我々は感謝NIDDKからスタンフォード小児研究基金とK08DK087895 - 01からのパイロット助成金の支援を認める。

1。マウスの膀胱壁インジェクションマウス系統、年齢、および性別の選択は、実験的なニーズによって決定されます。これは老化の前に免疫学的成熟度のウィンドウであるため、我々は、生後8〜12週間の間にマウスを使用してください。一般的なガイドラインとして、マウスはストレス誘発性の交絡因子を避けるために、実験操作の前に少なくとも一週間に到着してください。 <ol><li>石鹸と水で手術台の表面を清掃してください。 </li><li>慈徳のワイプまたは消毒ワイプで手術台の表面を拭いてください。 </li><li>手術に使用する前にきれいな手術器具をオートクレーブ。 </li><li>さらに、使用直前に熱いビーズ滅菌器で手術器具を殺菌するだけでなく、手術中に動物の間で。 </li><li>最初の使用前と最終的な外科手術の最後に無水アルコールで繰り返し吸引と注射で100μLハミルトンシリンジと29または30ゲージ（1 / 2インチ長い）針を清掃してください。 </li><li>それぞれの動物の間でリン酸緩衝生理食塩水、滅菌とハミルトンシリンジを洗浄し、すすいでください。 </li><li> 2〜5パーセントの間にイソフルランセットを使用して、イソフルラン誘導チャンバ内にそれらを置くことによってマウスを麻酔。 </li><li>全身麻酔が達成されると、正常な体温を維持するためにウォームなパッドの下で仰臥位での室と場所からマウスを取り外します。 </li><li> （適切な麻酔を維持するために、必要に応じて1〜3パーセントから滴定）イソフルラン気化を含むノズルで、動物の鼻を配置することにより、メンテナンスの麻酔を達成する。 </li><li>バリカンで腹部の皮膚を剃る。 </li><li>手術中の糞便汚染を防ぐために、肛門をカバーするために使い捨て、滅菌外科用ドレープを使用してください。 </li><li>手術野（下腹部）をカバーする2番目の使い捨て、滅菌外科用ドレープを使用してください。 </li><li> Betadineに浸したガーゼの3つの部分で準備腹部。このステップを3回繰り返します。 </li><li>拡大のための解剖顕微鏡を使って、ハサミで下の正中腹部切開を行います。 </li><li>膀胱を公開。 </li><li>膀胱がいっぱいになっている場合、部分的にドームでの穏やかな下方圧力によってそれを解凍。 </li><li>上を向いて針のベベルと膀胱のドーム（壁内注射）の壁に、29または30ゲージの注射針と注射器を用いて、試料溶液を（最大50μL）注入。 </li><li>膀胱壁に試料溶液を注入する注射器のプランジャーを押してください。極めて限局したブレブは、膀胱壁への細胞の正常な注射の指示です。 </li><li> 、シリンジを取り外します創傷クリップで切開を閉じて、マウスが温暖化のパッドに回復することができます。 </li></ol> 2。代表的な結果： 漏れ流体をしないとサイズで安定したまま極めて限局したブレブは、膀胱壁（図1）への細胞の成功注入の指示です。組織学的分析は、膀胱壁に注入された細胞の存在を確認するために実行することができます。 <img src="/files/ftp_upload/2523/2523fig1.jpg" alt="図1"/> 図1。説明のためにインドのインクを使用して成功した膀胱壁のインジェクションの例。 <img src="/files/ftp_upload/2523/2523fig2.jpg" alt="図2"/> 図2実験手順の模式図。

<ol>
	<li>Dinney, C. P., Fishbeck, R., Singh, R. K., Eve, B., Pathak, S., Brown, N., Xie, B., Fan, D., Bucana, C., Fidler, I. J., Killion, 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=Isolation+and+Characterization+of+Metastatic+Variants+from+Human+Transitional+Cell+Carcinoma+Passaged+by+Orthotopic+Implantation+in+Athymic+Nude+Mice.">Isolation and Characterization of Metastatic Variants from Human Transitional Cell Carcinoma Passaged by Orthotopic Implantation in Athymic Nude Mice.</a> The Journal of Urology. 154, 1532-1538 (1995).</li><li>Singh, A. V., Franke, A. A., Blackburn, G. L., Zhou, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Soy+Phytochemicals+Prevent+Orthotopic+Growth+and+Metastasis+of+Bladder+Cancer+in+Mice+by+Alterations+of+Cancer+Cell+Proliferation+and+Apoptosis+and+Tumor+Angiogenesis.">Soy Phytochemicals Prevent Orthotopic Growth and Metastasis of Bladder Cancer in Mice by Alterations of Cancer Cell Proliferation and Apoptosis and Tumor Angiogenesis.</a> Cancer Res. 66, 1851-1858 (2006).</li><li>Yanagiuchi, A., Miyake, H., Nomi, M., Takenaka, A., Fujisawa, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modulation+of+the+Microenvironment+by+Growth+Factors+Regulates+the+In+Vivo+Growth+of+Skeletal+Myoblasts.">Modulation of the Microenvironment by Growth Factors Regulates the In Vivo Growth of Skeletal Myoblasts.</a> BJU International. 103, 1569-1573 (2009).</li><li>Miyake, H., Hara, I., Yamanaka, K., Gohji, K., Arakawa, S., Kamidono, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Overexpression+of+Bcl-2+Enhances+Metastatic+Potential+of+Human+Bladder+Cancer+Cells.">Overexpression of Bcl-2 Enhances Metastatic Potential of Human Bladder Cancer Cells.</a> British Journal of Cancer. 79, 1651-1656 (1999).</li><li>Chancellor, M. B., Yokoyama, T., Tirney, S., Mattes, C. E., Ozawa, H., Yoshimura, N., Groat, W. C. d. e., Huard, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preliminary+Results+of+Myoblast+Injection+into+the+Urethra+and+Bladder+Wall:+a+Possible+method+for+the+Treatment+of+Stress+Urinary+Incontinence+and+Impaired+Detrusor+Contractility.">Preliminary Results of Myoblast Injection into the Urethra and Bladder Wall: a Possible method for the Treatment of Stress Urinary Incontinence and Impaired Detrusor Contractility.</a> Neurourol Urodyn. 19, 279-287 (2000).</li><li>Dinney, C. P., Tanguay, S., Bucana, C. D., Eve, B. Y., Fidler, I. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intravesical+Liposomal+Muramyl+Tripeptide+Phosphatidylethanolamine+Treatment+of+Human+Bladder+Carcinoma+Growing+in+Nude+Mice.">Intravesical Liposomal Muramyl Tripeptide Phosphatidylethanolamine Treatment of Human Bladder Carcinoma Growing in Nude Mice.</a> J Interferon Cytokine Res. 15, 585-592 (1995).</li><li>Mohamedali, K. A., Kedary, D., Sweeney, P., Kamaty, A., Davisy, D. W., Evey, B. Y., Huangy, S., Thorpez, P. E., Dinney, C. P., Rosenblum, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Vascular-targeting+Fusion+toxin+VEGF121/rGel+Inhibits+the+Growth+of+Orthotopic+Human+Bladder+Carcinoma+Tumors.">The Vascular-targeting Fusion toxin VEGF121/rGel Inhibits the Growth of Orthotopic Human Bladder Carcinoma Tumors.</a> Neoplasia. 7, 912-920 (2005).</li><li>Slaton, J. W., Perrotte, P., Inoue, K., Dinney, C. P., Fidler, I. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interferon-&#945;-mediated+Down-Regulation+of+Angiogenesis-related+Genes+and+Therapy+of+Bladder+Cancer+Are+Dependent+on+Optimization+of+Biological+Dose+and+Schedule.">Interferon-&#945;-mediated Down-Regulation of Angiogenesis-related Genes and Therapy of Bladder Cancer Are Dependent on Optimization of Biological Dose and Schedule.</a> Clinical Cancer Research. 5, 2726-2734 (1999).</li><li>Yokoyama, T., Huard, J., Pruchnic, R., Yoshimura, N., Qu, Z., Cao, B., De Groat, W. C., Kumon, H., Chancellor, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle-Derived+Cell+Transplantation+and+Differentiation+into+Lower+Urinary+Tract+Smooth+Muscle.">Muscle-Derived Cell Transplantation and Differentiation into Lower Urinary Tract Smooth Muscle.</a> Urology. 57, 826-831 (2001).</li><li>Chan, E., Patel, A., Heston, W., Larchian, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+Orthotopic+Models+for+Bladder+Cancer+Research.">Mouse Orthotopic Models for Bladder Cancer Research.</a> BJU International. 104, 1286-1291 (1988).</li><li>Russell, P. J., Jelbart, M., Wills, E., Singh, S., Wass, J., Witherspoon, J., Raghavan, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Establishment+and+characterization+of+a+new+human+bladder+cancer+cell+line+showing+features+of+squamous+and+glandular+differentiation.">Establishment and characterization of a new human bladder cancer cell line showing features of squamous and glandular differentiation.</a> Int. J. Cancer. 41, 74-74 (1988).</li><li>Ahlering, T. E., Dubeau, L., Jones, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+in+vivo+model+to+study+invasion+and+metastasis+of+human+bladder+carcinoma.">A new in vivo model to study invasion and metastasis of human bladder carcinoma.</a> Cancer Res. 41, 6660-6660 (1987).</li><li>Heaney, J. A., Omellas, E. P., Daly, J. J., Lin, J. C., Rout, G. R. <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+growth+of+human+bladder+cancer+cell+lines.">In vivo growth of human bladder cancer cell lines.</a> Invest. Urol. 15, 380-380 (1978).</li><li>Hadaschik, B., Black, P., Sea, J., Metwalli, A., Fazli, L., Dinney, C., Gleave, M., So, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+validated+mouse+model+for+orthotopic+bladder+cancer+using+transurethral+tumor+inoculation+and+bioluminescence+imaging.">A validated mouse model for orthotopic bladder cancer using transurethral tumor inoculation and bioluminescence imaging.</a> BJU International. 100, 1377-1384 (2007).</li></ol>

利害の衝突は宣言されません。

マウスの膀胱壁の注入は、膀胱壁の定義の領域に特定の細胞の移植が可能になります。この手法は、膀胱癌、平滑筋、および幹細胞生物学のマウスモデルのための広いアプリケーションを持っています。彼らの成長と分化を生理学的に関連する解剖学的文脈で検討することができるように定義することにより、マウスの膀胱壁の注入は、同所場所に膀胱癌や幹細胞の導入を促進する。実際に?...

<table border="1"><tbody><tr><td> 試薬の名前 </td><td> 会社 </td><td> カタログ番号 </td></tr><tr><td>イソフルラン（Aerrane） </td><td>バクスター</td><td> NDC 10019-773-60 </td></tr><tr><td> 29または30ゲージの針</td><td>ハミルトン</td><td> 7803から06または7803から07 </td></tr><tr><td> 100マイクロリットルのシリンジ</td><td>ハミルトン</td><td> 7656〜01 </td></tr></tbody></table>

mouse bladder wall injection

マウスの膀胱壁の注入は、同所の幹細胞、平滑筋、および癌の生物学を含む膀胱の現象を研究するために便利なテクニックです。注射を開始する前に、外科領域は、石けんと水と消毒液で洗浄する必要があります。手術用機器は使用前に、各動物の間で滅菌する必要があります。各マウスは、吸入イソフルラン麻酔（誘導2-5％、メンテナンス1-3％）とハサミで正中腹部切開を行うことによって露出は膀胱の下に配置されます。膀胱がいっぱいになっている場合、それは2本の指で圧迫穏やかなことで、部分的に減圧される。興味の細胞懸濁液は、intramurally 29または30ゲージの注射針と1ccのまたはより小さいシリンジを使用して膀胱のドームの壁に注入されます。傷は、傷のクリップと温暖化のパッドに回復させたマウスを用いて閉じられます。膀胱壁の注入は、練習で習得することができる繊細な顕微技術です。

Watch this Scientific Journal Video about Mouse Bladder Wall Injection at JoVE.com

Mouse Bladder Wall Injection

マウスの膀胱壁の注入は、同所膀胱の幹細胞と癌の生物学を研究するために有用なアプローチである。この繊細な顕微法は慎重に手法と実践を習得することができます。

マウスの膀胱壁インジェクション

Mouse bladder wall injection is a useful technique to orthotopically study bladder phenomena, including stem cell, smooth muscle, and cancer biology. ...

Research

JoVE Journal

Medicine

23.8K ビュー.  Stanford University School of Medicine. マウスの膀胱壁の注入は、同所膀胱の幹細胞と癌の生物学を研究するために有用なアプローチである。この繊細な顕微法は慎重に手法と実践を習得することができます。

ビデオ: Mouse Bladder Wall Injection

An Orthotopic Model of Murine Bladder Cancer

In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and subsequent cell adhesion. After their bladders are transurethrally catheterized and drained, animals are again catheterized to permit insertion of a platinum wire into bladders without damaging the urethra or bladder. The catheters are made of Teflon to serve as an insulator for the wire, which will conduct electrical current into the bladder to create a burn injury. An electrocautery unit is used to deliver 2.5W to the exposed end of the wire, burning away extracellular layers and providing attachment sites for carcinoma cells that are delivered in suspension to the bladder through a subsequent catheterization. Cells remain in the bladder for 90 minutes, after which the catheters are removed and the bladders allowed to drain naturally. The development of tumor is monitored via ultrasound. Specific attention is paid to the catheterization technique in the accompanying video.

Bladder tumors are established in female mice in a minimally invasive fashion through catheterization, local cauterization, and subsequent adhesion of carcinoma cells to the burn sites.

マウス膀胱癌の同所性モデル

In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and ...

医学

Hyperinsulinemic-euglycemic Clamps in Conscious, Unrestrained Mice

Type 2 diabetes is characterized by a defect in insulin action. The hyperinsulinemic-euglycemic clamp, or insulin clamp, is widely considered the "gold standard" method for assessing insulin action in vivo. During an insulin clamp, hyperinsulinemia is achieved by a constant insulin infusion. Euglycemia is maintained via a concomitant glucose infusion at a variable rate. This variable glucose infusion rate (GIR) is determined by measuring blood glucose at brief intervals throughout the experiment and adjusting the GIR accordingly. The GIR is indicative of whole-body insulin action, as mice with enhanced insulin action require a greater GIR. The insulin clamp can incorporate administration of isotopic 2[14C]deoxyglucose to assess tissue-specific glucose uptake and [3-3H]glucose to assess the ability of insulin to suppress the rate of endogenous glucose appearance (endoRa), a marker of hepatic glucose production, and to stimulate the rate of whole-body glucose disappearance (Rd).
The miniaturization of the insulin clamp for use in genetic mouse models of metabolic disease has led to significant advances in diabetes research. Methods for performing insulin clamps vary between laboratories. It is important to note that the manner in which an insulin clamp is performed can significantly affect the results obtained. We have published a comprehensive assessment of different approaches to performing insulin clamps in conscious mice1 as well as an evaluation of the metabolic response of four commonly used inbred mouse strains using various clamp techniques2. Here we present a protocol for performing insulin clamps on conscious, unrestrained mice developed by the Vanderbilt Mouse Metabolic Phenotyping Center (MMPC; URL: www.mc.vanderbilt.edu/mmpc). This includes a description of the method for implanting catheters used during the insulin clamp. The protocol employed by the Vanderbilt MMPC utilizes a unique two-catheter system3. One catheter is inserted into the jugular vein for infusions. A second catheter is inserted into the carotid artery, which allows for blood sampling without the need to restrain or handle the mouse. This technique provides a significant advantage to the most common method for obtaining blood samples during insulin clamps which is to sample from the severed tip of the tail. Unlike this latter method, sampling from an arterial catheter is not stressful to the mouse1. We also describe methods for using isotopic tracer infusions to assess tissue-specific insulin action. We also provide guidelines for the appropriate presentation of results obtained from insulin clamps.

The hyperinsulinemic-euglycemic clamp, or insulin clamp, is the gold standard for assessing insulin action in vivo. A method for performing insulin clamps in mice is described. This includes a method for arterial catheterization that permits experiments to be performed in conscious, unrestrained mice with minimal stress.

意識、無拘束マウスにおける高インスリン血症、正常血糖クランプ

Type 2 diabetes is characterized by a defect in insulin action. The hyperinsulinemic-euglycemic clamp, or insulin clamp, is widely considered the "gold ...

The Use of Cystometry in Small Rodents: A Study of Bladder Chemosensation

The lower urinary tract (LUT) functions as a dynamic reservoir that is able to store urine and to efficiently expel it at a convenient time. While storing urine, however, the bladder is exposed for prolonged periods to waste products. By acting as a tight barrier, the epithelial lining of the LUT, the urothelium, avoids re-absorption of harmful substances. Moreover, noxious chemicals stimulate the bladder's nociceptive innervation and initiate voiding contractions that expel the bladder's contents. Interestingly, the bladder's sensitivity to noxious chemicals has been used successfully in clinical practice, by intravesically infusing the TRPV1 agonist capsaicin to treat neurogenic bladder overactivity1. This underscores the advantage of viewing the bladder as a chemosensory organ and prompts for further clinical research. However, ethical issues severely limit the possibilities to perform, in human subjects, the invasive measurements that are necessary to unravel the molecular bases of LUT clinical pharmacology. A way to overcome this limitation is the use of several animal models2. Here we describe the implementation of cystometry in mice and rats, a technique that allows measuring the intravesical pressure in conditions of controlled bladder perfusion.
	After laparotomy, a catheter is implanted in the bladder dome and tunneled subcutaneously to the interscapular region. Then the bladder can be filled at a controlled rate, while the urethra is left free for micturition. During the repetitive cycles of filling and voiding, intravesical pressure can be measured via the implanted catheter. As such, the pressure changes can be quantified and analyzed. Moreover, simultaneous measurement of the voided volume allows distinguishing voiding contractions from non-voiding contractions3.
	Importantly, due to the differences in micturition control between rodents and humans, cystometric measurements in these animals have only limited translational value4. Nevertheless, they are quite instrumental in the study of bladder pathophysiology and pharmacology in experimental pre-clinical settings. Recent research using this technique has revealed the key role of novel molecular players in the mechano- and chemo-sensory properties of the bladder.

Cystometry is an efficient technique to measure bladder function of small animals in vivo. The bladder is continuously infused at rates controlled through an intravesical catheter, whereas the urethra is left free for micturition. This allows for repetitive filling and emptying of the bladder, while intravesical pressure and voided volume are recorded.

小型げっ歯類における膀胱内圧測定の使用：膀胱Chemosensationの研究

	The lower urinary tract (LUT) functions as a  dynamic reservoir that is able to store urine and to efficiently expel it at a  convenient time. While ...

An Orthotopic Bladder Tumor Model and the Evaluation of Intravesical saRNA Treatment

We present a novel method for treating bladder cancer with intravesically delivered small activating RNA (saRNA) in an orthotopic xenograft mouse bladder tumor model. The mouse model is established by urethral catheterization under inhaled general anesthetic. Chemical burn is then introduced to the bladder mucosa using intravesical silver nitrate solution to disrupt the bladder glycosaminoglycan layer and allows cells to attach. Following several washes with sterile water, human bladder cancer KU-7-luc2-GFP cells are instilled through the catheter into the bladder to dwell for 2 hours. Subsequent growth of bladder tumors is confirmed and monitored by in vivo bladder ultrasound and bioluminescent imaging. The tumors are then treated intravesically with saRNA formulated in lipid nanoparticles (LNPs). Tumor growth is monitored with ultrasound and bioluminescence. All steps of this procedure are demonstrated in the accompanying video.

Establishing an orthotopic bladder tumor model to evaluate antitumor effects of intravesically delivered saRNA and monitoring tumor growth by ultrasound and bioluminescent imaging.

同所性膀胱腫瘍モデルと膀胱内Sarnaの治療の評価

We present a novel method for treating bladder cancer with intravesically delivered small activating RNA (saRNA) in an orthotopic xenograft mouse bladder ...

Subretinal Injection of Gene Therapy Vectors and Stem Cells in the Perinatal Mouse Eye

The loss of sight affects approximately 3.4 million people in the United States and is expected to increase in the upcoming years.1 Recently, gene therapy and stem cell transplantations have become key therapeutic tools for treating blindness resulting from retinal degenerative diseases. Several forms of autologous transplantation for age-related macular degeneration (AMD), such as iris pigment epithelial cell transplantation, have generated encouraging results, and human clinical trials have begun for other forms of gene and stem cell therapies.2 These include RPE65 gene replacement therapy in patients with Leber's congenital amaurosis and an RPE cell transplantation using human embryonic stem (ES) cells in Stargardt's disease.3-4 Now that there are gene therapy vectors and stem cells available for treating patients with retinal diseases, it is important to verify these potential therapies in animal models before applying them in human studies. The mouse has become an important scientific model for testing the therapeutic efficacy of gene therapy vectors and stem cell transplantation in the eye.5-8 In this video article, we present a technique to inject gene therapy vectors or stem cells into the subretinal space of the mouse eye while minimizing damage to the surrounding tissue.

This surgical technique illustrates the injection of gene therapy vectors and stem cells into the subretinal space of the mouse eye.

マウスの眼の中に遺伝子治療ベクターと幹細胞の網膜下注入

The loss of sight affects approximately 3.4 million people in the United States and is expected to increase in the upcoming years.1 Recently, gene therapy ...

Reduction in Left Ventricular Wall Stress and Improvement in Function in Failing Hearts using Algisyl-LVR

Injection of Algisyl-LVR, a treatment under clinical development, is intended to treat patients with dilated cardiomyopathy. This treatment was recently used for the first time in patients who had symptomatic heart failure. In all patients, cardiac function of the left ventricle (LV) improved significantly, as manifested by consistent reduction of the LV volume and wall stress. Here we describe this novel treatment procedure and the methods used to quantify its effects on LV wall stress and function.
Algisyl-LVR is a biopolymer gel consisting of Na+-Alginate and Ca2+-Alginate. The treatment procedure was carried out by mixing these two components and then combining them into one syringe for intramyocardial injections. This mixture was injected at 10 to 19 locations mid-way between the base and apex of the LV free wall in patients.
Magnetic resonance imaging (MRI), together with mathematical modeling, was used to quantify the effects of this treatment in patients before treatment and at various time points during recovery. The epicardial and endocardial surfaces were first digitized from the MR images to reconstruct the LV geometry at end-systole and at end-diastole. Left ventricular cavity volumes were then measured from these reconstructed surfaces.
Mathematical models of the LV were created from these MRI-reconstructed surfaces to calculate regional myofiber stress. Each LV model was constructed so that 1) it deforms according to a previously validated stress-strain relationship of the myocardium, and 2) the predicted LV cavity volume from these models matches the corresponding MRI-measured volume at end-diastole and end-systole. Diastolic filling was simulated by loading the LV endocardial surface with a prescribed end-diastolic pressure. Systolic contraction was simulated by concurrently loading the endocardial surface with a prescribed end-systolic pressure and adding active contraction in the myofiber direction. Regional myofiber stress at end-diastole and end-systole was computed from the deformed LV based on the stress-strain relationship.

This article describes procedures for implanting a novel hydrogel in failing hearts and quantifying its effect on left ventricular wall stress and function. These procedures have been successfully applied in dogs and humans.

Algisyl-LVRを使っハーツ失敗関数内で左心室壁応力と改善の削減

Injection of  Algisyl-LVR, a treatment under clinical development, is intended to treat  patients with dilated cardiomyopathy. This treatment was recently ...

An Orthotopic Bladder Cancer Model for Gene Delivery Studies

Bladder cancer is the second most common cancer of the urogenital tract and novel therapeutic approaches that can reduce recurrence and progression are needed. The tumor microenvironment can significantly influence tumor development and therapy response. It is therefore often desirable to grow tumor cells in the organ from which they originated. This protocol describes an orthotopic model of bladder cancer, in which MB49 murine bladder carcinoma cells are instilled into the bladder via catheterization. Successful tumor cell implantation in this model requires disruption of the protective glycosaminoglycan layer, which can be accomplished by physical or chemical means. In our protocol the bladder is treated with trypsin prior to cell instillation. Catheterization of the bladder can also be used to deliver therapeutics once the tumors are established. This protocol describes the delivery of an adenoviral construct that expresses a luciferase reporter gene. While our protocol has been optimized for short-term studies and focuses on gene delivery, the methodology of mouse bladder catheterization has broad applications.

Implantation of cancer cells into the organ of origin can serve as a useful preclinical model to evaluate novel therapies. MB49 bladder carcinoma cells can be grown within the bladder following intravesical instillation. This protocol demonstrates catheterization of the mouse bladder for the purpose of tumor implantation and adenoviral delivery.

遺伝子送達研究のための同所膀胱癌モデル

Bladder cancer is the second most common cancer of the urogenital tract and novel therapeutic approaches that can reduce recurrence and progression are ...

Minimally Invasive Establishment of Murine Orthotopic Bladder Xenografts

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

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

マウス同所性膀胱異種移植片の最小侵襲性の確立

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

Urinary Bladder Distention Evoked Visceromotor Responses as a Model for Bladder Pain in Mice

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition characterized by increased urgency and frequency of urination, as well as nocturia and general pelvic pain, especially upon bladder filling or voiding. Despite years of research, the cause of IC/BPS remains elusive and treatment strategies are unable to provide complete relief to patients. In order to study nervous system contributions to the condition, many animal models have been developed to mimic the pain and symptoms associated with IC/BPS. One such murine model is urinary bladder distension (UBD). In this model, compressed air of a specific pressure is delivered to the bladder of a lightly anesthetized animal over a set period of time. Throughout the procedure, wires in the superior oblique abdominal muscles record electrical activity from the muscle. This activity is known as the visceromotor response (VMR) and is a reliable and reproducible measure of nociception. Here, we describe the steps necessary to perform this technique in mice including surgical manipulations, physiological recording, and data analysis. With the use of this model, the coordination between primary sensory neurons, spinal cord secondary afferents, and higher central nervous system areas involved in bladder pain can be unraveled. This basic science knowledge can then be clinically translated to treat patients suffering from IC/BPS.

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition characterized in part by pelvic pain. In order to study nervous system contributions to the condition, a physiological model of urinary bladder pain is used in mice and rats.

マウスの膀胱痛に対するモデルとしての膀胱膨満誘発内臓運動応答

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition ...

Ultrahigh Resolution Mouse Optical Coherence Tomography to Aid Intraocular Injection in Retinal Gene Therapy Research

HR-SD-OCT is utilized to monitor the progression of photoreceptor degeneration in live mouse models, assess the delivery of therapeutic agents into the subretinal space, and to evaluate toxicity and efficacy in vivo. HR-SD-OCT uses near infrared light (800-880 nm) and has optics specifically designed for the unique optics of the mouse eye with sub-2-micron axial resolution. Transgenic mouse models of outer retinal (photoreceptor) degeneration and controls were imaged to assess the disease progression. Pulled glass microneedles were used to deliver sub retinal injections of adeno-associated virus (AAV) or nanoparticles (NP) via a trans-scleral and trans-choroidal approach. Careful positioning of the needle into the subretinal space was required prior to a calibrated pressure injection, which delivers fluid into the sub retinal space. Real time subretinal surgery was conducted on our retinal imaging system (RIS). HR-SD-OCT demonstrated progressive uniform retinal degeneration due to expression of a toxic mutant human mutant rhodopsin (P347S) (RHOP347S) transgene in mice. HR-SD-OCT allows rigorous quantification of all the retinal layers. Outer nuclear layer (ONL) thickness and photoreceptor outer segment length (OSL) measurements correlate with photoreceptor vitality, degeneration, or rescue. The RIS delivery system allows real-time visualization of subretinal injections in neonatal (~P10-14) or adult mice, and HR-SD-OCT immediately determines success of delivery and maps areal extent. HR-SD-OCT is a powerful tool that can evaluate the success of subretinal surgery in mice, in addition to measuring vitality of photoreceptors in vivo. HR-SD-OCT can also be used to identify uniform animal cohorts to evaluate the extent of retinal degeneration, toxicity, and therapeutic rescue in preclinical gene therapy research studies.

Here we demonstrate a novel approach to using high resolution spectral-domain optical coherence tomography (HR-SD-OCT) to assist delivery of gene therapy agents into the subretinal space, assess its areal coverage, and characterize photoreceptor vitality.

超高解像度マウス光コヒーレンストモグラフィー網膜遺伝子治療研究における眼内注入を支援するために

HR-SD-OCT is utilized to monitor the progression of photoreceptor degeneration in live mouse models, assess the delivery of therapeutic agents into the ...