우리는 기꺼이 NIDDK에서 스탠포드 소아 연구 기금과 K08DK087895 - 01의 파일럿 부여의 지원을 인정합니다.

1. 마우스 블레더 월 주입 마우스 변형, 연령 및 성별의 선택은 실험 요구에 의해 결정됩니다. 이것이 노화하기 전에 면역 성숙의 윈도우이기 때문에 우리는, 나이, 8, 12 주 정도 마우스를 사용합니다. 일반적인 지침으로, 생쥐가 스트레스 유발 혼란함을 주죠 요인을 방지하기 위해 실험 조작하기 전에 적어도 한 주에 도착해야합니다. <ol><li> 비누와 물로 수술 테이블 표면을 청소하십시오. </li><li> Cide의 Swipes 또는 살균 잎사귀와 수술 테이블 표면을 닦아주십시오. </li><li> 수술에 사용하기 전에 깨끗한 수술 악기 압력솥. </li><li> 또한, 사용하는 즉시 사전 핫 구슬 살균기와 수술 도구를 소독뿐만 아니라, 수술하는 동안 동물 사이. </li><li> 100 μL 해밀턴 주사기 29 또는 30 게이지 (긴 2분의 1 인치) 처음 사용하기 전에 절대 알코올로 반복 흡인과 주사로하고 최종 수술의 끝에 바늘을 청소하십시오. </li><li> 각 동물 간의 생리 인산염 버퍼 소독과 해밀턴 주사기를 씻고 린스. </li><li> 2~5% 사이 isoflurane 세트와 isoflurane 유도 챔버에서 그들을 배치하여 생쥐를 마취. </li><li> 전신 마취를 얻을되면 정상적인 체온을 유지하기 위해 밑에 따뜻한 패드와 부정사 위치에 챔버와 장소에서 마우스를 제거합니다. </li><li> (적절한 마취를 유지하기 위해 필요한 1~3%에서 titrated) isoflurane 기화가 들어있는 노즐​​에서 동물의 주둥이를 배치하여 유지 보수 마취를 얻을 수 있습니다. </li><li> 가위로 복부 피부를 면도. </li><li> 수술하는 동안 배설물로 오염을 방지하기 위해 항문을 충당하기 위해 일회용, 살균 외과 드레이프를 사용하십시오. </li><li> 외과 분야 (낮은 복부)를 충당하기 위해 두 번째 일회용, 살균 외과 드레이프를 사용하십시오. </li><li> Betadine - 젖었 거즈 3 가지로 준비 복부. 이 단계를 세 번 반복합니다. </li><li> 확대를위한 해부 현미경을 사용하여 가위로 더 낮은 복부 정중선 절개를합니다. </li><li> 방광을 폭로. </li><li> 방광이 가득 경우, 부분적으로 돔에서 완만한 하향 압력에 의해 그것을 압축 해제. </li><li> 위쪽을 향하고 바늘의 베벨과 방광 돔 (교내 사출)의 벽에, 29 또는 30 게이지 바늘과 주사기를 사용하여 샘​​플 솔루션을 (최대 50 μL) 주사. </li><li> 방광 벽에 ​​샘플 솔루션을 주입하는 주사기의 플런저를 밀어. 잘화된 물집이 방광 벽에있는 세포의 성공적인 주입의 표시입니다. </li><li> , 주사기를 제거 상처 클립으로 절개를 닫고 마우스가 온난 패드에 복구할 수 있습니다. </li></ol> 2. 대표 결과 : 누출 액을하지 않으며 크기의 안정 유지 잘화된 물집은 방광 벽 (그림 1)에 세포의 성공적인 주입의 표시입니다. Histological 분석은 방광 벽에 ​​주입된 세포의 존재를 확인하기 위해 수행할 수 있습니다. <img src="/files/ftp_upload/2523/2523fig1.jpg" alt="그림 1"/> 그림 1. 보여주기위한 인도 잉크를 사용하여 성공적인 방광 벽 주입의 예. <img src="/files/ftp_upload/2523/2523fig2.jpg" alt="그림 2"/> 그림 2. 실험 절차의 도식.

<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>

관심 없음 충돌 선언하지 않습니다.

마우스 방광 벽 주입은 방광 벽 정의된 영역에 특정 세포의 주입을 허용합니다. 이 기술은 방광 암, 부드러운 근육, 그리고 줄기 세포 생물학의 마우스 모델에 대한 다양한 응용 프로그램을하고 있습니다. 자신의 성장과 분화가 생리학 관련 해부학 맥락에서 공부를 할 수 있도록 정의함으로써, 마우스 방광 벽 주입은 orthotopic 위치에 방광 암 또는 줄기 세포의 도입을 용이하게합니다. 사실, Dinney ?...

<table border="1"><tbody><tr><td> 시약의 이름 </td><td> 회사 </td><td> 카탈로그 번호 </td></tr><tr><td> Isoflurane (Aerrane) </td><td> 백스터 </td><td> NDC 10019-773-60 </td></tr><tr><td> 29 또는 30 게이지 바늘 </td><td> 해밀턴 </td><td> 7803-06 또는 7803-07 </td></tr><tr><td> 100 microliter 주사기 </td><td> 해밀턴 </td><td> 7656-01 </td></tr></tbody></table>

mouse bladder wall injection

마우스 방광 벽 주입은 orthotopically 줄기 세포, 평활근, 그리고 암 생물학을 포함하여 방광 현상​​을 연구하기위한 유용한 기술입니다. 주사를 시작하기 전에, 수술 지역은 비누와 물로 및 살균 솔루션을 청소해야합니다. 수술 장비는 사용하기 전에 각각의 동물 사이에 소독해야합니다. 각각의 마우스는 흡입 isoflurane 마취 배치 (유도에 대한 2~5%, 유지 보수 1-3 %)와 방광 가위로 복부 정중선 절개하여 노출됩니다. 방광이 가득있다면, 두 손가락 사이 압박에 의해 부분적으로 부드럽게 풀립니다. 관심의 세포 현탁액은 intramurally 29 또는 30 게이지 바늘 1 CC 또는 작은 주사기를 사용하여 방광의 돔 벽에에 주입합니다. 상처는 상처 후 클립과 온난 패드를 복구하는 허용 마우스를 사용 닫힙니다. 방광 벽 주입은 연습 마스터 수있는 민감한 microsurgical 기법입니다.

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

Mouse Bladder Wall Injection

마우스 방광 벽 주입은 orthotopically 방광 줄기 세포와 암 생물학을 연구하는 유용한 방법입니다. 이 섬세한 microsurgical 방법은 신중 기술과 연습 마스터하실 수 있습니다.

마우스 블레더 월 주입

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

23.9K 조회수.  Stanford University School of Medicine. 마우스 방광 벽 주입은 orthotopically 방광 줄기 세포와 암 생물학을 연구하는 유용한 방법입니다. 이 섬세한 microsurgical 방법은 신중 기술과 연습 마스터하실 수 있습니다.

비디오: 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.

Murine 방광 암의 Orthotopic 모델

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

연구

의학

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.

작은 쥐의 Cystometry의 사용 : 방광 Chemosensation의 연구

	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.

Orthotopic 방광 종양 모델 및 Intravesical saRNA 트리 트먼트의 평가

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.

마우스 눈에 유전자 치료 벡터와 줄기 세포의 Subretinal 주입

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-LVR을 사용하여 마음을 실패의 기능에서 왼쪽 심실 벽 스트레스와 개선의 감소

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.

유전자 전달 연구 동소 방광암 모델

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.

쥐과 동 소성 방광 이종 이식의 최소 침습 설립

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

Ultrasound Imaging-guided Intracardiac Injection to Develop a Mouse Model of Breast Cancer Brain Metastases Followed by Longitudinal MRI

Breast cancer brain metastasis, occurring in 30% of breast cancer patients at stage IV, is associated with high mortality. The median survival is only 6 months. It is critical to have suitable animal models to mimic the hemodynamic spread of the metastatic cells in the clinical scenario. Here, we are introducing the use of small animal ultrasound imaging to guide an accurate injection of brain tropical breast cancer cells into the left ventricle of athymic nude mice. Longitudinal MRI is used to assessing intracranial initiation and growth of brain metastases. Ultrasound-guided intracardiac injection ensures not only an accurate injection and hereby a higher successful rate but also significantly decreased mortality rate, as compared to our previous manual procedure. In vivo high resolution MRI allows the visualization of hyperintense multifocal lesions, as small as 310 &#181;m in diameter on T2-weighted images at 3 weeks post injection. Follow-up MRI reveals intracranial tumor growth and increased number of metastases that distribute throughout the whole brain.

A breast cancer brain metastasis mouse model is established with ultrasound imaging-guided intracardiac injection of MDA-MB231/Br-GFP cells. Development of multifocal intracranial metastases has been monitored longitudinally using high-resolution 9.4 T MRI.

종 MRI에 이어 유방암 뇌 전이 마우스 모델을 개발하기 위해 초음파 영상 유도 심장 내 주입

Breast cancer brain metastasis, occurring in 30% of breast cancer patients at stage IV, is associated with high mortality. The median survival is only 6 ...

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.

쥐의 방광 통증 모델로 방광 팽창 유발 내장 운동 응답

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.

초고 해상도 마우스 광학 일관성 단층 촬영 망막 유전자 치료 연구에서 안 구내 주사를 돕기 위해

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