この映像公開はDK 68055プログラム·プロジェクト無償 &quot;腸内系の病理&quot;のための糖尿病および消化器腎臓病研究所（NIDDK）からの資金によって可能となった。クリストファーCreedonはロチェスター公立学校の指導プログラムによってサポートされていました。 我々は、事務補助やロスガトスリサーチ社（カリフォルニア州マウンテンビュー）から博士ダグラスベーアのために氏クリスティZodrow、技術支援のための氏ゲイリー·ストルツに感謝します。

1。マウスのトレーニングと馴化<ol><li>分析に先立ち、試験条件にそれらを慣らすために、一定のエアフローと2から4時間の試験室ですべてのマウスを置く。これが顕著にそうでなければ胃排出遅延の異常検出が発生する可能性があり、ストレスレベルを低減します。マウスの胃排出実験が実行されていたかのように同じように扱う。オクタン酸を追加することなく、卵黄を（後述）を準備し、各マウスに0.2グラムを養う。 </li><li>マウスは、十分に訓練されているまで（通常は2〜3倍）、このプロセスを繰り返します。環境条件が同じに保たれているようにしたマウスは、通常、簡単に限り、慣らされています。 </li></ol> 注：非慣れたマウスは慣れたマウスは素早く静かに彼らの新しい環境と休息に定住しながら、チャンバーへの転送後、約1時間の周りに移動して、排便すると頻繁に排尿し続けています。 ノーTE：実験中：そのような過剰な排尿、排便、卵を食べることへの関心の欠如など、慣れの損失の兆候がモニター動物。このような場合は再度前胃排出データを取得するには空室に1-2回habituatingを検討してください。この実験をしている間に一貫性が極めて重要である。物事を正確に同じ方法を行うと、毎回は、信頼性と再現性のある結果を得るための唯一の方法です。これは彼らのケージメイトからマウスを分離しない、毎日同じ時刻に（ 例えば、インスリン）絶対に必要でない場合、マウスを絶食すると同時に、胃排出試験を開始し、マウスを扱うのと同じ方法で治療を与えることが含まれています。 2。試験食を含む同位体の調製<ol><li> 50mlファルコンチューブに卵黄の5グラムを量り始めてください。新鮮な試験食を準備するために、各実験当日、これらの手順を繰り返します。 </li><li> concentrとオクタン酸10μlを追加ファルコンチューブにスパチュラで1分間激しく卵とミックスを含む50mlファルコンチューブに2μlの/ gのation。 </li><li>卵をガラスビーカーに移し、それが凝固し、その一貫性は小さいボールを作るのに適しているまで、ブンゼンバーナーで加熱されています。これは、典型的には約30秒かかります。 </li></ol> 注：卵黄のボールはマウス1匹当たり0.2グラムの重量を量るべきである。これは、すべてのマウスでは累積投与量を一定に保つことが重要です。 3。実験の開始<ol><li>食糞を防止するために、金属の &quot;メッシュ底&quot;断食ラック上（12時間）速いマウスは一晩、一回訓練を受けたと胃排出の準備が整いました。彼らは飲料水への無料アクセスを提供していることを確認してください。糖尿病マウスは、現在の実験で使用されているので、それらは以上16時間絶食させてはいけません。 </li><li>胃が空室を設定することで起動します。空気乾燥してきたクリーンチャンバーとカバーを使用しています。また、任意のアナライザまたはCO 2空気供給源にチャンバーを接続するチューブが水分を含まなければなりません。水が分析装置により読み出された信号に干渉する可能性が</li><li>一定の空気の流れを提供するインレットチューブにチャンバーを接続します。その後マシンにチャンバーから出口管を接続してください。チューブを閉じて空気の流れをオンにします。 </li></ol> 注：彼らは簡単に閉じ、しっかりと密封されるようにカバーの蓋の端にワセリンの非常に少量を適用します。このタイトなシールは、マウスによって生成されたすべての二酸化炭素を回収することが必要である。 4。実験手順<ol><li>各マウスの重量を量ることから始めます。体重はその継続的健康の指標です。次に、適切なチャンバー内に各マウスを置きます。それは空気が、この時点でチャンバーに流入していることはもちろん重要です。 </li><li>測定を開始するには、マウスが調整前室に順応させて空気レベルをING。 </li><li>マウスは数分かかることがあり、落ち着いて見えると、各マウス室用空気流量を調整します。これは、それぞれのマウスとは異なる場合があります。典型的には、空気の流れがCO 2が使用されているどのような機器で検出可能なレベルに達すると、レベルは健康的な空気の離職率を確保するのに十分に低いままであることを確認するために呼気ことを確認するために、実験開始時に調整されます。我々は、万人あたり1,000〜1,500部品との間の最初のCO 2レベルを使用します。 </li><li>調整が困難な場合は、エア漏れをチェックします。その後、チャンバのそれぞれの処理を繰り返しますし、通気を行った調整は、CO 2レベルを修正したかどうかを確認するために測定の別のラウンドを監視します。それは、マウスを供給する前に読んで着実にベースラインを得ることが重要である。我々は、自己校正機能を備えたマシンを使用しています。これがない場合はケース·キャリブレーションをチェックする必要があります。 </li><li>これが達成されると、卵の食事の管理最初にマウスと記録時間に各マウスは食べ物を受け取ります。 </li><li>我々は、各マウスの13 CO 2の濃縮曲線をフィッティングのために十分な値を得るために、4時間の手順を実行します。マウスでCO 2濃度は、マウスにはまだ安全であることを確認するために、すべての30〜60分を確認してください。 </li><li>テストが終わった後のマウスはすぐに食べ始めることができるように、テストの終了前に食品を含む新しい箱を用意します。 </li></ol>

<ol>
	<li>Yeung, C. K., McCurrie, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+method+to+investigate+the+inhibitory+effects+of+drugs+on+gastric+emptying+in+the+mouse+in+vivo.">A simple method to investigate the inhibitory effects of drugs on gastric emptying in the mouse in vivo.</a> J. Pharmacol. Toxicol. Methods. 45, 235-240 (2001).</li><li>Osinski, M. A., Seifert, T. R., Cox, B. F., Gintant, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+improved+method+of+evaluation+of+drug-evoked+changes+in+gastric+emptying+in+mice.">An improved method of evaluation of drug-evoked changes in gastric emptying in mice.</a> J. Pharmacol. Toxicol. Methods. 47, 115-120 (2002).</li><li>Ghoos, Y. F., 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=Measurement+of+gastric+emptying+rate+of+solids+by+means+of+a+carbon-labeled+octanoic+acid+breath+test.">Measurement of gastric emptying rate of solids by means of a carbon-labeled octanoic acid breath test.</a> Gastroenterology. 104, 1640-1647 (1993).</li><li>Symonds, E., Butler, R., Omari, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+breath+tests+can+detect+alterations+in+gastric+emptying+in+the+mouse.">Noninvasive breath tests can detect alterations in gastric emptying in the mouse.</a> Eur. J. Clin. Invest. 32, 341-344 (2002).</li><li>Symonds, E. L., Butler, R. N., Omari, T. I. <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+gastric+emptying+in+the+mouse+using+the+[13C]-octanoic+acid+breath+test.">Assessment of gastric emptying in the mouse using the [13C]-octanoic acid breath test.</a> Clin. Exp. Pharmacol. Physiol. 27, 671-675 (2000).</li><li>Verhulst, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+ghrelin+in+the+relationship+between+hyperphagia+and+accelerated+gastric+emptying+in+diabetic+mice.">Role of ghrelin in the relationship between hyperphagia and accelerated gastric emptying in diabetic mice.</a> Gastroenterology. 135, 1267-1276 (2008).</li><li>Schoonjans, R., 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+13C-octanoic+acid+breath+test:+validation+of+a+new+noninvasive+method+of+measuring+gastric+emptying+in+rats.">The 13C-octanoic acid breath test: validation of a new noninvasive method of measuring gastric emptying in rats.</a> Neurogastroenterol. Motil. 14, 287-293 (2002).</li><li>Choi, K. M., 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=Determination+of+gastric+emptying+in+nonobese+diabetic+mice.">Determination of gastric emptying in nonobese diabetic mice.</a> Am. J. Physiol. Gastrointest. Liver Physiol. 293, 1039-1045 (2007).</li><li>Atkinson, M. A., Leiter, E. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+NOD+mouse+model+of+type+1+diabetes:+as+good+as+it+gets?.">The NOD mouse model of type 1 diabetes: as good as it gets?.</a> Nat. Med. 5, 601-604 (1999).</li><li>Camilleri, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+practice.+Diabetic+gastroparesis.">Clinical practice. Diabetic gastroparesis.</a> N. Engl. J. Med. 356, 820-829 (2007).</li><li>Choi, K. M., 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=Heme+oxygenase-1+protects+interstitial+cells+of+Cajal+from+oxidative+stress+and+reverses+diabetic+gastroparesis.">Heme oxygenase-1 protects interstitial cells of Cajal from oxidative stress and reverses diabetic gastroparesis.</a> Gastroenterology. 135, 2055-2064 (2008).</li><li>Kashyap, P. 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=Carbon+monoxide+reverses+diabetic+gastroparesis+in+NOD+mice.">Carbon monoxide reverses diabetic gastroparesis in NOD mice.</a> Am. J. Physiol. GI. G298, G1013-G1019 (2010).</li></ol>

著者らは、彼らが競合する経済的利益を持っていないことを宣言します。

記載された技術は、本明細書で、マウスにおける固体胃排出能の in vivo測定で繰り返して、非侵襲することができます。このシステムでは、動物は、それらを移動して自由に回転できるように、測定室に拘束されないという利点があります。これは不慣れな環境であるため、マウスはまだ訓練され、胃内容排出へのストレスの影響を防ぐために試験室に慣れなければならない。?...

本稿では、マウスにおける胃排出能の非侵襲的測定にかかわる技術的および方法論的な考慮事項について説明します。ここに記載されたプロトコールに従うことによって、研究者は確実にかつ再現性胃排出に関する薬理学的薬剤の影響を研究し、基礎疾患や欠陥6,8の治療への胃内容排出の応答に従って、疾患の発症に起因する空胃の変化を追うことができ、 11,12。以前の出版物では、13 Cオクタン酸呼気検査の適用は、ヒトおよび動物3,8で胃排出を測定するのに便利な方法であることが示された。本稿では詳細に説明し、6から8を介して信頼性のあるデータを得るために必要な手続き糖尿病マウスにおける胃排出能の縦断的研究のために必要な数ヶ月。以前に公開されている方法と比較した場合、このプロトコルに従うことの利点は、治験責任医師がデータobtaineを保証することができるということですdは、信頼性と再現性になります。また、ここで記載されたガスのサンプルを収集し、分析するための自​​動化されたシステムは、研究で同時に追跡することができる動物の数が増加します。全体的に、この論文の目的は、テストに対するマウスの馴化を維持し、それが得られた結果のばらつきを軽減する重要な要因を特定することです。 胃排出能の in vivoでの測定では、マウスを一晩絶食させると一定のエアフローを透明なプラスチックの試験室に入れた。マウスをチューブに慣らさ取得した後、ベースラインは、13 CO 2濃度が決定され、気流がそれに応じて調整され吐き出さ。次に、我々は、13 C標識オクタン酸と混合卵黄から成る試験食を投与する。マウスを絶食と訓練されているので、それらは一般的には2分以内に試験食を食べます。投与オクタン酸は胃で吸収されませんが、十二指腸で取り上げられることになるdは解放され、周囲の空気中の13 CO 2の濃縮で、その結果、吐き出され、13 CO 2、に肝臓で代謝されてしまいます。大気試料は決められた時間間隔で収集され、炭素同位体分析器によって分析される。このプロセス全体の律速段階は、胃排出能であり、13 CO 2の肺の排泄は、直接標識食事の胃排出能に対応しています。 <img src="/files/ftp_upload/50301/50301fig1.jpg" alt="図1" fo:content-width="6in" fo:src="/files/ftp_upload/50301/50301fig1highres.jpg"/> 図1胃排出装置の模式図。一晩絶食後、マウスを移動すると、それらを自由に回転できるように透明なチャンバー内に配置されます。入口管は、新鮮で一定の空気の流入を可能にし、コンセントは13のC-to-12を測定するために同位体分析装置につながる呼気中のC比。チャンバは[13 C] -オクタン酸を含む食品の配達のための中央のポートを持っています。

<table border="1"><tbody><tr><td> NAME </td><td> 会社 </td><td> CATのNUMBER </td><td> コメント </td></tr><tr><td>一定の空気供給、流れの源ならびに組成</td><td>研究施設の中心的な空気供給</td><td></td><td></td></tr><tr><td>エアインレット、エアアウトレット、食品管理の開口部を有し、130ミリリットルのサンプリング室 空気供給用のプラスチックチューブ</td><td>社内に構築</td><td></td><td></td></tr><tr><td>オクタン酸</td><td>ケンブリッジアイソトープラボラトリーズ（アンドーバー、マサチューセッツ州） </td><td> CLM-293から1 </td><td></td></tr><tr><td>卵の食事を準備するには： <ul><li>小さなビーカー</li><li> 50mlのプラスチックチューブ</li><li>ブンゼンバーナー</li><li>卵</li><li>へら</li></ul></td><td>すべてのサプライヤ</td><td></td><td>してみてください卵の栄養成分とおいしさは、食事の摂取と胃排出能に影響を与えることができるので、卵サプライヤーとの整合性</td></tr><tr><td>二酸化炭素同位体分析器</td><td>ロスガトスリサーチ社（カリフォルニア州マウンテンビュー） </td><td></td><td></td></tr></tbody></table>

assessment of gastric emptying in non-obese diabetic mice using a [13c]-octanoic acid breath test

マウスにおける胃排出試験は、最も一般的に使用される技術は、動物との食事を1,2の死後回復を殺す必要とするので、同じ動物における胃排出変化に追従することができないことによって制限されています。このアプローチでは、年齢や疾患の進行と胃排出能の変化を判断するのは長期的な研究を防ぎます。人間3に一般的に使用される[13 C] -オクタン酸呼気試験は、マウス4-6およびラット7で使用するために変更され、我々は以前に、このテストは薬剤への応答との間に胃排出能の変化に対する信頼性と応答性であることを示したされています糖尿病疾患の進行8。このビデオプレゼンテーションでは、この変更されたテストの原理と実用的な実装について説明する。以前の研究と同様に、NOD LTJマウスは、1型糖尿病9のモデルを使用されています。これらのマウスの割合はdiabetの合併症、胃不全麻痺の症状を起こすES胃10の機械的閉塞せずに遅延胃排出が特徴。 本論文では、試験食を準備し、4時間胃排出データと方法で得られたデータを分析する方法を取得する方法、テストのためにマウスを訓練する方法を示しています。本研究で用いた炭素同位体分析器は、同時に12匹までから空気サンプルの自動サンプリングに適しています。この技術は、糖尿病やその他の長期にわたる疾患をもつマウスの大きいグループから胃排出の長手方向のフォローアップができます。

三つの異なるマウスから設定の代表的なデータを図2に示します。黒いグラフは通常の胃排出能を持つマウスからのデータポイントを表します。それは、時間の関数として表される時間当たり投与量の割合として表される呼気に回収される13 Cの割合を示しています。青い曲線は、40分のT半値で加速胃排出能を持つマウスからのものであり、赤い曲線は、168分のT半値で空に...

Watch this Scientific Journal Video about Assessment of Gastric Emptying in Non-obese Diabetic Mice Using a [13C]-octanoic Acid Breath Test at JoVE.com

Assessment of Gastric Emptying in Non-obese Diabetic Mice Using a [13C]-octanoic Acid Breath Test

非侵襲による胃排出の決定[<sup&gt; 13</sup&gt;女性のNOD LTJマウスで胃を追跡するためのC] - オクタン酸呼気検査。

[を使用した非肥満糖尿病マウスにおける胃排出能の評価<sup&gt; 13</sup&gt; C] - オクタン酸呼気テスト

Gastric emptying studies in mice have been limited by the inability to follow gastric emptying changes in the same animal since the most commonly used ...

assessment-gastric-emptying-non-obese-diabetic-mice-using-13c

Research

JoVE Journal

Medicine

Assessment of Gastric Emptying in Non-obese Diabetic Mice Using a [13C]-octanoic Acid Breath Test

13.5K ビュー.  Mayo Clinic. 非侵襲による胃排出の決定[ 13女性のNOD LTJマウスで胃を追跡するためのC] - オクタン酸呼気検査。

ビデオ: Assessment of Gastric Emptying in Non-obese Diabetic Mice Using a [13C]-octanoic Acid Breath Test

Making Sense of Listening: The IMAP Test Battery

The ability to hear is only the first step towards making sense of the range of information contained in an auditory signal. Of equal importance are the abilities to extract and use the information encoded in the auditory signal. We refer to these as listening skills (or auditory processing AP). Deficits in these skills are associated with delayed language and literacy development, though the nature of the relevant deficits and their causal connection with these delays is hotly debated.
When a child is referred to a health professional with normal hearing and unexplained difficulties in listening, or associated delays in language or literacy development, they should ideally be assessed with a combination of psychoacoustic (AP) tests, suitable for children and for use in a clinic, together with cognitive tests to measure attention, working memory, IQ, and language skills. Such a detailed examination needs to be relatively short and within the technical capability of any suitably qualified professional. Current tests for the presence of AP deficits tend to be poorly constructed and inadequately validated within the normal population. They have little or no reference to the presenting symptoms of the child, and typically include a linguistic component. Poor performance may thus reflect problems with language rather than with AP. To assist in the assessment of children with listening difficulties, pediatric audiologists need a single, standardized child-appropriate test battery based on the use of language-free stimuli.
We present the IMAP test battery which was developed at the MRC Institute of Hearing Research to supplement tests currently used to investigate cases of suspected AP deficits. IMAP assesses a range of relevant auditory and cognitive skills and takes about one hour to complete. It has been standardized in 1500 normally-hearing children from across the UK, aged 6-11 years. Since its development, it has been successfully used in a number of large scale studies both in the UK and the USA. IMAP provides measures for separating out sensory from cognitive contributions to hearing. It further limits confounds due to procedural effects by presenting tests in a child-friendly game-format. Stimulus-generation, management of test protocols and control of test presentation is mediated by the IHR-STAR software platform. This provides a standardized methodology for a range of applications and ensures replicable procedures across testers. IHR-STAR provides a flexible, user-programmable environment that currently has additional applications for hearing screening, mapping cochlear implant electrodes, and academic research or teaching.

A test battery (IMAP) for performing an in-depth assessment of auditory and cognitive abilities contributing to listening skills is described. It is quick to administer, child-friendly and free from linguistic confounds. Stimulus generation and protocol management are controlled via a software platform (IHR-STAR) to ensure replicable procedures.

IMAPのテストバッテリー：リスニングの意味を理解する

The ability to hear is only the first step towards making sense of the range of information contained in an auditory signal. Of equal importance are the ...

医学

Roux-en-Y Gastric Bypass Operation in Rats

Currently, the most effective therapy for the treatment of morbid obesity to induce significant and maintained body weight loss with a proven mortality benefit is bariatric surgery1,2. Consequently, there has been a steady rise in the number of bariatric operations done worldwide in recent years with the Roux-en-Y gastric bypass (gastric bypass) being the most commonly performed operation3. Against this background, it is important to understand the physiological mechanisms by which gastric bypass induces and maintains body weight loss. These mechanisms are yet not fully understood, but may include reduced hunger and increased satiation4,5, increased energy expenditure6,7, altered preference for food high in fat and sugar8,9, altered salt and water handling of the kidney10 as well as alterations in gut microbiota11. Such changes seen after gastric bypass may at least partly stem from how the surgery alters the hormonal milieu because gastric bypass increases the postprandial release of peptide-YY (PYY) and glucagon-like-peptide-1 (GLP-1), hormones that are released by the gut in the presence of nutrients and that reduce eating12.
During the last two decades numerous studies using rats have been carried out to further investigate physiological changes after gastric bypass. The gastric bypass rat model has proven to be a valuable experimental tool not least as it closely mimics the time profile and magnitude of human weight loss, but also allows researchers to control and manipulate critical anatomic and physiologic factors including the use of appropriate controls. Consequently, there is a wide array of rat gastric bypass models available in the literature reviewed elsewhere in more detail 13-15. The description of the exact surgical technique of these models varies widely and differs e.g. in terms of pouch size, limb lengths, and the preservation of the vagal nerve. If reported, mortality rates seem to range from 0 to 35%15. Furthermore, surgery has been carried out almost exclusively in male rats of different strains and ages. Pre- and postoperative diets also varied significantly.
Technical and experimental variations in published gastric bypass rat models complicate the comparison and identification of potential physiological mechanisms involved in gastric bypass. There is no clear evidence that any of these models is superior, but there is an emerging need for standardization of the procedure to achieve consistent and comparable data. This article therefore aims to summarize and discuss technical and experimental details of our previously validated and published gastric bypass rat model.

Numerous studies using gastric bypass rat models have been recently conducted to uncover the underlying physiological mechanisms of Roux-en-Y gastric bypass operations. This article aims to demonstrate and discuss the technical and experimental details of our published gastric bypass rat model to understand advantages and limitations of this experimental tool.

ラットにおけるルー-en-Y法胃バイパス手術

Currently, the most effective therapy for the treatment of morbid obesity to induce significant and maintained body weight loss with a proven mortality ...

Fast and Accurate Exhaled Breath Ammonia Measurement

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that uses a quantum cascade based laser. The monitor is coupled to a sampler that measures mouth pressure and carbon dioxide. The system is temperature controlled and specifically designed to address the reactivity of this compound. The sampler provides immediate feedback to the subject and the technician on the quality of the breath effort. Together with the quick response time of the monitor, this system is capable of accurately measuring exhaled breath ammonia representative of deep lung systemic levels. 
Because the system is easy to use and produces real time results, it has enabled experiments to identify factors that influence measurements. For example, mouth rinse and oral pH reproducibly and significantly affect results and therefore must be controlled. Temperature and mode of breathing are other examples. As our understanding of these factors evolves, error is reduced, and clinical studies become more meaningful. This system is very reliable and individual measurements are inexpensive. 
The sampler is relatively inexpensive and quite portable, but the monitor is neither. This limits options for some clinical studies and provides rational for future innovations.

Ammonia is an important physiologic metabolite relevant to various disease and wellness states. It is also a difficult molecule to measure in breath, which demands particular precautions be taken to obtain accurate results. Not all factors influencing ammonia are known, but progress can be difficult without accounting for these factors.

高速かつ正確な呼気アンモニア測定

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that ...

Open Tracheostomy Gastric Acid Aspiration Murine Model of Acute Lung Injury Results in Maximal Acute Nonlethal Lung Injury

Acid pneumonitis is a major cause of sterile acute lung injury (ALI) in humans. Acid pneumonitis spans the clinical spectrum from asymptomatic to acute respiratory distress syndrome (ARDS), characterized by neutrophilic alveolitis, and injury to both alveolar epithelium and vascular endothelium. Clinically, ARDS is defined by acute onset of hypoxemia, bilateral patchy pulmonary infiltrates and non-cardiogenic pulmonary edema. Human studies have provided us with valuable information about the physiological and inflammatory changes in the lung caused by ARDS, which has led to various hypotheses about the underling mechanisms. Unfortunately, difficulties determining the etiology of ARDS, as well as a wide range of pathophysiology have resulted in a lack of critical information that could be useful in developing therapeutic strategies.
Translational animal models are valuable when their pathogenesis and pathophysiology accurately reproduce a concept proven in both in vitro and clinical settings. Although large animal models (e.g., sheep) share characteristics of the anatomy of human trachea-bronchial tree, murine models provide a host of other advantages including: low cost; short reproductive cycle lending itself to greater data acquisition; a well understood immunologic system; and a well characterized genome leading to the availability of a variety of gene deletion and transgenic strains. A robust model of low pH induced ARDS requires a murine ALI that targets mainly the alveolar epithelium, secondarily the vascular endothelium, as well as the small airways leading to the alveoli. Furthermore, a reproducible injury with wide differences between different injurious and non-injurious insults is important.
The murine gastric acid aspiration model presented here using hydrochloric acid employs an open tracheostomy and recreates a pathogenic scenario that reproduces the low pH pneumonitis injury in humans. Additionally, this model can be used to examine interaction of a low pH insult with other pulmonary injurious entities (e.g., food particles, pathogenic bacteria).

This protocol induces acute lung injury in a mouse that has close fidelity to the pathogenesis of acid pneumonitis observed in humans. We generate a maximal acute nonlethal low pH lung injury and account for differences in rodent-human anatomic respiratory structure using an open tracheostomy coupled with circumferential pressure release.

最大急性非致死肺損傷における急性肺損傷の結果のオープン気管切開胃酸吸引マウスモデル

Acid pneumonitis is a major cause of sterile acute lung injury (ALI) in humans. Acid pneumonitis spans the clinical spectrum from asymptomatic to acute ...

Techniques of Sleeve Gastrectomy and Modified Roux-en-Y Gastric Bypass in Mice

Obesity is a major public health issue, with a prevalence of 4 to 28% for men and 6.2 to 36.5% for women in Europe (from 2003 to 2008). Morbid obesity is frequently associated with metabolic complications, such as type 2 diabetes, hypertension, and dyslipidemia, reducing life expectancy and quality. In the absence of any effective noninvasive treatments, bariatric surgery is a valuable therapeutic option for patients with morbid obesity (body mass index (BMI) &#62;40 kg/m2), leading to long-term, sustained weight loss and improvements in metabolic complications. However, the underlying cellular and molecular mechanisms sustaining the beneficial effects of bariatric surgery are not yet fully understood. Due to the numerous genetically-modified strains available, the mouse model is the most convenient animal model to explore the molecular mechanisms behind the pleiotropic beneficial effects of bariatric surgeries. Here, we detailed the optimized healthcare methods and surgical protocols in mice for the two most widely-used bariatric surgeries: the sleeve gastrectomy and the modified Roux-en-Y gastric bypass. Deciphering the molecular mechanisms underlying the therapeutic effects of bariatric surgeries offers the promise of identifying new therapeutics targets.

Bariatric surgery is the most efficient way to reduce body weight and the deadly metabolic complications (diabetes, obesity, and dyslipidemia) frequently associated with morbid obesity. Mouse models of bariatric surgery represent a unique asset for deciphering molecular mechanisms behind the beneficial effects of these surgeries on diabetes, hypertension, and dyslipidemia.

スリーブ胃切除およびマウスにおける変更されたルーワイ胃バイパスのテクニック

Obesity is a major public health issue, with a prevalence of 4 to 28% for men and 6.2 to 36.5% for women in Europe (from 2003 to 2008). Morbid obesity is ...

A Blood-based Test for the Detection of ROS1 and RET Fusion Transcripts from Circulating Ribonucleic Acid Using Digital Polymerase Chain Reaction

We have developed novel methods for the isolation and characterization of tumor-derived circulating ribonucleic acid (cRNA) for blood-based liquid biopsy. Robust detection of cRNA recovered from blood represents a solution to a critical unmet need in clinical diagnostics. The test begins with the collection of whole blood into blood collection tubes containing preservatives that stabilize cRNA. Cell-free, exosomal, and platelet-associated RNA is isolated from plasma in this test system. The cRNA is reverse transcribed to complementary DNA (cDNA) and amplified using digital polymerase chain reaction (dPCR). Samples are evaluated for both the target biomarker as well as a control gene. Test validation included limit of detection, accuracy, and robustness studies with analytic samples. The method developed as a result of these studies reproducibly detect multiple fusion variants for ROS1 (C-Ros proto-oncogene 1; 8 variants) and RET (rearranged during transfection proto-oncogene; 8 variants). The sample processing workflow has been optimized so that test results can consistently be generated within 72 hours of sample receipt.

Detection of circulating ribonucleic acid (cRNA) from blood is an unmet need in clinical diagnostics. Here we describe methods that characterize cRNA from non-small cell lung cancer patients using sensitive and specific digital polymerase chain reaction. The tests meet design requirements to detect fusion variants within 72 hours.

血液ベース ROS1 の検出のためのテストし、デジタル ポリメラーゼ連鎖反応を使用してリボ核酸を循環から融合の成績証明書を RET

We have developed novel methods for the isolation and characterization of tumor-derived circulating ribonucleic acid (cRNA) for blood-based liquid biopsy. ...

Protocol to Create Chronic Wounds in Diabetic Mice

Chronic wounds develop as a result of defective regulation in one or more complex cellular and molecular processes involved in proper healing. They impact ~6.5M people and cost ~$40B/year in the US alone. Although a significant effort has been invested in understanding how chronic wounds develop in humans, fundamental questions remain unanswered. Recently, we developed a novel mouse model for diabetic chronic wounds that have many characteristics of human chronic wounds. Using db/db-/- mice, we can generate chronic wounds by inducing high levels of oxidative stress (OS) in the wound tissue immediately after wounding, using a one-time treatment with inhibitors specific to the antioxidant enzymes catalase and glutathione peroxidase. These wounds have high levels of OS, develop biofilm naturally, become fully chronic within 20 days after treatment and can remain open more for more than 60 days. This novel model has many features of diabetic chronic wounds in humans and therefore can contribute significantly to advancing fundamental understanding of how wounds become chronic. This is a major breakthrough because chronic wounds in humans cause significant pain and distress to patients and result in amputation if unresolved. Moreover, these wounds are very expensive and time-consuming to treat, and lead to significant loss of personal income to patients. Advancements in this field of study through the use of our chronic wound model can significantly improve health care for millions who suffer under this debilitating condition. In this protocol, we describe in great detail the procedure to cause acute wounds to become chronic, which has not been done before.

Chronic wounds are developed from acute wounds on a diabetic mouse model by inducing high levels of oxidative stress after a full-thickness cutaneous wound. The wound is treated with inhibitors for catalase and glutathione peroxidase, resulting in impaired healing and biofilm development by bacteria present in the skin microbiome.

糖尿病マウスの慢性創傷を作成するためのプロトコル

Chronic wounds develop as a result of defective regulation in one or more complex cellular and molecular processes involved in proper healing. They impact ...

Non-invasive Assessment of Dorsiflexor Muscle Function in Mice

Assessment of skeletal muscle contractile function is an important measurement for both clinical and research purposes. Numerous conditions can negatively affect skeletal muscle. This can result in a loss of muscle mass (atrophy) and/or loss of muscle quality (reduced force per unit of muscle mass), both of which are prevalent in chronic disease, muscle-specific disease, immobilization, and aging (sarcopenia). Skeletal muscle function in animals can be evaluated by a range of different tests. All tests have limitations related to the physiological testing environment, and the selection of a specific test often depends on the nature of the experiments. Here, we describe an in vivo, non-invasive technique involving a helpful and easy assessment of force frequency-curve (FFC) in mice that can be performed on the same animal over time. This permits monitoring of disease progression and/or efficacy&#160;of a potential therapeutic treatment.

Measurement of rodent skeletal muscle contractile function is a useful tool that can be used to track disease progression as well as efficacy of therapeutic intervention. We describe here the non-invasive, in vivo assessment of the dorsiflexor muscles that can be repeated over time in the same mouse.

マウスで内がえし筋機能の非侵襲的評価

Assessment of skeletal muscle contractile function is an important measurement for both clinical and research purposes. Numerous conditions can negatively ...

Assessment and Characterization of Hyaloid Vessels in Mice

In the eye, the embryonic hyaloid vessels nourish the developing lens and retina and regress when the retinal vessels develop. Persistent or failed regression of hyaloid vessels can be seen in diseases such as persistent hyperplastic primary vitreous (PHPV), leading to an obstructed light path and impaired visual function. Understanding the mechanisms underlying the hyaloid vessel regression may lead to new molecular insights into the vascular regression process and potential new ways to manage diseases with persistent hyaloid vessels. Here we describe the procedures for imaging hyaloid in live mice with optical coherence tomography (OCT) and fundus fluorescein angiography (FFA) and a detailed technical protocol of isolating and flat-mounting hyaloid ex vivo for quantitative analysis. Low-density lipoprotein receptor-related protein 5 (LRP5) knockout mice were used as an experimental model of persistent hyaloid vessels, to illustrate the techniques. Together, these techniques may facilitate a thorough assessment of hyaloid vessels as an experimental model of vascular regression and studies on the mechanism of persistent hyaloid vessels.

This protocol describes both in vivo and ex vivo methods to fully visualize and characterize hyaloid vessels, a model of vascular regression in mouse eyes, using optical coherence tomography and fundus fluorescein angiography for the live imaging and ex vivo isolation and subsequent flat mount of hyaloid for quantitative analysis.

マウスにおけるヒアロイド血管の評価と特徴付け

In the eye, the embryonic hyaloid vessels nourish the developing lens and retina and regress when the retinal vessels develop. Persistent or failed ...

Using a Chemical Biopsy for Graft Quality Assessment

Kidney transplantation is a life-saving treatment for a large number of people with end-stage renal dysfunction worldwide. The procedure is associated with an increased survival rate and greater quality of patient&#8217;s life when compared to conventional dialysis. Regrettably, transplantology suffers from a lack of reliable methods for organ quality assessment. Standard diagnostic techniques are limited to macroscopic appearance inspection or invasive tissue biopsy, which do not provide comprehensive information about the graft. The proposed protocol aims to introduce solid phase microextraction (SPME) as an ideal analytical method for comprehensive metabolomics and lipidomic analysis of all low molecular compounds present in kidneys allocated for transplantation. The small size of the SPME probe enables performance of a chemical biopsy, which enables extraction of metabolites directly from the organ without any tissue collection. The minimum invasiveness of the method permits execution of multiple analyses over time: directly after organ harvesting, during its preservation, and immediately after revascularization at the recipient&#8217;s body. It is hypothesized that the combination of this novel sampling method with a high-resolution mass spectrometer will allow for discrimination of a set of characteristic compounds that could serve as biological markers of graft quality and indicators of possible development of organ dysfunction.

The protocol presents utilization of the chemical biopsy approach followed by comprehensive metabolomic and lipidomic analysis for quality assessment of kidney grafts allocated for transplantation.

移植質評価のための化学バイオプシーの使用

Kidney transplantation is a life-saving treatment for a large number of people with end-stage renal dysfunction worldwide. The procedure is associated ...