The work was supported by grants from NIH/NINDS (NS054883), Craig H. Neilsen Foundation (280072), and the Veterans Administration and Canadian Spinal Research Organization. We thank the Rat Resource and Research Center, University of Missouri, Columbia, Missouri, for providing GFP rats.

全ての動物プロトコールは施設内動物管理使用委員会（IACUC）によって承認された。実験動物のケアと安全のためのNIHガイドラインは厳密に従っている。外科的処置による動物は、十分に痛みや不快感を最小限に抑えるために処理した。 1脊髄Curgeryおよび細胞移植<ol><li>すべての手術の前にオートクレーブ手術器具。異なる動物での手続きの間に商品から病原体や微生物の汚染物質を除去するためにホットビーズ滅菌器（ファイン科学ツール）を使用します。手術中に無菌手術用手袋、ガウン、ドレープを使用してください。各外科的処置のための無菌手術法を採用しています。 </li><li>腹腔内注射（ 腹腔内 ）を介して投与ケタミンの組合せ（2ミリリットル/ kg）を（25 mg / ml）を、キシラジン（1.3 mg / ml）で、およびアセプロマジン（0.25 mg / ml）を用いて雌のフィッシャー344ラットを麻酔。 </li><li>後ろの領域を剃るとベタジンとエタノールで繰り返し肌をきれいにします。</li><li> ＃10ブレードを用いて皮膚をカットし、ぶっきらぼうに筋肉層を分析。 、椎骨を分離するのT3椎骨を公開し、先の細い骨鉗子を使用して、背側椎弓切除術を実行するためにメスを使用してください。 </li><li>長手方向に硬膜を切開し、脊髄断端の間に約1mm吻側尾ギャップを作成するために虹彩切除鋏と微小誤嚥の組み合わせを用いてT4レベルで脊髄を横断する。 </li><li> 3-0 Vycrilと完全に出血を停止し、その後、縫合糸の筋肉まで2分と創傷クリップで皮膚を閉じ - 約1待ちます。 </li><li>皮下手術直後乳酸リンゲル（5ミリリットル）、ブプレノルフィン（0.035ミリグラム/ kg）であり、アンピシリン（33 mgの/ kg）を注入し、目を覚ましまで暖かいインキュベーター内でラットを維持する。 </li><li>注入リンゲル1日2回、3日間または痛みや苦痛の徴候が消えるまで、アンピシリン一日一回、10日までのソリューション、およびブプレノルフィン。 Fまで他の動物の会社に動物を返さないでくださいully回復した。 </li><li>手動で反射排尿の確立まで、約2週間1日2回、膀胱を空にしてから、必要に応じて生存全体で一日一回、膀胱を空にします。 </li><li>二週間後、上述したように、SCIラットを再麻酔し、脊髄損傷部位を再度露出させる。病変部位に移植された細胞を保持するために閉じた硬膜を保管してください。 </li><li>病変空洞の震源地と複数の噴射に病変の吻側および尾側インターフェースに、細胞液10μl（3.5×10 5μl）を、E14ユビキタスGFPトランスジェニックラットの胚から採取し、フィブリノゲンに埋め込 ​​まれ、トロンビン5,6を注入ハミルトンシリンジに接続さ40μmの内径引っ張らガラスマイクロピペットを使用して、サイ​​ト、。 </li><li>筋肉層を縫合し、創傷クリップで皮膚を閉じます。 </li><li> 1.8 - ステップ1.6で説明したように手術後の注入と膀胱のケアを行います。 </li><li>一週間であること前部灌流は、脊髄7に腹腔内に逆行ラベル交感神経節前ニューロンへの0.5％フルオロゴールド（FG、蒸留水0.4 ml）を注入する。 </li></ol> 2トランスミッタ注入<ol><li>滅菌のために室温で（10時間まで）で2時間 - 少なくとも1 transmittersin 2％グルタルアルデヒド溶液（蒸留水96ミリリットルの50％glutaraldhyde 4 ml）に浸す。 </li><li>徹底的に滅菌0.9％食塩水で3回の送信機を洗浄し、使用（わずか1時間）まで生理食塩水に保管してください。 </li><li> E14細胞移植後8週間（10週間損傷後）、細胞移植およびナイーブ対照ラットの有無にかかわらず、SCIを受けたラットreanesthetize。 </li><li>腹部と後肢を剃る。ベタジンで皮膚をきれいにしてください。仰臥位で手術台にラットを置きます。 </li><li> ＃15ブレードを使用して右側の腹側腹部と太ももの内側の皮膚を切開。 </li><li>秒を切ってubcutaneous結合組織は、小さなハサミを使用して大腿血管や神経の束を露出させる。 </li><li>静脈と湾曲した先端の微細鉗子を用いて神経から大腿動脈を分離します。 </li><li>動脈の下に3絹縫合糸を入れて、それぞれの縫合糸の緩い結び目を作る。 </li><li>その後のカテーテル挿入のための血管拡張を誘発するために動脈の表面に0.1ミリリットルリドカイン（2％）を適用します。 </li><li>近位に緩い絹縫合糸を延伸して恒久的な絹の結び目と一時ブロックと遠位に容器を固定します。 </li><li> 20ゲージの湾曲した針を用いて動脈を穿刺し、カテーテル挿入ツールを使用してテレメータカテーテル（長さ8cm）の先端を挿入します。 </li><li>これにより、胸部大動脈に先端を置く4センチメートルまで吻側にカテーテルを挿入します。 </li><li>大腿動脈の周りに3絹縫合糸を結ぶことにより、血管内にカテーテルを固定する。 </li><li>胸郭とtの尾側縁部との間の横腹に沿って皮下ポケットを作る彼鈍はさみで膝の範囲の中で最も頭側延長。 </li><li>過度の運動を避けるために、送信機を周囲の結合組織にポケットや縫合に送信機本体を挿入します。 </li><li> 6号絹糸で皮膚を縫合。 </li><li>皮下手術直後乳酸リンゲル（5ミリリットル）、ブプレノルフィン（0.035ミリグラム/ kg）であり、アンピシリン（33 mgの/ kg）を注入し、目を覚ましまで暖かいインキュベーター内でラットを維持する。 </li></ol> 3基礎平均動脈圧（MAP）および心拍数（HR）の記録<ol><li>早ければ、送信機移植後1日として、受信機パッドの上に単一の動物を入れて、上の送信機の電源を入れます。動物を慣らすと心血管パラメータを安定させるために15分 - 約10のを待ちます。 </li><li>少なくとも1時間、コンピュータ化されたデータ収集システムのパルス動脈圧から導出されたレコードの休止MAPおよびHR、。データごとに5秒を収集します。 </li><li> ANを監視imals継続的に可視痙攣発生時のデータポイントを削除します。各動物について、平均データ点は平均値を得る。 </li></ol> 4。結腸直腸拡張によって誘発される自律神経反射異常<ol><li>トランスミッタ受信機内部の食物ペレットに付属のタオルでNSC-グラフトまたはSCIのコントロールラットを拘束。通常、ラットは処置の間に協調的に滞在することができます。 </li><li>約2cmのために、直腸にラテックスバルーンが先端に付いたカテーテルを挿入し、テープ8尾に固定します。 </li><li>上の送信機の電源を入れ、10を待つ - 血圧が予備挿入ベースラインに戻ることができるように、15分。 </li><li>約30mmHgの圧力を生成するために、ゆっくりと1分間空気1.4mlの10秒間に渡りバルーンの膨張によって、結腸直腸膨張を誘発する。 </li><li>レコードマップおよびHR 1分前、途中で1分間、および結腸直腸拡張後の1分;サンプルデータ3分間の手順の間ごとに3秒。 </li><li>2つの試験の間に少なくとも15分間の回復間隔で動物あたり3試験 - 2を実行します。 </li><li>がない場合はさらなる評価上記の二重線量麻酔の組み合わせで過量投与した動物（ 腹腔内 ）。 4％パラホルムアルデヒドに続いて食塩水で動物を灌流する。 </li><li>各動物について、前及び結腸直腸膨張の間に値を平均;各試験のベースラインおよび膨満によって誘発されるMAPおよびHRの変化との差を計算し、 2を超える平均 - 3試験の平均値を得た。 </li></ol> 5。送信機のクリーニング<ol><li>麻酔後であるが、灌流の前に、動物の体からトランスミッターを外します。クリーニングまで蒸留水を満たしたビーカーに直ちに浸し;テレメータ装置の乾燥を避ける。 </li><li>室温で24時間、1％ターグ-A-ザイム、酵素洗浄溶液（10g / Lの水）にテレメータを転送する。 </li><li>のReGel鈍30ゲージの針を用いて送信機のカテーテルの先端のReGel注射器に接続されている。 </li><li>慎重に折り畳まれた軟組織を使用して送信機を乾燥させ、元のプラスチックトレイに格納します。 </li></ol>

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V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Autonomic+assessment+of+animals+with+spinal+cord+injury:+tools,+techniques+and+translation.">Autonomic assessment of animals with spinal cord injury: tools, techniques and translation.</a> Spinal Cord. 47, 2-35 (2009).</li><li>Mayorov, D. N., Adams, M. A., Krassioukov, 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=Telemetric+blood+pressure+monitoring+in+conscious+rats+before+and+after+compression+injury+of+spinal+cord.">Telemetric blood pressure monitoring in conscious rats before and after compression injury of spinal cord.</a> J Neurotrauma. 18, 727-736 (2001).</li><li>Hou, S., Tom, V. 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著者らは、開示することは何もない。

伝統的に、流体充填されたカニューレを動脈に挿入し、各動物11における端末スナップショットとして心臓血管のパラメータを記録するために圧力変換器に接続されている。連続して長い時間のための心血管系のパフォーマンスを監視するには、無線遠隔測定システムは、多くの研究室で使用される。このより洗練されたツールは、自由に動物を移動させること、意識的に血圧を記録...

心血管機能障害は、高レベルでの脊髄損傷（SCI）後に発生します。それは、無秩序な血圧および心拍数安静時、起立性低血圧、運動誘発性低血圧、および損傷レベル1,2下の感覚刺激に反応して高血圧や圧反射媒介徐脈のエピソードを特徴自律神経反射異常で明らかにされている。これらの症状は、脊髄損傷患者の日常生活を妨害する。したがって、SCIと実験的な治療法と動物での心血管系の変化の調査のための効果的なツールを確立することが重要である。 動物において心臓血管機能を調べるために、いくつかの技術は、血圧および心拍数をモニターするために使用されている。非侵襲尾カフ末梢血圧3を測定するために使用することができるのに対し、中央の心血管パラメータは、カニューレ挿入およびテレメトリー記録することができる。他の方法、テレと比較するとmetryは、それが自由に動く動物および心血管機能4の長期モニタリングに連続記録を可能にする主な利点を有している。 SCIの動物モデルにおいて、実験的刺激後の末梢血圧の変化が検出されるのに十分な大きさでなくてもよい。したがって、適切な心臓監視技術は、SCIを持つ動物のために選択する必要があります。 本研究では、無線遠隔測定システムは、完全な脊髄離断後の成体ラットにおける血管機能をモニターするために導入した。ラットは、損傷部位に同系ラットの胚14日目（E14）脳幹由来神経幹細胞（BS-NSCの）の移植片を受けた。けがのない移植ナイーブラットは、損傷していないラットを対照とした。遠隔測定の手順は、送信機滅菌注入（ 図1）、基底心血管パラメータの記録、結腸直腸拡張によって誘発される応答、および送信機の洗浄とを含むストレージ。

<table border="0" cellpadding="0" cellspacing="0" width="580">
	<tbody>
		<tr height="21">
			<td colspan="2" height="21" style="height:21px;width:315px;">Table of Specific Materials/Equipment:</td>
			<td style="width:103px;"></td>
			<td style="width:164px;"></td>
		</tr>
		<tr height="20">
			<td height="41" rowspan="2" style="height:41px;width:120px;">Reagents</td>
			<td rowspan="2" style="width:195px;">Company</td>
			<td rowspan="2" style="width:103px;">Catalogue</td>
			<td rowspan="2" style="width:164px;">Comment</td>
		</tr>
		<tr height="21">
		</tr>
		<tr height="62">
			<td height="83" rowspan="2" style="height:83px;width:120px;">Fibrinogen (rat)</td>
			<td rowspan="2" style="width:195px;">Sigma</td>
			<td rowspan="2" style="width:103px;">F6755-25MG</td>
			<td rowspan="2" style="width:164px;">2 hr at 37oC to dissovle</td>
		</tr>
		<tr height="21">
		</tr>
		<tr height="59">
			<td height="80" rowspan="2" style="height:80px;width:120px;">Thrombin (rat)</td>
			<td rowspan="2" style="width:195px;">Sigma</td>
			<td rowspan="2" style="width:103px;">T5772-100UN</td>
			<td rowspan="2" style="width:164px;">Dissovle in 10 mM CaCl2</td>
		</tr>
		<tr height="21">
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:120px;">1% Terg-A-Zyme</td>
			<td style="width:195px;">Sigma</td>
			<td style="width:103px;">Z273287</td>
			<td style="width:164px;">Enzymatic solution for telemeter cleaning</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:120px;">Fluorogold</td>
			<td style="width:195px;">Fluorochrome</td>
			<td style="width:103px;"></td>
			<td style="width:164px;">Dissovle in distilled water and avoid light</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:120px;">Telemeter&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160; (PA-C40)</td>
			<td style="width:195px;">Data Sciences International</td>
			<td style="width:103px;"></td>
			<td style="width:164px;"></td>
		</tr>
		<tr height="105">
			<td height="105" style="height:105px;width:120px;">Telementric recording and analysis system</td>
			<td style="width:195px;">Data Sciences International</td>
			<td style="width:103px;"></td>
			<td style="width:164px;">Signal stimulator, Data Exchange Matrix, receivers, Ambient pressure reference monitor</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:120px;">Balloon-tipped catheter</td>
			<td style="width:195px;">Edward Lifesciences</td>
			<td style="width:103px;">111F7-P</td>
			<td style="width:164px;">For colorectal distension</td>
		</tr>
	</tbody>
</table>

a radio-telemetric system to monitor cardiovascular function in rats with spinal cord transection and embryonic neural stem cell grafts

ハイ胸部または頸髄損傷（SCI）は、心臓血管機能障害につながることができます。心臓血管のパラメータを監視するために、または緑色蛍光タンパク質を発現する胚性脳幹由来神経幹細胞の移植なしT4脊髄離断を受けたラットの大腿動脈に無線送信機に接続されたカテーテルを移植した。そのようなカニューレ挿入またはテールカフなどの他の方法に比べて、遠隔測定が連続的に自由に移動する動物における血圧および心拍数を監視することが有利である。また、長期複数のデータの取得が可能である。脊髄損傷ラットでは、結腸直腸拡張に応じて、非拘束状態と自律神経反射異常の下で基礎心血管データが正常に記録された。送信機は脊髄離断前に移植される場合に加えて、SCIの前と後の心血管パラメータは同じラットに比較することができる。記載されtelemetの1つの制限ryの手順は、大腿動脈内注入は、同側の後肢への血液供給に影響を与えることができることである。

上記の遠隔測定技術を用いて、成功した脊髄損傷動物における心血管パラメータを記録した。 HRは以前の報告9と一致して、ナイーブ動物と比較して増加したのに対し、単独のSCIと動物では、MAPは、有意に減少した。 BS-NSC移植した動物では、MAPおよびHRはナイーブ動物（ 図2）で測定されたレベルに近づいた。侵害刺激は、MAPの上昇およびHR 3,8の減少をもたらした?...

Watch this Scientific Journal Video about A Radio-telemetric System to Monitor Cardiovascular Function in Rats with Spinal Cord Transection and Embryonic Neural Stem Cell Grafts at JoVE.com

A Radio-telemetric System to Monitor Cardiovascular Function in Rats with Spinal Cord Transection and Embryonic Neural Stem Cell Grafts

私たちは、8週間、胚性脳幹神経幹細胞は、病変部位に移植後T4脊髄離断ラットの心臓血管のパラメータを記録するために無線遠隔測定システムを使用するためのプロトコルを提示する。テレメトリーを正確に意識的に自由に移動し、脊髄損傷ラットにおける心血管機能を評価するための高度な技術です。

ラジオ遠隔測定システムは、脊髄離断および胚性神経細胞移植片幹ラットの心臓血管機能を監視する

High thoracic or cervical spinal cord injury (SCI) can lead to cardiovascular dysfunction. To monitor cardiovascular parameters, we implanted a catheter ...

a-radio-telemetric-system-to-monitor-cardiovascular-function-rats

Research

JoVE Journal

Medicine

10.4K ビュー.  Drexel University College of Medicine. 私たちは、8週間、胚性脳幹神経幹細胞は、病変部位に移植後T4脊髄離断ラットの心臓血管のパラメータを記録するために無線遠隔測定システムを使用するためのプロトコルを提示する。テレメトリーを正確に意識的に自由に移動し、脊髄損傷ラットにおける心血管機能を評価するための高度な技術です。 

ビデオ: A Radio-telemetric System to Monitor Cardiovascular Function in Rats with Spinal Cord Transection and Embryonic Neural Stem Cell Grafts

Assessing Endothelial Vasodilator Function with the Endo-PAT 2000

The endothelium is a delicate monolayer of cells that lines all blood vessels, and which comprises the systemic and lymphatic capillaries. By virtue of the panoply of paracrine factors that it secretes, the endothelium regulates the contractile and proliferative state of the underlying vascular smooth muscle, as well as the interaction of the vessel wall with circulating blood elements. Because of its central role in mediating vessel tone and growth, its position as gateway to circulating immune cells, and its local regulation of hemostasis and coagulation, the the properly functioning endothelium is the key to cardiovascular health. Conversely, the earliest disorder in most vascular diseases is endothelial dysfunction.
In the arterial circulation, the healthy endothelium generally exerts a vasodilator influence on the vascular smooth muscle. There are a number of methods to assess endothelial vasodilator function. The Endo-PAT 2000 is a new device that is used to assess endothelial vasodilator function in a rapid and non-invasive fashion. Unlike the commonly used technique of duplex ultra-sonography to assess flow-mediated vasodilation, it is totally non-operator-dependent, and the equipment is an order of magnitude less expensive. The device records endothelium-mediated changes in the digital pulse waveform known as the PAT ( peripheral Arterial Tone) signal, measured with a pair of novel modified plethysmographic probes situated on the finger index of each hand. Endothelium-mediated changes in the PAT signal are elicited by creating a downstream hyperemic response. Hyperemia is induced by occluding blood flow through the brachial artery for 5 minutes using an inflatable cuff on one hand. The response to reactive hyperemia is calculated automatically by the system. A PAT ratio is created using the post and pre occlusion values. These values are normalized to measurements from the contra-lateral arm, which serves as control for non-endothelial dependent systemic effects. Most notably, this normalization controls for fluctuations in sympathetic nerve outflow that may induce changes in peripheral arterial tone that are superimposed on the hyperemic response.
In this video we demonstrate how to use the Endo-PAT 2000 to perform a clinically relevant assessment of endothelial vasodilator function.

A noninvasive procedure to assess endothelial function is demonstrated using the Endo-PAT 2000.

遠藤- PAT 2000で内皮血管拡張機能を評価する

The endothelium is a delicate monolayer of cells that lines all blood vessels, and which comprises the systemic and lymphatic capillaries. By virtue of ...

医学

Acute and Chronic Tactile Sensory Testing after Spinal Cord Injury in Rats

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory testing in rat SCI models is needed. However, until recently, no single testing approach had been validated for SCI so that standardized methods have not been implemented across labs. Additionally, available testing methods could not be implemented acutely or when severe motor impairments existed, preventing studies of the development of SCI-induced allodynia3. Here we present two validated sensory testing methods using von Frey Hair (VFH) monofilaments which quantify changes in tactile sensory thresholds after SCI4-5. One test is the well-established Up-Down test which demonstrates high sensitivity and specificity across different SCI severities when tested chronically5. The other test is a newly-developed dorsal VFH test that can be applied acutely after SCI when allodynia develops, prior to motor recovery4-5. Each VFH monofilament applies a calibrated force when touched to the skin of the hind paw until it bends. In the up-down method, alternating VFHs of higher or lower forces are used on the plantar L5 dermatome to delineate flexor withdrawal thresholds. Successively higher forces are applied until withdrawal occurs then lower force VFHs are used until withdrawal ceases. The tactile threshold reflects the force required to elicit withdrawal in 50% of the stimuli. For the new test, each VFH is applied to the dorsal L5 dermatome of the paw while the rat is supported by the examiner. The VFH stimulation occurs in ascending order of force until at least 2 of 3 applications at a given force produces paw withdrawal. Tactile sensory threshold is the lowest force to elicit withdrawal 66% of the time. Acclimation, testing and scoring procedures are described. Aberrant trials that require a retest and typical trials are defined. Animal use was approved by Ohio State University Animal Care and Use Committee.

We describe two tactile sensory testing methods for acute or chronic periods of spinal cord injury in rats. These validated procedures can detect the development and maintenance of allodynia-like sensations.

ラット脊髄損傷後の急性および慢性の触覚官能検査

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory ...

A Contusion Model of Severe Spinal Cord Injury in Rats

The translational potential of novel treatments should be investigated in severe spinal cord injury (SCI) contusion models. A detailed methodology is described to obtain a consistent model of severe SCI. Use of a stereotactic frame and computer controlled impactor allows for creation of reproducible injury. Hypothermia and urinary tract infection pose significant challenges in the post-operative period. Careful monitoring of animals with daily weight recording and bladder expression allows for early detection of post-operative complications. The functional results of this contusion model are equivalent to transection models. The contusion model can be utilized to evaluate the efficacy of both neuroprotective and neuroregenerative approaches.

A contusion model of severe spinal cord injury is described. Detailed pre-operative, operative and post-operative steps are described to obtain a consistent model.

ラットにおける重度の脊髄損傷挫傷モデル

The translational potential of novel treatments should be investigated in severe spinal cord injury (SCI) contusion models. A detailed methodology is ...

Synergetic Use of Neural Precursor Cells and Self-assembling Peptides in Experimental Cervical Spinal Cord Injury

Spinal cord injuries (SCI) cause serious neurological impairment and psychological, economic, and social consequences for patients and their families. Clinically, more than 50% of SCI affect the cervical spine1. As a consequence of the primary injury, a cascade of secondary mechanisms including inflammation, apoptosis, and demyelination occur finally leading to tissue scarring and development of intramedullary cavities2,3. Both represent physical and chemical barriers to cell transplantation, integration, and regeneration. Therefore, shaping the inhibitory environment and bridging cavities to create a supportive milieu for cell transplantation and regeneration is a promising therapeutic target4. Here, a contusion/compression model of cervical SCI using an aneurysm clip is described. This model is more clinically relevant than other experimental models, since complete transection or ruptures of the cord are rare. Also in comparison to the weight drop model, which in particular damage the dorsum columns, circumferential compression of the spinal cord appears advantageous. Clip closing force and duration can be adjusted to achieve different injury severity. A ring spring facilitates precise calibration and constancy of clip force. Under physiological conditions, synthetic self-assembling peptides (SAP) self-assemble into nanofibers and thus, are appealing for application in SCI5. They can be injected directly into the lesion minimizing damage to the cord. SAPs are biocompatible structures erecting scaffolds to bridge intramedullary cavities and thus, equip the damaged cord for regenerative treatments. K2(QL)6K2 (QL6) is a novel SAP introduced by Dong et al.6 In comparison to other peptides, QL6 self-assembles into &#946;-sheets at neutral pH6.14 days after SCI, after the acute stage, SAPs are injected into the center of the lesion and neural precursor cells (NPC) are injected into adjacent dorsal columns. In order to support cell survival, transplantation is combined with continuous subdural administration of growth factors by osmotic micro pumps for 7 days.

Treating cervical spinal cord injury with both self-assembling peptides (SAP) and neural precursor cells (NPC), together with growth factors, is a promising approach to promote regeneration and recovery. A contusion/compression aneurysm clip rat model of cervical SCI and combined treatment involving SAP injection and NPC transplantation is established.

実験的子宮頸脊髄損傷における神経前駆細胞と自己組織化ペプチドの相乗使用

Spinal cord injuries (SCI) cause serious neurological impairment and psychological, economic, and social consequences for patients and their families. ...

Neural Stem Cell Transplantation in Experimental Contusive Model of Spinal Cord Injury

Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can be used both to investigate the biological responses to injury and to test potential therapies. Contusion or compression injury delivered to the surgically exposed spinal cord are the most widely used models of the pathology. In this report the experimental contusion is performed by using the Infinite Horizon (IH) Impactor device, which allows the creation of a reproducible injury animal model through definition of specific injury parameters. Stem cell transplantation is commonly considered a potentially useful strategy for curing this debilitating condition. Numerous studies have evaluated the effects of transplanting a variety of stem cells. Here we demonstrate an adapted method for spinal cord injury followed by tail vein injection of cells in CD1 mice. In short, we provide procedures for: i) cell labeling with a vital tracer, ii) pre-operative care of mice, iii) execution of a contusive spinal cord injury, and iv) intravenous administration of post mortem neural precursors. This contusion model can be utilized to evaluate the efficacy and safety of stem cell transplantation in a regenerative medicine approach.

Spinal cord injury is a traumatic condition that causes severe morbidity and high mortality. In this work we describe in detail a contusion model of spinal cord injury in mice followed by a transplantation of neural stem cells.

脊髄損傷の実験的挫傷モデルにおける神経幹細胞移植

Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can ...

Photothrombosis-induced Focal Ischemia as a Model of Spinal Cord Injury in Mice

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) below the site of injury. The secondary ischemia that develops following the initial mechanical insult is a serious complication of the SCI and severely impairs the function and viability of surviving neuronal and non-neuronal cells in the SC. In addition, ischemia is also responsible for the growth of lesion during chronic phase of injury and interferes with the cellular repair and healing processes. Thus there is a need to develop a spinal cord ischemia model for studying the mechanisms of ischemia-induced pathology. Focal ischemia induced by photothrombosis (PT) is a minimally invasive and very well established procedure used to investigate the pathology of ischemia-induced cell death in the brain. Here, we describe the use of PT to induce an ischemic lesion in the spinal cord of mice. Following retro-orbital sinus injection of Rose Bengal, the posterior spinal vein and other capillaries on the dorsal surface of SC were irradiated with a green light resulting in the formation of a thrombus and thus ischemia in the affected region. Results from histology and immunochemistry studies show that PT-induced ischemia caused spinal cord infarction, loss of neurons and reactive gliosis. Using this technique a highly reproducible and relatively easy model of SCI in mice can be achieved that would serve the purpose of scientific investigations into the mechanisms of ischemia induced cell death as well as the efficacy of neuroprotective drugs. This model will also allow exploration of the pathological changes that occur following SCI in live mice like axonal degeneration and regeneration, neuronal and astrocytic Ca2+ signaling using two-photon microscopy.

Photothrombosis is a minimally invasive and highly reproducible procedure to induce focal ischemia in the spinal cord and serves as a model of spinal cord injury in mice.

マウスの脊髄損傷のモデルとしての光血栓誘発性局所虚血

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) ...

A Neonatal Mouse Spinal Cord Compression Injury Model

Spinal cord injury (SCI) typically causes devastating neurological deficits, particularly through damage to fibers descending from the brain to the spinal cord. A major current area of research is focused on the mechanisms of adaptive plasticity that underlie spontaneous or induced functional recovery following SCI. Spontaneous functional recovery is reported to be greater early in life, raising interesting questions about how adaptive plasticity changes as the spinal cord develops. To facilitate investigation of this dynamic, we have developed a SCI model in the neonatal mouse. The model has relevance for pediatric SCI, which is too little studied. Because neural plasticity in the adult involves some of the same mechanisms as neural plasticity in early life1, this model may potentially have some relevance also for adult SCI. Here we describe the entire procedure for generating a reproducible spinal cord compression (SCC) injury in the neonatal mouse as early as postnatal (P) day 1. SCC is achieved by performing a laminectomy at a given spinal level (here described at thoracic levels 9-11) and then using a modified Yasargil aneurysm mini-clip to rapidly compress and decompress the spinal cord. As previously described, the injured neonatal mice can be tested for behavioral deficits or sacrificed for ex vivo physiological analysis of synaptic connectivity using electrophysiological and high-throughput optical recording techniques1. Earlier and ongoing studies using behavioral and physiological assessment have demonstrated a dramatic, acute impairment of hindlimb motility followed by a complete functional recovery within 2 weeks, and the first evidence of changes in functional circuitry at the level of identified descending synaptic connections1.

This article describes a method for generating a reproducible spinal cord compression injury (SCI) in the neonatal mouse. The model provides an advantageous platform for studying mechanisms of adaptive plasticity that underlie spontaneous functional recovery.

新生児マウス脊髄圧縮損傷モデル

Spinal cord injury (SCI) typically causes devastating neurological deficits, particularly through damage to fibers descending from the brain to the spinal ...

Vein Interposition Model: A Suitable Model to Study Bypass Graft Patency

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. Research models for bypass graft failure are essential for a better understanding of pathobiological and pathophysiological processes during graft patency loss. Large animal models, such as pigs or sheep, resemble human anatomical structures but require special facilities and equipment. This video describes a rat vein interposition model to investigate vein graft patency loss. Rats are inexpensive and easy to handle. Compared to mouse models, the convenient size of rats permits better operability and enables a sufficient amount of material to be obtained for further diverse analysis. In brief, the inferior epigastric vein of a donor rat is harvested and used to replace a segment of the femoral artery. Anastomosis is conducted via single stitches and sealed with fibrin glue. Graft patency can be monitored non-invasively using duplex sonography. Myointimal hyperplasia, which is the main cause for graft patency loss, develops progressively over time and can be calculated from histological cross sections.

This video demonstrates a model to study the development of myointimal hyperplasia after venous interposition surgery in rats.

静脈介在モデル：バイパスグラフト開存を研究するための適切なモデル

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. ...

Studying the Hypothalamic Insulin Signal to Peripheral Glucose Intolerance with a Continuous Drug Infusion System into the Mouse Brain

Insulin regulates systematic metabolism in the hypothalamus and the peripheral insulin response. An inflammatory reaction in peripheral adipose tissues contributes to type 2 diabetes mellitus (T2DM) development and appetite regulation in the hypothalamus. Chemokine CCL5 and C-C chemokine receptor type 5 (CCR5) levels have been suggested to mediate arteriosclerosis and glucose intolerance in type 2 diabetes mellitus (T2DM). In addition, CCL5 plays a neuroendocrine role in the hypothalamus by regulating food intake and body temperature, thus, prompting us to investigate its function in hypothalamic insulin signaling and the regulation of peripheral glucose metabolism.
The micro-osmotic pump brain infusion system is a quick and precise way to manipulate CCL5 function and study its effect in the brain. It also provides a convenient alternative approach to generating a transgenic knockout animal. In this system, CCL5 signaling was blocked by intracerebroventricular (ICV) infusion of its antagonist, MetCCL5, using a micro-osmotic pump. The peripheral glucose metabolism and insulin responsiveness was detected by the Oral Glucose Tolerance Test (OGTT) and Insulin Tolerance Test (ITT). Insulin signaling activity was then analyzed by protein blot from tissue samples derived from the animals.
After 7-14 days of MetCCL5 infusion, the glucose metabolism and insulin responsiveness was impaired in mice, as seen in the results of the OGTT and ITT. The IRS-1 serine302 phosphorylation was increased and the Akt activity was reduced in mice hypothalamic neurons following CCL5 inhibition. Altogether, our data suggest that blocking CCL5 in the mouse brain increases the phosphorylation of IRS-1 S302 and interrupts hypothalamic insulin signaling, leading to a decrease in insulin function in peripheral tissues as well as the impairment of glucose metabolism.

This protocol studies the role of chemokine (C-C motif) ligand 5 (CCL5) in the hypothalamus by delivering an antagonist, MetCCL5, into the mouse brain using a micro-osmotic pump brain infusion system. This transient inhibition of CCL5 activity interrupted hypothalamic insulin signaling, leading to glucose intolerance and peripheral systemic insulin sensitivity.

マウスの脳内へ連続的な薬剤注入システムと末梢グルコース不耐症に視床下部のインスリン シグナルを勉強

Insulin regulates systematic metabolism in the hypothalamus and the peripheral insulin response. An inflammatory reaction in peripheral adipose tissues ...

Balloon-based Injury to Induce Myointimal Hyperplasia in the Mouse Abdominal Aorta

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine balloon denudation model, which is comparable with established vessel injury models in large animals. The aorta denudation model with balloon catheters mimics the clinical setting and leads to comparable pathobiological and physiological changes. Briefly, after performing a horizontal incision in the aorta abdominalis, a balloon catheter will be inserted into the vessel, inflated, and introduced retrogradely. Inflation of the balloon will lead to intima injury and overdistension of the vessel. After removing the catheter, the aortic incision will be closed with single stiches. The model shown in this article is reproducible, easy to perform, and can be established quickly and reliably. It is especially suitable for evaluating expensive experimental therapeutic agents, which can be applied in an economical fashion. By using different knockout-mouse strains, the impact of different genes on MH development can be assessed.

This article demonstrates a murine model to study the development of myointimal hyperplasia (MH) after aortic balloon injury.

マウスの腹部大動脈 Myointimal 過形成を誘導するために気球を利用した傷害

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine ...