酔。 </li><li>脱毛クリームを使用して、胸の毛を削除します。徹底的に暖かい水と綿棒でクリームを削除します。 </li><li> 37℃に維持動物処理台に仰臥位でマウスを固定します。足に電極ゲルを適用し、電極とパッドにそれらをテープで固定します。 </li><li>マウスの胸部に超音波ゲルを適用します。 40 MHzのMS550Dプローブおよび高周波数超音波システムと、大動脈弓のと同様に、左右の頸動脈上にカラードプラとパルス波ドプラ走査を行います。 </li><li> Bモードを使用して、左心室を探します。 Mモードを使用して、収縮期および拡張期の壁厚を測定することができます。使用シネストアおよび必要に応じてデータや画像を保存するためのフレームストア。 </li><li> UB-モードを歌う、分岐点の前に左総頸動脈を見つけます。流れを評価するために、パルス波やカラードップラーを使用してください。類似した時点で、右頸動脈に繰り返します。後で分析するためにシネストアとフレームストアモードで画像を保存します。 </li><li>マウスは左側臥位になるように、左端に動物ハンドリングプラットフォームを傾けます。大動脈弓を見つけるために、XとYをマウス、胸骨の位置でヘッドに平行な位置からのオフセットを30°にプローブを配置し、操作します。 </li><li>くびれの位置を確認します。血液は、2つの頚動脈が大動脈弓から分岐点との間の狭い水路を流れるされるように、くびれのサイトをより見やすくするために、カラードプラモードを使用します。 </li><li>ピーク流速を測定するために、パルス波やカラードップラーを使用してください。使用シネストアおよび必要に応じてデータや画像を保存するためのフレームストア。 </li><li>動物が腹臥位に加熱パッド上で回復させます。それは完全に麻酔から回復した後に12時間の明/暗サイクルの部屋で、元のケージに動物を返します。 </li></ol>

<ol>
	<li>Rockman, H. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Segregation+of+atrial-specific+and+inducible+expression+of+an+atrial+natriuretic+factor+transgene+in+an+in+vivo+murine+model+of+cardiac+hypertrophy.">Segregation of atrial-specific and inducible expression of an atrial natriuretic factor transgene in an in vivo murine model of cardiac hypertrophy.</a> Proc Natl Acad Sci U S A. 88 (18), 8277-8281 (1991).</li><li>de Almeida, A. C., van Oort, R. J., Wehrens, X. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transverse+aortic+constriction+in+mice.">Transverse aortic constriction in mice.</a> J Vis Exp. (38), (2010).</li><li>Hu, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Minimally+invasive+aortic+banding+in+mice:+effects+of+altered+cardiomyocyte+insulin+signaling+during+pressure+overload.">Minimally invasive aortic banding in mice: effects of altered cardiomyocyte insulin signaling during pressure overload.</a> Am J Physiol Heart Circ Physiol. 285 (3), H1261-H1269 (2003).</li><li>Shimizu, I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Excessive+cardiac+insulin+signaling+exacerbates+systolic+dysfunction+induced+by+pressure+overload+in+rodents.">Excessive cardiac insulin signaling exacerbates systolic dysfunction induced by pressure overload in rodents.</a> J Clin Invest. 120 (5), 1506-1514 (2010).</li><li>Tarnavski, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+surgical+models+in+cardiovascular+research.">Mouse surgical models in cardiovascular research.</a> Methods Mol Biol. 573, 115-137 (2009).</li><li>Zhang, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+modified+murine+model+for+the+study+of+reverse+cardiac+remodelling.">A modified murine model for the study of reverse cardiac remodelling.</a> Exp Clin Cardiol. 18 (2), e115-e117 (2013).</li><li>Hartley, C. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doppler+estimation+of+reduced+coronary+flow+reserve+in+mice+with+pressure+overload+cardiac+hypertrophy.">Doppler estimation of reduced coronary flow reserve in mice with pressure overload cardiac hypertrophy.</a> Ultrasound Med Biol. 34 (6), 892-901 (2008).</li><li>Scherrer-Crosbie, M., Thibault, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Echocardiography+in+translational+research:+of+mice+and+men.">Echocardiography in translational research: of mice and men.</a> J Am Soc Echocardiogr. 21 (10), 1083-1092 (2008).</li><li>Faerber, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+of+heart+failure+by+minimally+invasive+aortic+constriction+in+mice:+reduced+peroxisome+proliferator-activated+receptor+gamma+coactivator+levels+and+mitochondrial+dysfunction.">Induction of heart failure by minimally invasive aortic constriction in mice: reduced peroxisome proliferator-activated receptor gamma coactivator levels and mitochondrial dysfunction.</a> J Thorac Cardiovasc Surg. 141 (2), 492-500 (2011).</li><li>Li, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+Cardiac+Morphological+and+Functional+Changes+in+Mouse+Model+of+Transverse+Aortic+Constriction+by+Echocardiographic+Imaging.">Assessment of Cardiac Morphological and Functional Changes in Mouse Model of Transverse Aortic Constriction by Echocardiographic Imaging.</a> J Vis Exp. (112), (2016).</li></ol>

The authors declare that they have no competing financial interests.

ロックマンらは、心臓に圧負荷を研究するための方法として、大動脈結紮するための第1の方法を考案しました。これは、高血圧や心不全1の細胞および分子の結果を明らかに開始するための研究を可能にしました。改良された方法、低侵襲横断大動脈狭窄（MTAC）は、従来の横断大動脈狭窄9の欠点がない、左心室圧負荷および心不全の誘導のために適し、時...

<html>この記事の全体的な目的は、マウスの大動脈狭窄（TAC）は、より単純で好都合手術を生産するように改変することができる方法を横断実証することです。 TACは、1991年1に導入され、操作ゲノム2、3、4、5、6を特に小動物において、左心室および心不全に肥大につながる細胞および分子経路を研究するために分子生物学を有効にしました。所望の圧力過負荷の製造におけるその有効性にもかかわらず、技術は、最終的手順を行う際に、その低い生存率により困難によって制限されます。 TACを行う外科医は、気管内挿管、開胸術を実行することができ、および肺虚脱1を防止するために人工呼吸を使用しなければなりません。 コンテンツは&quot;&gt;多くの場合、簡単かつ低侵襲性の方法が望まれている。その結果、低侵襲横断大動脈狭窄（MTAC）は3を考案した。結果（具体的には、左心室肥大）が同じであるが、手順が有意に少ない装置を含みます動物へとダメージが少ない。MTACが90％以上の大きな生存率を、持っている、とだけ15〜25分の動作時間を持つことが望ましい方法であることが判明している。成功した手術後の死亡のまれな例があまりにに起因することができます驚異的な左心室過負荷や、腎臓などの重要な臓器への血液供給の致命的な減少をもたらす大動脈弓の-tightくびれ、。 手順が完了した後、心エコー検査は、バンディングおよび心臓形態および機能の成功を評価するために使用することができます。高周波超音波システムは、直径を測定し、左右の変化を流すために利用することができます頚動脈、ピークフロー速度測定のための大動脈弓の狭窄を可視化するため、および左心室壁の厚さと機能パラメータ7,8を決定します。安楽死の後、器官はまた、ヘマトキシリン及びエオシン染色およびピクロシリウスコラーゲン染色で組織病理学的研究のために収集することができ、後者の線維を観察するために特に有用です。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1 inch 30 Gauge Needles</td>
			<td>BD, Franklin Lakes, NJ 07417, USA.&#160;</td>
			<td>305128</td>
			<td>Curved as in the procedure and Figure 1</td>
		</tr>
		<tr>
			<td>27 Gauge Needle</td>
			<td>BD, Franklin Lakes, NJ 07417, USA.&#160;</td>
			<td>301629</td>
			<td>Make blunt and smooth for spacer</td>
		</tr>
		<tr>
			<td>6/0 Dafilon</td>
			<td>B. Braun, 34212 Melsungen, Hessen, Germany.</td>
			<td>C0933066</td>
			<td></td>
		</tr>
		<tr>
			<td>6/0 Silkam</td>
			<td>B. Braun, 34212 Melsungen, Hessen, Germany.</td>
			<td>C0762067</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile gloves</td>
			<td>A.R. Medicom, Inc (Asia), Hong Kong.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton Applicator</td>
			<td>Mannings, Quarry Bay, Hong Kong.</td>
			<td></td>
			<td>Local Shopping Center</td>
		</tr>
		<tr>
			<td>Delapitory Cream</td>
			<td>Veet (Hong Kong), Kwun Tong, Kowloon, Hong Kong</td>
			<td></td>
			<td>Local Shopping Center</td>
		</tr>
		<tr>
			<td>Lexer-Baby Scissor</td>
			<td>FST, British Columbia V7H 0A6, Canada.</td>
			<td>14078-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Curved Iris Forcep</td>
			<td>FST, British Columbia V7H 0A6, Canada.</td>
			<td>11065-07</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro Olsen-Hegar Needle Holder</td>
			<td>WPI, Sarasota, FL 34240, USA.&#160;</td>
			<td>501989</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereo Micrioscope</td>
			<td>WPI, Sarasota, FL 34240, USA.&#160;</td>
			<td>PZMIII-BS</td>
			<td></td>
		</tr>
		<tr>
			<td>GenTeal Gel</td>
			<td>Novartis, East Hanover, NJ 07936, USA.&#160;</td>
			<td></td>
			<td>Local Pharmacy</td>
		</tr>
		<tr>
			<td>Heating Pad</td>
			<td>Kent Scientific Corporation, Torrington, CT 06790, USA.&#160;</td>
			<td>DCT-20</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical tape</td>
			<td>Laboratory Animal Unit</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamine and Xylazine</td>
			<td>Laboratory Animal Unit</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Betadine</td>
			<td>Laboratory Animal Unit</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Buprenorphine</td>
			<td>Laboratory Animal Unit</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

minimally invasive transverse aortic constriction in mice

低侵襲性の横大動脈狭窄（MTAC）は、標準開胸横大動脈狭窄（TAC）より、マウスにおける横大動脈の狭窄のためのより望ましい方法です。横方向の大動脈狭窄は、左心室における高圧の誘導のための機能性に優れた方法であるが、それは気管挿管と人工呼吸の使用のために、より困難で長い手順です。 TACは、新しい方法、MTACとして、しばしばも少ない存続可能ではありません、どちらも換気セットアップで肋骨と肋間筋の切断や気管挿管を必要とします。胸腔にアクセスするために開胸とは対照的にMTACでは、大動脈弓は、前首の正中切開を介して到達されます。甲状腺は、胸骨切痕を明らかにするために引き戻されます。胸骨はその後、第2リブレベルに削減され、大動脈弓は、結合組織と胸腺を分離することによって、単純に到達します。からそこに、縫合糸は、アーチの周りに包まれ、スペーサで結ばれ、その後、胸骨カットし、皮膚を閉じることができますすることができます。 MTACは、左心室の高血圧症を誘発するより速く、より侵襲性の低い方法であり、ハイスループット研究の可能性を可能にします。狭窄の成功は、大動脈弓と左右の頸動脈血管の寸法の流速を決定するために、高周波経胸腔心エコー検査法、特にカラードップラーとパルス波ドップラーを使用して確認することができ、かつ左心室機能と形態。成功したくびれはまた、間質および血管周囲線維症の心筋細胞肥大などの重要な組織病理学的変化を、トリガされます。ここでは、MTACの手順は、頸動脈中の得られた流れの変化は、心エコー検査、全体の形態、および心臓内の組織病理学的変化に調べることができる方法を示す、記載されています。

MTACは、タイムリーに、左心室の大幅な高血圧の誘導を可能にします。 ポリウレタン管を適所に曲げられて保持されるように4 30ゲージ針- 図1に見られるように、プロシージャのための主要なツールの製造は、2を必要とする、シンプルで最小です。外科手術自体はTACと比較して、容易に行うことができるよう?...

Watch this Scientific Journal Video about Minimally Invasive Transverse Aortic Constriction in Mice at JoVE.com

Minimally Invasive Transverse Aortic Constriction in Mice

気管挿管と人工呼吸器の使用を排除しながら、低侵襲性横大動脈狭窄（MTAC）は、通常の横方向の大動脈狭窄（TAC）の必需品を節約します。これは、特に翻訳研究における左心室過負荷のハイスループット研究のために非常に望ましい方法であることが分かります。

マウスにおける低侵襲横大動脈狭窄

Minimally invasive transverse aortic constriction (MTAC) is a more desirable method for the constriction of the transverse aorta in mice than standard ...

minimally-invasive-transverse-aortic-constriction-in-mice

Research

JoVE Journal

Medicine

20.3K ビュー.  The University of Hong Kong. 気管挿管と人工呼吸器の使用を排除しながら、低侵襲性横大動脈狭窄（MTAC）は、通常の横方向の大動脈狭窄（TAC）の必需品を節約します。これは、特に翻訳研究における左心室過負荷のハイスループット研究のために非常に望ましい方法であることが分かります。 

ビデオ: Minimally Invasive Transverse Aortic Constriction in Mice

In vitro Measurements of Tracheal Constriction Using Mice

Transgenic and knockout mice have been powerful tools for the investigation of the physiology and pathophysiology of airways1,2. In vitro tensometry of isolated tracheal preparations has proven to be a useful assay of airway smooth muscle (ASM) contractile response in genetically modified mice. These in vitro tracheal preparations are relatively simple, provide a robust response, and retain both functional cholinergic nerve endings and muscle responses, even after long incubations.
Tracheal tensometry also provides a functional assay to study a variety of second messenger signaling pathways that affect contraction of smooth muscle. Contraction in trachea is primarily mediated by parasympathetic, cholinergic nerves that release acetylcholine onto ASM (Figure 1). The major ASM acetylcholine receptors are muscarinic M2 and M3 which are Gi/o and Gq coupled receptors, respectively3,4,5. M3 receptors evoke contraction by coupling to Gq to activate phospholipase C, increase IP3 production and IP3-mediated calcium release from the sarcoplasmic reticulum3,6,7. M2/Gi/o signaling is believed to enhance contractions by inhibition of adenylate cyclase leading to a decrease in cAMP levels5,8,9,10. These pathways constitute the so called "pharmaco-contraction coupling" of airway smooth muscle11. In addition, cholinergic signaling through M2 receptors (and modulated by M3 signaling) involves pathways that depolarize the ASM which in turn activate L-type, voltage-dependent calcium channels (Figure 1) and calcium influx (so called "excitation-contraction coupling")4,7. More detailed reviews on signaling pathways controlling airway constriction can be found4,12. The above pathways appear to be conserved between mice and other species. However, mouse tracheas differ from other species in some signaling pathways. Most prominent is their lack of contractile response to histamine and adenosine13,14, both well-known ASM modulators in humans and other species5,15.
Here we present protocols for the isolation of murine tracheal rings and the in vitro measurement of their contractile output. Included are descriptions of the equipment configuration, trachea ring isolation and contractile measurements. Examples are given for evoking contractions indirectly using high potassium stimulation of nerves and directly by depolarization of ASM muscle to activate voltage-dependent calcium influx (1. high K+, Figure 1). In addition, methods are presented for stimulations of nerves alone using electric field stimulation (2. EFS, Figure 1), or for direct stimulation of ASM muscle using exogenous neurotransmitter applied to the bath (3. exogenous ACH, Figure 1). This flexibility and ease of preparation renders the isolated trachea ring model a robust and functional assay for a number of signaling cascades involved in airway smooth muscle contraction.

In vitro Measurements of Tracheal Constriction Using Mice

Transgenic mice have been extremely useful in ascribing physiological function to genes. As such, research in general, and functional studies of airway, in particular, have undergone a remarkable shift toward murine models. Here we provide protocols for in vitro trachea constriction studies to evaluate smooth muscle function in murine airway.

マウスを用いた気管狭窄のin vitroでの測定で

Transgenic and knockout mice have been powerful tools for the investigation of the physiology and pathophysiology of airways1,2. In vitro tensometry of ...

医学

Ascending Aortic Constriction in Rats for Creation of Pressure Overload Cardiac Hypertrophy Model

Ascending aortic constriction is the most common and successful surgical model for creating pressure overload induced cardiac hypertrophy and heart failure. Here, we describe a detailed surgical procedure for creating pressure overload and cardiac hypertrophy in rats by constriction of the ascending aorta using a small metallic clip. After anesthesia, the trachea is intubated by inserting a cannula through a half way incision made between two cartilage rings of trachea. Then a skin incision is made at the level of the second intercostal space on the left chest wall and muscle layers are cleared to locate the ascending portion of aorta. The ascending aorta is constricted to 50&#8211;60% of its original diameter by application of a small sized titanium clip. Following aortic constriction, the second and third ribs are approximated with prolene sutures. The tracheal cannula is removed once spontaneous breathing was re-established. The animal is allowed to recover on the heating pad by gradually lowering anesthesia. The intensity of pressure overload created by constriction of the ascending aorta is determined by recording the pressure gradient using trans-thoracic two dimensional Doppler-echocardiography. Overall this protocol is useful to study the remodeling events and contractile properties of the heart during the gradual onset and progression from compensated cardiac hypertrophy to heart failure stage.

We describe a stepwise procedure for creating pressure overload and left ventricular hypertrophy in Wistar rats by constriction of the ascending aorta using a small metallic clip. This model is extensively used for studying remodeling changes during cardiac hypertrophy and for identifying and evaluating strategies for regression of such changes.

圧負荷心肥大モデルの作成のためのラットの大動脈狭窄昇順

Ascending aortic constriction is the most common and successful surgical model for creating pressure overload induced cardiac hypertrophy and heart ...

Minimally Invasive Establishment of Murine Orthotopic Bladder Xenografts

Orthotopic bladder cancer xenografts are the gold standard to study molecular cellular manipulations and new therapeutic agents&#160;in vivo. Suitable cell lines are inoculated either by intravesical instillation (model of nonmuscle invasive growth) or intramural injection into the bladder wall (model of invasive growth). Both procedures are complex and highly time-consuming. Additionally, the superficial model has its shortcomings due to the lack of cell lines that are tumorigenic following instillation. Intramural injection, on the other hand, is marred by the invasiveness of the procedure and the associated morbidity for the host mouse.
With these shortcomings in mind, we modified previous methods to develop a minimally invasive approach for creating orthotopic bladder cancer xenografts. Using ultrasound guidance we have successfully performed percutaneous inoculation of the bladder cancer cell lines UM-UC1, UM-UC3 and UM-UC13 into 50 athymic nude. We have been able to demonstrate that this approach is time efficient, precise and safe. With this technique, initially a space is created under the bladder mucosa with PBS, and tumor cells are then injected into this space in a second step. Tumor growth is monitored at regular intervals with bioluminescence imaging and ultrasound. The average tumor volumes increased steadily in in all but one of our 50 mice over the study period.
In our institution, this novel approach, which allows bladder cancer xenograft inoculation in a minimally-invasive, rapid and highly precise way, has replaced the traditional model.

The established technique to inoculate primary invasive orthotopic bladder cancer xenografts requires laparotomy and mobilization of the bladder. This procedure inflicts significant morbidity on the mice, is technically challenging and time-consuming. We therefore developed a high-precision, percutaneous approach utilizing ultrasound guidance.

マウス同所性膀胱異種移植片の最小侵襲性の確立

Orthotopic bladder cancer xenografts are the gold standard to study molecular cellular manipulations and new therapeutic agents&#160;in vivo. Suitable ...

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 ...

Assessment of Cardiac Morphological and Functional Changes in Mouse Model of Transverse Aortic Constriction by Echocardiographic Imaging

Transverse aortic constriction (TAC) in mice has been used as a valuable model to study mechanisms of cardiac hypertrophy and heart failure1. A reliable noninvasive method is essential to assess real-time cardiac morphological and functional changes in animal models of heart disease. Transthoracic echocardiography represents an important tool for noninvasive assessment of cardiac structure and function2. Here we used a high-resolution ultrasound imaging system to monitor myocardial remodeling and heart failure progression over time in a mouse model of TAC. B-mode, M-mode, and Doppler imaging were used to precisely assess cardiac hypertrophy, ventricular dilatation, and functional deterioration in mice following TAC. Color and pulse wave (PW) Doppler imaging was used to noninvasively measure pressure gradient across the aortic constriction created by TAC and to assess transmitral blood flow in mice. Thus transthoracic echocardiographic imaging provides comprehensive noninvasive measurements of cardiac dimensions and function in mouse models of heart disease.

The goal of this protocol is to noninvasively assess cardiac structural and functional changes in a mouse model of heart disease created by transverse aortic constriction, using B- and M-mode echocardiography and color/pulse wave Doppler imaging.

心臓形態学的および心エコーイメージングによって横大動脈狭窄のマウスモデルにおける機能の変更の評価

Transverse aortic constriction (TAC) in mice has been used as a valuable model to study mechanisms of cardiac hypertrophy and heart failure1. A reliable ...

Technique of Minimally Invasive Transverse Aortic Constriction in Mice for Induction of Left Ventricular Hypertrophy

Transverse aortic constriction (TAC) in mice is one of the most commonly used surgical techniques for experimental investigation of pressure overload-induced left ventricular hypertrophy (LVH) and its progression to heart failure. In the majority of the reported investigations, this procedure is performed with intubation and ventilation of the animal which renders it demanding and time-consuming and adds to the surgical burden to the animal. The aim of this protocol is to describe a simplified technique of minimally invasive TAC without intubation and ventilation of mice. Critical steps of the technique are emphasized in order to achieve low mortality and high efficiency in inducing LVH.
Male C57BL/6 mice (10-week-old, 25-30 g, n=60) were anesthetized with a single intraperitoneal injection of a mixture of ketamine and xylazine. In a spontaneously breathing animal following a 3-4 mm upper partial sternotomy, a segment of 6/0 silk suture threaded through the eye of a ligation aid was passed under the aortic arch and tied over a blunted 27-gauge needle. Sham-operated animals underwent the same surgical preparation but without aortic constriction. The efficacy of the procedure in inducing LVH is attested by a significant increase in the heart/body weight ratio. This ratio is obtained at days 3, 7, 14 and 28 after surgery (n = 6 - 10 in each group and each time point). Using our technique, LVH is observed in TAC compared to sham animals from day 7 through day 28. Operative and late (over 28 days) mortalities are both very low at 1.7%.
In conclusion, our cost-effective technique of minimally invasive TAC in mice carries very low operative and post-operative mortalities and is highly efficient in inducing LVH. It simplifies the operative procedure and reduces the strain put on the animal. It can be easily performed by following the critical steps described in this protocol.

The aim of this protocol is to describe step-by-step the technique of minimally invasive transverse aortic constriction (TAC) in mice. By elimination of intubation and ventilation which are mandatory for the commonly used standard procedure, minimally invasive TAC simplifies the operative procedure and reduces the strain put on the animal.

左室肥大の誘導マウスにおける低侵襲の横大動脈狭窄のテクニック

Transverse aortic constriction (TAC) in mice is one of the most commonly used surgical techniques for experimental investigation of pressure ...

A Minimally Invasive Model to Analyze Endochondral Fracture Healing in Mice Under Standardized Biomechanical Conditions

Bone healing models are necessary to analyze the complex mechanisms of fracture healing to improve clinical fracture treatment. During the last decade, an increased use of mouse models in orthopedic research was noted, most probably because mouse models offer a large number of genetically-modified strains and special antibodies for the analysis of molecular mechanisms of fracture healing. To control the biomechanical conditions, well-characterized osteosynthesis techniques are mandatory, also in mice. Here, we report on the design and use of a closed bone healing model to stabilize femur fractures in mice. The intramedullary screw, made of medical-grade stainless steel, provides through fracture compression an axial and rotational stability compared to the mostly used simple intramedullary pins, which show a complete lack of axial and rotational stability. The stability achieved by the intramedullary screw allows the analysis of endochondral healing. A large amount of callus tissue, received after stabilization with the screw, offers ideal conditions to harvest tissue for biochemical and molecular analyses. A further advantage of the use of the screw is the fact that the screw can be inserted into the femur with a minimally invasive technique without inducing damage to the soft tissue. In conclusion, the screw is a unique implant that can ideally be used in closed fracture healing models offering standardized biomechanical conditions.

This protocol describes a minimally invasive osteosynthesis technique using an intramedullary screw for standardized stabilization of femur fractures, which can be used to analyze endochondral bone healing in mice.

癒しの標準化された生体力学的条件下でマウスの軟骨破壊を分析する低侵襲モデル

Bone healing models are necessary to analyze the complex mechanisms of fracture healing to improve clinical fracture treatment. During the last decade, an ...

A Closed-chest Model to Induce Transverse Aortic Constriction in Mice

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to perform a partial thoracotomy to visualize the transverse aortic arch. However, the surgical trauma caused by the thoracotomy in open-chest models changes the respiratory physiology as the ribs are dissected and left unattached after chest closure. To prevent this, we established a minimally invasive, closed chest approach via lateral thoracotomy. Herein we approach the aortic arch via the 2nd intercostal space without entering the chest cavities, leaving the mouse with a less traumatic injury to recover from. We perform this operation using standard laboratory settings for open chest TAC procedures with equal survival rates. Apart from maintaining physiological breathing patterns due to the closed chest approach, the mice seem to benefit by showing rapid recovery, as the less invasive technique appears to facilitate a fast healing process and to reduce immune response after trauma.

Here, we present a protocol of Transverse Aortic constriction (TAC) via a lateral thoracotomy. This technique is a minimally invasive, closed chest surgical procedure aiming to simulate pressure overload and heart failure in mice utilizing standard TAC laboratory settings.

マウスで横方向の大動脈狭窄を誘導するために閉じて胸モデル

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to ...

A Pre-clinical Rat Model for the Study of Ischemia-reperfusion Injury in Reconstructive Microsurgery

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many areas of biomedical research due to its cost-effectiveness and its translation to humans. This protocol describes a method to create a preclinical free skin flap model in rats with ischemia-reperfusion injury. The described 3 cm x 6 cm rat free skin flap model is easily obtained after the placement of several vascular ligatures and the section of the vascular pedicle. Then, 8 h after the ischemic insult and completion of the microsurgical anastomosis, the free skin flap develops the tissue damage. These ischemia-reperfusion injury-related damages can be studied in this model, making it a suitable model for evaluating therapeutic agents to address this pathophysiological process. Furthermore, two main monitoring techniques are described in the protocol for the assessment of this animal model: transit-time ultrasound technology and laser speckle contrast analysis.

Here, we describe a pre-clinical animal model for studying the pathophysiology of ischemia-reperfusion injury in reconstructive microsurgery. This free skin flap model based on the superficial caudal epigastric vessels in the rat may also allow for the evaluation of different therapies and compounds to counteract ischemia-reperfusion injury-related damage.

再建微小外科における虚血再灌流損傷の研究のための前臨床ラットモデル

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many ...

Evaluation of Capnography Sampling Line Compatibility and Accuracy when Used with a Portable Capnography Monitor

Capnography is commonly used to monitor patient&#8217;s ventilatory status. While sidestream capnography has been shown to provide a reliable assessment of end-tidal CO2 (ETCO2), its accuracy is commonly validated using commercial kits composed of a capnography monitor and its matching disposable nasal cannula sampling lines. The purpose of this study was to assess the compatibility and accuracy of cross-paired capnography sampling lines with a single portable bedside capnography monitor. A series of 4 bench tests were performed to evaluate the tensile strength, rise time, ETCO2 accuracy as a function of respiratory rate, and ETCO2 accuracy in the presence of supplemental O2. Each bench test was performed using specialized, validated equipment to allow for a full evaluation of sampling line performance. The 4 bench tests successfully differentiated between sampling lines from different commercial sources and suggested that due to increased rise time and decreased ETCO2 accuracy, not all nasal cannula sampling lines provide reliable clinical data when cross-paired with a commercial capnography monitor. Care should be taken to ensure that any cross-pairing of capnography monitors and disposable sampling lines is fully validated for use across respiratory rates and supplemental O2 flow rates commonly encountered in clinical settings.

The goal of this study was to evaluate the accuracy of capnography sampling lines used in conjunction with a portable bedside capnography monitor. Sampling lines from 7 manufacturers were evaluated for tensile strength, rise time, and ETCO2 accuracy as a function of respiratory rate or supplemental oxygen flow rate.

ポータブル・キャプノグラフィー・モニターで使用した場合の、キャポグラフィー・サンプリング・ラインの適合性と精度の評価

Capnography is commonly used to monitor patient&#8217;s ventilatory status. While sidestream capnography has been shown to provide a reliable assessment ...