This work was supported by funding from the March of Dimes FY12-133 (S.K.E.) and the National Institutes of Health R01HD037831 (S.K.E.). We would like to thank Dr. Deborah Frank for critical review of the manuscript.

注：国立衛生研究所が定める動物の管理と使用のためのガイドラインを遵守し、すべての動物の手順。すべてのプロトコルは、セントルイスのワシントン大学の動物実験委員会によって承認された。 注：このプロトコルの具体的な材料や設備材料および装置テーブルを一覧表示されます。 雌マウスの1時限繁殖<ol><li>男性との2時間の時間ウィンドウで生後2と6ヶ月の間に成人女性を繁殖。具体的な繁殖時間限り、実験の全体​​のための一貫性が保たれているように必要とされていません。 </li><li>交尾栓の存在によって妊娠を確認してください。プラグは初日後性交0（dpc0）と見られた日と時間を指定します。圧力記録を開始する際に特定の時間を覚えています。 </li><li> dpc8-13に手術を行います。正確なDPCはマウス系統に依存します。 </li><li>マウスは、手術の前にPE妊娠しているかどうかを判断するには麻酔を投与する前に、撮像システムに結合された40 MHzのリニアアレイプローブ腹部のrform音波イメージング。これは妊娠嚢の可視化を可能にします。超音波システムが利用できない場合、マウスの体重を使用する。 注：dpc0とdpc8と腹部の膨満の間に少なくとも2グラムの体重増加は妊娠の良い指標である。 </li></ol> 2.テレメトリ手術準備<ol><li>手術前24時間以内の楽器、ガーゼ、綿棒、外科用ドレープ、及びピペットチップを滅菌するオートクレーブを使用してください。 </li><li> 70％エタノールで清掃手術表面。 </li><li> （げっ歯類に特化した）加熱パッドとビーズ滅菌器の電源をオンにします。 </li></ol> 3.テレメトリ手術<ol><li>麻酔は、1~2リットル/分の酸素流量で4~5％のイソフルランを使用。 </li><li>マウスは麻酔の兆候を示すまで待ってから、ブプレノルフィンを皮下注射する（鎮痛剤：20グラム当たり0.1ミリリットル）。 </li><li>膀胱へのダウンリブの底からマウスの腹部を剃る。 </li><li>予め温め加熱パッド上で手術テーブルにマウスを持参し、乾燥を防ぐために眼軟膏をマウスの目をカバーしています。 </li><li>無菌テクニックを開始します。滅菌手袋を着用し、ヨウ素とエタノールスクラブを配置している無菌面としてラッパーを保つ。 </li><li>パッケージまたは生理食塩水からテレメータを取り外し、滅菌ガーゼの上に配置するために滅菌ピンセットを使用しています。 <ol><li>ゲルとカテーテル先端を埋めるために（テレメータサプライヤーによって提供される）注射器を使用してください。穏やかにカテーテル内にゲルを描画するのに十分な負圧を生成するカテーテルを絞る。 </li></ol></li><li>ピペットで無菌のチップを置きます。ピペット2外科接着剤のμL及び滅菌表面にピペットを設定する。 </li><li>ヨウ素とした後、70％エタノールでエリアを綿棒、手術部位を清掃する。 </li><li>十分な大きさの手術ドレープの穴をカットマウスオーバー切開と場所のために必要な面積のために。 </li><li>マウスが完全に切開を行う前に麻酔をかけていることを確認するために足ピンチ（ペダル応答）を実行します。 </li><li>小さな垂直正中切開を行います。次に、基本的な体壁筋を通して類似の切開を作る。 </li><li>優しく体腔から子宮を引き出し、生存可能であり、ほとんどの子犬を含む子宮角を見つけます。 <ol><li>オプション - miRNAの追加については、テレメータの挿入前に子宮の筋肉層にウイルスベクターを注入する。 </li></ol></li><li>エッジスプリングはさみをカットする3ミリメートルと子宮角の先端に小さな切開を行います。 </li><li>子宮壁と胎児の嚢の間、圧迫することなく、カテーテルを通します。その脱落を減らすために、いくつかの袋を過ぎてカテーテルを挿入します。胎児の嚢は、挿入によって妨害されていないことを確認してください。 </li><li> Uにカテーテルを接着するテレメータ挿入部位に外科的接着剤をピペットterineホーンと子宮角の外にスライドするからカテーテルを防ぐ。剛性になるために接着剤のために数秒待ってから、慎重に体腔内子宮をバックに配置します。 </li><li>カテーテル挿入部位の反対側に体腔内テレメータを置く。 </li><li> 6.0吸収性縫合糸で閉じ体腔を縫合。まず、財布ステッチ、その後三から四個々のステッチを行う。 </li><li>同じ技術を使用して皮膚を縫合するが、6-0非吸収性縫合糸。 </li><li>スワブジブカイン（局所鎮痛剤）切開部位上に軟膏。 </li><li>動物を再水和を助けるために滅菌生理食塩水を皮下の0.3〜0.4ミリリットルを注入。 </li><li>それは完全に目を覚ましになるまで、手術後の動物を監視します。それは重く、麻酔したマウスを窒息することができ、自由に寝具、特定のタイプのように、テーブルの上にその頭を保持できるようになるまで戻って、そのケージにマウスを置かないでください。 30〜45分間麻酔をかけてきたマウスの場合、人は目を期待するべきであるEマウスは1時間と15分までに完全に目を覚ましすべき。 </li><li>追加の手術を行う場合は、70％エタノールとビーズで楽器を拭く再使用する前に滅菌する。 </li></ol> 4.手術後の回復<ol><li>ログ内の毎日の手術後の進行状況を監視します。体重増加や損失をカタログや無気力、出血、または外科的切開の不完全な閉鎖を監視。 <ol><li>マウス軟化しマウス飼料（〜アップルソースの一貫性）や特殊な回復ダイエット食品フィード。 </li><li>縫合糸を調べ、それらが引き出されていないことを確認しますが、過度にマウスを処理しないようにしてください。 </li></ol></li></ol> 5.テレメトリレコーディング<ol><li> （マウスが手術から回復した少なくとも2〜3日後）に興味のある日には、テレメーターをオンにするには、マウスの近くに磁石を配置します。レシーバをマウスケージを置きます。テレメーターが有効化されている場合、受信機上のライトがオンにする必要があります。 </li><li>真新しいtelemeのキャリブレーション<html>提供メーカー固有のキャリブレーションとTERS。再利用テレメータキャリブレーション仕様は、ソフトウェアに保持されます。 </li><li>ソフトウェアの中では、「動物」を右クリックして録音を開始し、最大500 Hzのレートでサンプリングを可能にする「連続サンプリングを開始、「クリックしてください。ソフトウェアは保存し、トレースするように設定されていることを確認します。 注：サンプリングマウスのアイコンが緑色に変わったら、圧力が記録されている。配達中にマウスを乱すことは中断したり、労働を変えるので、慎重に観察することができます。 </li><li>興味の配達や時間経過後、「動物」を右クリックして記録し、「すべて、録音を停止する」を選択することを停止する。テレメータ電池は一般的に唯一の連続記録の1.5カ月続くように、バッテリ寿命を節約するためにテレメータをオフにマグネットを使用してください。 </li><li> CO 2過剰投与によりマウスを安楽死させるとテレメータを回復。子犬は、断頭により安楽死させる。 </li></ol>TLE &quot;&gt; 6。テレメータ滅菌<ol><li>室温で脱イオン水に一晩で1％の酵素洗剤の溶液中にテレメータを置きます。 </li><li>脱イオン水でテレメータをすすぎ、カテーテルを圧迫しないように注意して、すべての組織と接着剤を取り除く。 </li><li>滅菌するために、室温で24時間、2％グルタルアルデヒドでテレメータをインキュベートする。 </li><li>滅菌生理食塩水とのテレメータをすすぎ、次の注入まで滅菌生理食塩水に保管したり改装するために戻っメーカーに送信される。組織は滅菌後のカテーテルの内側に配置されている場合テレメータを改装する必要があります。典型的には、テレメータを改装する必要が前に4~5回使用することができる。 </li></ol>

<ol>
	<li>Slattery, M. M., Morrison, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preterm+delivery.">Preterm delivery.</a> Lancet. 360, 1489-1497 (2002).</li><li>Goldenberg, R. L., Culhane, J. F., Iams, J. D., Romero, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+and+causes+of+preterm+birth.">Epidemiology and causes of preterm birth.</a> Lancet. 371, 75-84 (2008).</li><li>Keelan, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacological+inhibition+of+inflammatory+pathways+for+the+prevention+of+preterm+birth.">Pharmacological inhibition of inflammatory pathways for the prevention of preterm birth.</a> Journal Of Reproductive Immunology. 88, 176-184 (2011).</li><li>Voltolini, C., 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=Understanding+spontaneous+preterm+birth:+from+underlying+mechanisms+to+predictive+and+preventive+interventions.">Understanding spontaneous preterm birth: from underlying mechanisms to predictive and preventive interventions.</a> Reproductive Sciences. 20, 1274-1292 (2013).</li><li>Ratajczak, C. K., Muglia, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insights+into+parturition+biology+from+genetically+altered+mice.">Insights into parturition biology from genetically altered mice.</a> Pediatric research. 64, 581-589 (2008).</li><li>Rajakumar, 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=Placental+HIF-1+alpha,+HIF-2+alpha,+membrane+and+soluble+VEGF+receptor-1+proteins+are+not+increased+in+normotensive+pregnancies+complicated+by+late-onset+intrauterine+growth+restriction.">Placental HIF-1 alpha, HIF-2 alpha, membrane and soluble VEGF receptor-1 proteins are not increased in normotensive pregnancies complicated by late-onset intrauterine growth restriction.</a> American Journal Of Physiology. Regulatory, Integrative And Comparative Physiology. 293, (2007).</li><li>Whitesall, S. E., Hoff, J. B., Vollmer, A. P., D'Alecy, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+simultaneous+measurement+of+mouse+systolic+arterial+blood+pressure+by+radiotelemetry+and+tail-cuff+methods.+American+journal+of+physiology.">Comparison of simultaneous measurement of mouse systolic arterial blood pressure by radiotelemetry and tail-cuff methods. American journal of physiology.</a> Heart And Circulatory Physiology. 286, H2408-H2415 (2004).</li><li>Mackay, L. B., Shi, L. B., Maul, H. .., Maner, W. L., Garfield, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+bilateral+pelvic+neurectomy+on+cervical+ripening+in+pregnant+rats.">The effect of bilateral pelvic neurectomy on cervical ripening in pregnant rats.</a> Journal Of Perinatal Medicine. 37, 263-269 (2009).</li><li>Pierce, S. L., Kutschke, W., Cabeza, R., England, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+measurement+of+intrauterine+pressure+by+telemetry:+a+new+approach+for+studying+parturition+in+mouse+models.">In vivo measurement of intrauterine pressure by telemetry: a new approach for studying parturition in mouse models.</a> Physiol Genomics. 42, 310-316 (2010).</li><li>Imamura, T., Luedke, C. E., Vogt, S. K., Muglia, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oxytocin+modulates+the+onset+of+murine+parturition+by+competing+ovarian+and+uterine+effects.">Oxytocin modulates the onset of murine parturition by competing ovarian and uterine effects.</a> American Journal Of Physiology. Regulatory, Integrative And Comparative Physiology. 279, R1061-R1067 (2000).</li><li>McCloskey, C., 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=The+inwardly+rectifying+K++channel+KIR7.1+controls+uterine+excitability+throughout+pregnancy.">The inwardly rectifying K+ channel KIR7.1 controls uterine excitability throughout pregnancy.</a> EMBO Mol Med. 6 (9), 1161-1174 (2014).</li></ol>

The authors have nothing to disclose.

子宮筋層収縮性の調査は、筋肉の緊張のex vivoでの測定値に依存してきた。この方法論は、動物を生きるために投与することができない、新開発の子宮収縮薬の早期テストのために役立ちます。しかし、 生体内でのアプローチは縦方向に労働の進行を理解する必要がある。 in vivoでの測定は、いくつかの目的を果たす。第一に、彼らは妊娠期間にわたって子宮収縮パターン...

早産は、先進国における周産期の罹患率および死亡率の主要な原因である。それは、周産期の罹患率の50％及び周産期死亡率1,2の75％の原因である。早産は多面的で、特発ことができます。多くの研究は、収縮性の一つに、静止組織から子宮筋層を変換する分子的および電気的な経路に浮上しているが、早期陣痛の正確な病態生理学は、とらえどころのないまま。内分泌、炎症、および遺伝子調節はすべて早産3,4に関連している。しかし、倫理的な懸念は、ヒトにおける早産のメカニズムの研究を実施する能力を制限する。 これらの制限を考えると、多くの研究者が分娩の生理学を研究するためにモデル系としてマウスになっている。マウスは、約3週間持続する短い妊娠期間を有し、容易に遺伝子操作することができる。さらに、複数の遺伝的マウスモデルが持っている労働5に不可欠なシグナル伝達経路を決定するために開発されて。マウスとヒトの分娩の主な違いにもかかわらず、マウスとヒトは、子宮内炎症や感染6を含む労働のために不可欠で同じメカニズムの多くを共有しています。このように、マウスは、子宮活動を調べるための貴重なツールとして役立つ。現在までに、マウスでの子宮収縮を測定するためのゴールドスタンダードは、ex vivo等尺性張力の録音されている。しかし、これは実験ごとに1つの妊娠の時点に限定され、その生理的環境から子宮の除去を必要とする。もう一つの重要な要因は、これらの調査のために必要な多数の動物である。最後に、この方法論は、労働と早産病態生理の誘導を調べる長期的な研究のために許可していません。 動脈圧を研究するために使用される放射テレメトリデバイスで行われた最近の進歩は、マウスの7とint型に変更ラット8でrauterine圧力が同じ技術を妊娠中のマウスにおける子宮内圧力の変化を研究するために使用することができるかどうかに疑問を懇願した。初期トラブルシューティングの後、メソッドが成功したマウスでの分娩誘発および進行を測定するために開発された。このin vivoでのアプローチは、強力な陣痛示す高圧状態に静止子宮の低圧状態からの遷移を測定することができる。この方法はまた、遺伝子発現9の生理学的影響を実証する、損なわ分娩および野生型マウスとトランスジェニックマウスの間に有意な圧力差を検出することができた。このプロトコルは、妊娠中のマウス子宮圧力を研究するためのin vivo方法では、リアルタイムで提供する。

<table border="0" cellpadding="0" cellspacing="0" width="702">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Name of Material/ Equipment</td>
			<td style="width:105px;">Company</td>
			<td style="width:147px;">Catalog Number</td>
			<td style="width:207px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Surgical Glue</td>
			<td style="width:105px;">3M Vetbond</td>
			<td style="width:147px;">1469SB</td>
			<td>Tissue Adhesive--stored in Drierite</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Absorbable 6-0 Sutures</td>
			<td style="width:105px;">Ethicon</td>
			<td style="width:147px;">J212H</td>
			<td>Vicryl</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Non-absorbable 6-0 Sutures</td>
			<td style="width:105px;">Ethicon</td>
			<td style="width:147px;">8706H</td>
			<td>Prolene</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;">Water Jacket Blanket + Heating Pad</td>
			<td style="width:105px;">Gaymer</td>
			<td style="width:147px;">T/Pump PN 11184-000 Blanket-66N111CC</td>
			<td>Specialized for rodents</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Bead sterilizer</td>
			<td style="width:105px;">Keller</td>
			<td style="width:147px;">Z378577</td>
			<td>Steri 250 Sterilizer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Disecting Microscope</td>
			<td style="width:105px;">Nikon</td>
			<td style="width:147px;">SMZ754</td>
			<td>Fibre optic gooseneck external light source</td>
		</tr>
		<tr height="46">
			<td height="46" style="height:46px;width:245px;">Sterile Surgical Gloves--Latex&#160;</td>
			<td style="width:105px;">Cardinal Health Triflex</td>
			<td style="width:147px;">2D7253</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:245px;">PhysioTel PA-C10 Pressure Transmitter</td>
			<td style="width:105px;">&#160;Data Sciences International</td>
			<td style="width:147px;">270-0135-001</td>
			<td style="width:207px;">TA11PA-C10</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:245px;">Telemeter Reciever</td>
			<td style="width:105px;">&#160;Data Sciences International</td>
			<td style="width:147px;">272-6001-001</td>
			<td style="width:207px;">RPC-1</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:245px;">Dataquest ART 4.3.2 Analysis Platinum</td>
			<td style="width:105px;">&#160;Data Sciences International</td>
			<td style="width:147px;">271-0147-141</td>
			<td>Analysis software</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Diet Recovery Gel</td>
			<td style="width:105px;">Clear H2O</td>
			<td style="width:147px;">72-01-5022</td>
			<td>Purified Soft Diet for Rodents</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:245px;">Tergazyme</td>
			<td style="width:105px;">Sigma-Aldrich</td>
			<td>Z273287</td>
			<td>Enzyme detergent</td>
		</tr>
	</tbody>
</table>

intrauterine telemetry to measure mouse contractile pressure in vivo

分子の電気信号の複雑な統合は、妊娠の終わりに収縮器官に静止子宮を変換するために必要とされる。子宮収縮の重要な調節因子の発見にもかかわらず、このプロセスはまだ十分に理解されていない。トランスジェニックマウスは、分娩を研究するための理想的なモデルを提供する。以前は、マウスに子宮収縮を研究する唯一の方法は、いくつかの理由のために最適ではex vivoでの等尺性張力の録音だった。子宮は、その生理的環境から除去されなければならない、調査の限られた時間的経過が可能であり、マウスを犠牲にしなければならない。放射遠隔測定の最近の開発は、マウスにおけるインビボ子宮内圧の長手方向の、リアルタイム測定を可能にした。送達が記載されるまでここで、子宮内テレメータの注入は、妊娠中期のマウスの子宮内の圧力変化を測定する。プレスの違いを比較することにより野生型およびトランスジェニックマウスとの間の小節がズレ、目的の遺伝子の生理​​的影響を明らかにすることができる。この技術は、早期陣痛を含む妊娠中に子宮筋障害を治療するために使用される治療薬の開発を促進すべきである。

インビボ子宮内圧力のリアルタイムの遠隔収集システムを用いて記録することができる。マウスの同時録画と対に圧力をサンプリングし、各子犬の配達の正確な時間をキャプチャし、母親の子宮内圧力に配達時間を相関させるために使用された。すべてのデータポイントは、圧力対時間プロット（ 図1A）を生成するために記録全体にプロットすることができる。こ?...

Watch this Scientific Journal Video about Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo at JoVE.com

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

This manuscript provides a protocol for implanting telemeters in the mouse for the purpose of measuring intrauterine pressures during pregnancy.

マウス収縮圧力を測定するために子宮内テレメトリ<em&gt;インビボ</em

A complex integration of molecular and electrical signals is needed to transform a quiescent uterus into a contractile organ at the end of pregnancy. ...

intrauterine-telemetry-to-measure-mouse-contractile-pressure-in-vivo

Research

JoVE Journal

Medicine

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

9.6K ビュー.  Washington University in St. Louis. This manuscript provides a protocol for implanting telemeters in the mouse for the purpose of measuring intrauterine pressures during pregnancy. 

ビデオ: Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

Pressure Controlled Ventilation to Induce Acute Lung Injury in Mice

Murine models are extensively used to investigate acute injuries of different organs systems (1-34). Acute lung injury (ALI), which occurs with prolonged mechanical ventilation, contributes to morbidity and mortality of critical illness, and studies on novel genetic or pharmacological targets are areas of intense investigation (1-3, 5, 8, 26, 30, 33-36). ALI is defined by the acute onset of the disease, which leads to non-cardiac pulmonary edema and subsequent impairment of pulmonary gas exchange (36). We have developed a murine model of ALI by using a pressure-controlled ventilation to induce ventilator-induced lung injury (2). For this purpose, C57BL/6 mice are anesthetized and a tracheotomy is performed followed by induction of ALI via mechanical ventilation. Mice are ventilated in a pressure-controlled setting with an inspiratory peak pressure of 45 mbar over 1 - 3 hours. As outcome parameters, pulmonary edema (wet-to-dry ratio), bronchoalveolar fluid albumin content, bronchoalveolar fluid and pulmonary tissue myeloperoxidase content and pulmonary gas exchange are assessed (2). Using this technique we could show that it sufficiently induces acute lung inflammation and can distinguish between different treatment groups or genotypes (1-3, 5). Therefore this technique may be helpful for researchers who pursue molecular mechanisms involved in ALI using a genetic approach in mice with gene-targeted deletion.

A murine model for ventilator induced lung injury is an important tool to study an acute lung injury in vivo. Here, we report an easy applicable in situ model for acute lung injury using high-pressure mechanical ventilation to induce acute failure of the lung.

マウスにおける急性肺障害を誘発するために圧力制御された換気

Murine models are extensively used to investigate acute injuries of different organs systems (1-34). Acute lung injury (ALI), which occurs with prolonged ...

医学

Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ultrasound and fetal echocardiography. Increasingly, these imaging modalities are being employed clinically to examine cardiac function and assess wellbeing in utero, thereby guiding timing of birth decisions. Here, we used a rabbit model of IUGR that allows analysis of cardiac function in a clinically relevant way. Using isoflurane induced anesthesia, IUGR is surgically created at gestational age day 25 by performing a laparotomy, exposing the bicornuate uterus and then ligating 40-50% of uteroplacental vessels supplying each gestational sac in a single uterine horn. The other horn in the rabbit bicornuate uterus serves as internal control fetuses. Then, after recovery at gestational age day 30 (full term), the same rabbit undergoes examination of fetal cardiac function. Anesthesia is induced with ketamine and xylazine intramuscularly, then maintained by a continuous intravenous infusion of ketamine and xylazine to minimize iatrogenic effects on fetal cardiac function. A repeat laparotomy is performed to expose each gestational sac and a microultrasound examination (VisualSonics VEVO 2100) of fetal cardiac function is performed. Placental insufficiency is evident by a raised pulsatility index or an absent or reversed end diastolic flow of the umbilical artery Doppler waveform. The ductus venosus and middle cerebral artery Doppler is then examined. Fetal echocardiography is performed by recording B mode, M mode and flow velocity waveforms in lateral and apical views. Offline calculations determine standard M-mode cardiac variables, tricuspid and mitral annular plane systolic excursion, speckle tracking and strain analysis, modified myocardial performance index and vascular flow velocity waveforms of interest. This small animal model of IUGR therefore affords examination of in utero cardiac function that is consistent with current clinical practice and is therefore useful in a translational research setting.

We describe examination of fetal cardiac function with contemporary functional fetal echocardiography and fetoplacental Doppler ultrasound using the VisualSonics VEVO 2100 microultrasound in a surgically induced model of intrauterine fetal growth restriction in a rabbit.

胎児心エコー検査と子宮内発育制限のウサギモデルにおけるパルス波ドップラー超音波

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ...

In vivo 19F MRI for Cell Tracking

In vivo 19F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. Here, we describe a general protocol for cell labeling, imaging, and image processing. The technique is applicable to various cell types and animal models, although here we focus on a typical mouse model for tracking murine immune cells. The most important issues for cell labeling are described, as these are relevant to all models. Similarly, key imaging parameters are listed, although the details will vary depending on the MRI system and the individual setup. Finally, we include an image processing protocol for quantification. Variations for this, and other parts of the protocol, are assessed in the Discussion section. Based on the detailed procedure described here, the user will need to adapt the protocol for each specific cell type, cell label, animal model, and imaging setup. Note that the protocol can also be adapted for human use, as long as clinical restrictions are met.

In vivo 19F MRI for Cell Tracking

We describe a general protocol for in vivo cell tracking using MRI in a mouse model with ex vivo labeled cells. A typical protocol for cell labeling, image acquisition processing and quantification is included.

細胞追跡のためのin vivoでの19 F MRI における

In vivo 19F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. ...

Accuracy in Dental Medicine, A New Way to Measure Trueness and Precision

Reference scanners are used in dental medicine to verify a lot of procedures. The main interest is to verify impression methods as they serve as a base for dental restorations. The current limitation of many reference scanners is the lack of accuracy scanning large objects like full dental arches, or the limited possibility to assess detailed tooth surfaces. A new reference scanner, based on focus variation scanning technique, was evaluated with regards to highest local and general accuracy. A specific scanning protocol was tested to scan original tooth surface from dental impressions. Also, different model materials were verified. The results showed a high scanning accuracy of the reference scanner with a mean deviation of 5.3 &#177; 1.1 &#181;m for trueness and 1.6 &#177; 0.6 &#181;m for precision in case of full arch scans. Current dental impression methods showed much higher deviations (trueness: 20.4 &#177; 2.2 &#181;m, precision: 12.5 &#177; 2.5 &#181;m) than the internal scanning accuracy of the reference scanner. Smaller objects like single tooth surface can be scanned with an even higher accuracy, enabling the system to assess erosive and abrasive tooth surface loss. The reference scanner can be used to measure differences for a lot of dental research fields. The different magnification levels combined with a high local and general accuracy can be used to assess changes of single teeth or restorations up to full arch changes.

Accuracy is a major demand in dental medicine. To verify accuracy, reference scanners are needed. This article presents a new reference scanner with an adjusted scanning method to acquire a broad variety of dental morphologies with high trueness and precision.

歯科医学、真度と精度を測定するための新しい方法の精度

Reference scanners are used in dental medicine to verify a lot of procedures. The main interest is to verify impression methods as they serve as a base ...

Measurement of the Pressure-volume Curve in Mouse Lungs

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential phenotypic changes are best assessed by measurement of the changes in lung elasticity. To best understand specific mechanisms underlying such pathologies in mice, it is essential to make functional measurements that can reflect the developing pathology. Although there are many ways to measure elasticity, the classical method is that of the total lung pressure-volume (PV) curve done over the whole range of lung volumes. This measurement has been made on adult lungs from nearly all mammalian species dating back almost 100 years, and such PV curves also played a major role in the discovery and understanding of the function of pulmonary surfactant in fetal lung development. Unfortunately, such total PV curves have not been widely reported in the mouse, despite the fact that they can provide useful information on the macroscopic effects of structural changes in the lung. Although partial PV curves measuring just the changes in lung volume are sometimes reported, without a measure of absolute volume, the nonlinear nature of the total PV curve makes these partial ones very difficult to interpret. In the present study, we describe a standardized way to measure the total PV curve. We have then tested the ability of these curves to detect changes in mouse lung structure in two common lung pathologies, emphysema and fibrosis. Results showed significant changes in several variables consistent with expected structural changes with these pathologies. This measurement of the lung PV curve in mice thus provides a straightforward means to monitor the progression of the pathophysiologic changes over time and the potential effect of therapeutic procedures.

Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung&#8217;s structural changes with developing pathology.

マウス肺における圧力 - 容積曲線の測定

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential ...

Trabecular Meshwork Response to Pressure Elevation in the Living Human Eye

The mechanical characteristics of the trabecular meshwork (TM) are linked to outflow resistance and intraocular pressure (IOP) regulation. The rationale behind this technique is the direct observation of the mechanical response of the TM to acute IOP elevation. Prior to scanning, IOP is measured at baseline and during IOP elevation. The limbus is scanned by spectral-domain optical coherence tomography at baseline and during IOP elevation (ophthalmodynamometer (ODM) applied at 30 g force). Scans are processed to enhance visualization of the aqueous humor outflow pathway using ImageJ. Vascular landmarks are used to identify corresponding locations in baseline and IOP elevation scan volumes. Schlemm canal (SC) cross-sectional area (SC-CSA) and SC length from anterior to posterior along its long axis are measured manually at 10 locations within a 1 mm segment of SC. Mean inner to outer wall distance (short axis length) is calculated as the area of SC divided by its long axis length. To examine the contribution of adjacent tissues to the effect IOP elevations, measurements are repeated without and with smooth muscle relaxation with instillation of tropicamide. TM migration into SC is resisted by TM stiffness, but is enhanced by the support of its attachment to adjacent smooth muscle within the ciliary body. This technique is the first to measure the living human TM response to pressure elevation in situ under physiological conditions within the human eye.

Trabecular meshwork (TM) migration into Schlemm&#8217;s canal space can be induced by acute pressure elevation by ophthalmodynamometer, and observed by spectral domain optical coherence tomography. The goal of this method is to quantify the morphometric response of the living outflow tract to acute pressure elevation in living tissues in situ.

生きている人間の目で圧力上昇を小柱網の対応

The mechanical characteristics of the trabecular meshwork (TM) are linked to outflow resistance and intraocular pressure (IOP) regulation. The rationale ...

Cardiac Catheterization in Mice to Measure the Pressure Volume Relationship: Investigating the Bowditch Effect

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these strategies is to examine their effects on heart function. There are several techniques to measure in vivo cardiac mechanics (e.g., echocardiography, pressure/volume relations, etc.). Compared to echocardiography, real-time left ventricular (LV) pressure/volume analysis via catheterization is more precise and insightful in assessing LV function. Additionally, LV pressure/volume analysis provides the ability to instantaneously record changes during manipulations of contractility (e.g., &#946;-adrenergic stimulation) and pathological insults (e.g., ischemia/reperfusion injury). In addition to the maximum (+dP/dt) and minimum (-dP/dt) rate of pressure change in the LV, an accurate assessment of LV function via several load-independent indexes (e.g., end systolic pressure volume relationship and preload recruitable stroke work) can be attained. Heart rate has a significant effect on LV contractility such that an increase in the heart rate is the primary mechanism to increase cardiac output (i.e., Bowditch effect). Thus, when comparing hemodynamics between experimental groups, it is necessary to have similar heart rates. Furthermore, a hallmark of many cardiomyopathy models is a decrease in contractile reserve (i.e., decreased Bowditch effect). Consequently, vital information can be obtained by determining the effects of increasing heart rate on contractility. Our and others data has demonstrated that the neuronal nitric oxide synthase (NOS1) knockout mouse has decreased contractility. Here we describe the procedure of measuring LV pressure/volume with increasing heart rates using the NOS1 knockout mouse model.

This article describes the measurement of murine left ventricular function via pressure/volume analysis at different heart rates.

ボーディッチ影響を調べる：圧力体積関係を測定するために、マウスにおける心臓カテーテル検査

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these ...

Development of an Algorithm to Perform a Comprehensive Study of Autonomic Dysreflexia in Animals with High Spinal Cord Injury Using a Telemetry Device

Spinal cord injury (SCI) is a debilitating neurological condition characterized by somatic and autonomic dysfunctions. In particular, SCI above the mid-thoracic level can lead to a potentially life-threatening hypertensive condition called autonomic dysreflexia (AD) that is often triggered by noxious or non-noxious somatic or visceral stimuli below the level of injury. One of the most common triggers of AD is the distension of pelvic viscera, such as during bladder and bowel distension or evacuation. This protocol presents a novel pattern recognition algorithm developed for a JAVA platform software to study the fluctuations of cardiovascular parameters as well as the number, severity and duration of spontaneously occurring AD events. The software is able to apply a pattern recognition algorithm on hemodynamic data such as systolic blood pressure (SBP) and heart rate (HR) extracted from telemetry recordings of conscious and unrestrained animals before and after thoracic (T3) complete transection. With this software, hemodynamic parameters and episodes of AD are able to be detected and analyzed with minimal experimenter bias.

The catheter of a telemetry device is implanted into the abdominal aorta in order to continuously collect beat-by-beat hemodynamic data from animals pre and post-high thoracic spinal cord transection. A novel JAVA software was employed to analyze hemodynamic parameters as well as frequency and intensity of spontaneous episodes of autonomic dysreflexia.

遠隔測定装置を使用した高脊髄損傷で動物に自律神経反射異常の総合的研究を行うためにアルゴリズムの開発

Spinal cord injury (SCI) is a debilitating neurological condition characterized by somatic and autonomic dysfunctions. In particular, SCI above the ...

Long-term Blood Pressure Measurement in Freely Moving Mice Using Telemetry

During the development of new vasoactive agents, arterial blood pressure monitoring is crucial for evaluating the efficacy of the new proposed drugs. Indeed, research focusing on the discovery of new potential therapeutic targets using genetically altered mice requires a reliable, long-term assessment of the systemic arterial pressure variation. Currently, the gold standard for obtaining long-term measurements of blood pressure in ambulatory mice uses implantable radio-transmitters, which require&#160;artery cannulation. This technique eliminates the need for tethering, restraining, or anesthetizing the animals which introduce stress and artifacts during data sampling. However, arterial blood pressure monitoring in mice via catheterization can be rather challenging due to the small size of the arteries. Here we present a step-by-step guide to illustrate the crucial key passages for a successful subcutaneous implantation of radio-transmitters and carotid artery cannulation in mice. We also include examples of long-term blood pressure activity taken from freely moving mice after a period of post-surgery recovery. Following this procedure will allow reliable direct blood pressure recordings from multiple animals simultaneously.

The goal of this protocol is to assess systemic blood pressure in conscious freely moving mice using implantable radio-telemetry devices.

テレメトリを使用して自由に動くマウスにおける長期血圧測定

During the development of new vasoactive agents, arterial blood pressure monitoring is crucial for evaluating the efficacy of the new proposed drugs. ...

Depletion of Mouse Cells from Human Tumor Xenografts Significantly Improves Downstream Analysis of Target Cells

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic patient tumors in different assays1. Human tumor xenografts represent the gold standard with respect to tumor biology, drug discovery, and metastasis research 2-4. Tumor xenografts can be derived from different types of material like tumor cell lines, tumor tissue from primary patient tumors4 or serially transplanted tumors. When propagated in vivo, xenografted tissue is infiltrated and vascularized by cells of mouse origin. Multiple factors such as the tumor entity, the origin of xenografted material, growth rate and region of transplantation influence the composition and the amount of mouse cells present in tumor xenografts. However, even when these factors are kept constant, the degree of mouse cell contamination is highly variable.
Contaminating mouse cells significantly impair downstream analyses of human tumor xenografts. As mouse fibroblasts show high plating efficacies and proliferation rates, they tend to overgrow cultures of human tumor cells, especially slowly proliferating subpopulations. Mouse cell derived DNA, mRNA, and protein components can bias downstream gene expression analysis, next-generation sequencing, as well as proteome analysis 5. To overcome these limitations, we have developed a fast and easy method to isolate untouched human tumor cells from xenografted tumor tissue. This procedure is based on the comprehensive depletion of cells of mouse origin by combining automated tissue dissociation with the benchtop tissue dissociator and magnetic cell sorting. Here, we demonstrate that human target cells can be can be obtained with purities higher than 96% within less than 20 min independent of the tumor type.

Human tumor xenografts are vascularized and infiltrated by cells of mouse origin during the growth phase in vivo. To circumvent the bias caused by these contaminating cells during downstream analysis, we developed a method allowing for comprehensive depletion of all mouse cells by magnetic cell sorting.

ヒト腫瘍異種移植片からマウス細胞の枯渇は大幅に標的細胞の下流分析を向上させます

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic ...