Bu araştırma NIH Intramural programından fon tarafından desteklenmektedir.

I. Hücre Hazırlık <ol><li> Hücreler implante edilmesi her 5 fareler için bir 80% konfluent 225 cm 3 şişeyi yaklaşık% 10 FBS ile DMEM birleştiği yetiştirilmektedir. </li><li> Hücre kapama ortamı aspire; kalsiyum veya magnezyum olmaksızın PBS ile 10 ml ile yıkanır. Aspirat PBS kapalı hücreler. </li><li> Tripsin 3 ml hücreleri Trypsinize, 10-15 dakika bekleyin ve medya 15 ml tripsin nötralize. </li><li> Pipet bir 50 ml konik içine tüm hücreleri ve medya. İki büyük şişeler biri 50 ml konik içine sığacak. </li><li> 4 ° C&#39;de 7 dakika boyunca 1600 rpm&#39;de hücreleri Spin Tamamen süpernatant aspire. Soğuk PBS 10 ml hücre süspansiyon hale getirin ve yavaşça aşağı pipetleme ve hücreleri durulayın. Hücre pelletini PBS ile eşit karıştırılarak emin olun. Süspansiyon bulanık olmalıdır. </li><li> Şu anda bir hücre sayımı coulter sayacı kullanılarak yapılır. Hücreler 4 ° C&#39;de 7 dakika boyunca 1600 rpm&#39;de santrifüje tabi tekrar edilir </li><li> Hücrelerinin 2. santrifüjleme gibi sonraPBS 1 ml korsan tüm PBS ve Pastör pipetiyle. </li><li> Hücreler 4 ° C&#39;de 10 dakika süre ile 3000 rpm&#39;de 1 ml PBS içinde bükülmüş edilir PBS aspire edilir, ve PBS U87 hücreleri için 5 ul başına 1x10 6 ila son konsantrasyon ayarlamak için eklenmektedir, diğer hücre çizgileri optimum tümör büyümesi için hücreleri, farklı sayıda ve konsantrasyon gerektirebilir. </li><li> Son hacim ve konsantrasyon elde edildikten sonra hücreler bir Eppendorf tüpüne buz üzerinde tutulur. </li></ol> II. Xenograftlarında <ol class="ualpha"><li> Farenin hazırlanması <ol><li> Fareler Ketamin / Ksilazin / Tuzlu bir enjeksiyon karışımını kullanarak anestezi uygulanmıştır. Karışım 120 Ketamine, 16 mg / spinal anestezi ve tuzlu su kg mg / kg &#39;dır. Fareler bireysel tartılmış ve ağırlık temelinde 10 g başına 0.1 ml verilmiştir. Anestezi 1 ml şırınga IP bağlı 26G iğne ile verilir. Hiçbir ağrı yanıt için bir uyku safhaya fareler için yaklaşık 15 dakika sürecektir. Bu ağrı yanıtı kontrol edebilirsinizonların haydut sıkıştırarak fareler. </li><li> Fare tamamen uyuşturulduktan sonra, göz merhemi kurumasını engellemek için gözlerini tatbik edilmiştir. </li><li> Fare ağız çubuğundaki çentik ön üst dişler ile stereotaksik yatağına yerleştirilir. Daha sonra oda stereotaksik yerleştirilir. </li><li> Burun koruma baş hareketini önlemek için, ve kulak çubukları hafifçe farelerin kulak kanalları yerleştirmek olan farenin burnunun üzerine yerleştirilir. Kafatası kırık ve / veya nefes almasını engelleyebilir fare gibi, burun çubuğu ya da çok sıkı kulak çubukları ya yerleştirme yapmak için dikkatli olun. </li><li> Fare kafası stereotaksik üzerindeki onay işareti kullanarak seviyesi bu süreçte yararlı olduğundan emin olun. </li><li> Hazırlık için fare, alkol ve providine iyot iyot alkol ve 3 uygulamanın 3 uygulama toplam alternatif farenin başına bolca uygulanır. % 70 olan bir alkol pedi biri nihai uygulama içinkafası. </li></ol></li><li> Cerrahi girişim <ol><li> Prosedüründe tüm enstrümanlar prosedürü başlamadan önce% 70 etanol araçları otoklav veya iliklerine yoluyla sterilize edilmelidir. </li><li> Bir insizyon çapraz geri kafatasının sol arka için sağ göz arkasında kafatası üstüne yapılır. Kesi 10 mm uzunluğunda 15 mm olmalıdır. </li><li> Bir Q-tip kullanarak, yavaş yavaş kafatası maruz ve kafatası ve cilt arasında membran silip için kesi ayırın. Aşırı kanama için herhangi bir kan kurulayın ve izlemek için bir Q-ucu kullanın. </li><li> Bregma görselleştirmek için kafatasının üst Açığa. Hamilton enjektörü noktasını kullanarak, bregma üzerine getirin. Kafatasında açılacak deliğin doğru pozisyonda 2 mm sağ ve bregma (Şekil 2) anterior 1 mm&#39;dir. Steril satın alındığında bir cerrahın keçeli işaretleyici, açılacak deliğinin yeri işaretlemek için kullanılır. Delik bir yakın bulunan unutmayınposterior sütür hattı. </li><li> Sonra, bir 18G iğne ile bir iğne vazo ya da beslenen bir 22G iğne noktası kullanarak işaretli konumda kafatasına küçük bir delik açın. </li><li> Hücrelerin 5 ul ile 10 ul cam Hamilton şırınga yerleştirin. Hamilton enjektörü ile ona bağlı 33G sabit iğneli bir künt 30G ile 10 ul olduğunu. Onları pipetleme ya bağlı bir 25G iğne ile 1 ml şırınga kullanılarak, ya yüklemeden önce hücreleri karıştırmak için emin olun. Hamilton enjektörü yüklendikten sonra, kafatasında delik ile sıraya kolunda geri koyun. </li><li> Iğne kafatası ile seviyesini bir kez, kafatasının yüzeyine iğne bunt sonuna indirin; bu kafatasının yüzeyinin altında 3 mm derinliğe kadar iğne indirin. </li><li> Iğne indirdi sonra, beyin etkileyen iğne herhangi bir majör travma önlemek için 2 dakika için orada kalacaktır. Hücreler implante 6-8 dakika toplam için dakikada 1 ul&#39;lik bir hızda olan, herhangi bir backflo dikkatli olmakw. Hücreler son implante edildikten sonra, bütün hücrelerin in getirilmesi için izin vermek üzere, bir 2 dakika süreyle bir yerde iğne terk </li><li> Beyinden iğne çıkarın ve bir Q-tip ile delik temizleyin. Her bir çözüm ve bu sırada 3x% 50 Actone,% 100 EtOH ve PBS ile iğne ve şırınga temizlemek için devam edin. </li><li> Kafatası üzerinde cilt bu noktada dikilerek üzere hazırdır. 5-0 kullanma PBS yaklaşık 3 dikiş ile kesi kapatın. </li><li> Fare bir ısı yastığı uyanmak için izin ver, ve gerekirse göz merhemi yeniden uygulayın. Kendi kendine hareket gösterir sonra fare kafesine iade edilebilir (yani başını yukarı kaldırılması veya omuzları hareket). </li></ol></li></ol> III. Radyasyon ve Uyuşturucu ile IC implant tedavisi <ol class="ualpha"><li> Radyasyon tedavisi <ol><li> Hücre hattı bağlı olarak, farelerin tedavisi 7 gün post-implant yerine 5 alabilir. Bölümümüz radyasyon tedavi için bir orthovoltage radyoterapi ünitesi kullanırMent. </li><li> Böyle BLI (Biyolüminesens Görüntüleme), MR veya BT gibi görüntüleme öncesinde randomizasyon (şekil 3A) tümör görselleştirmek için kullanılabilir. </li><li> Farelerde önceki gibi Ketamine / Xylazine / salin kokteyli ile anestezi uygulanmıştır, merhem ve göz gözlerine uygulanır. </li><li> Fareler farelerin başları merkezine doğru işaret edildiği bir pasta tekerlek gibi yapılandırılmış bir ölçüye radyasyon jig konumlandırılmıştır. Farelerin organları kurşun radyasyon korumalıdır. </li><li> İlaç tedavisi farklı teslimat yollarının, yani SQ, IP ve IV aracılığıyla önce ya da radyasyon da verilebilir. </li></ol></li></ol> IV. Sonuçlar Fare anestezi enjeksiyonu takiben 45-60 dakika içinde prosedürü uyanmak gerekir. Fare zor işlemden sonra uyanma ve süresi 1 aştı ½ saat işlemden sonra, fare kurtarmak olmayabilir ve ötanazi etkili bir alternatif olabilir. Eğer there aşırı kanama veya prosedür ötanazi sırasında herhangi bir noktada kırık kafatası olur tavsiye edilir. Tüm iyi ve hücre serisine bağlı olarak giderse prosedürü gerçekleşecek olan sonra, bir tümör 2-4 hafta içinde oluşturan başlamalıdır. Intrakraniyal xenograft modelleri kullanırken, terapötik etkinlik genellikle hayatta implantasyon sonrası 5 olarak ölçülür. Ancak, bu gibi bir IC modeli belirti ve bulgular önce ölüm meydana gelir. Fare görülebilir fiziksel belirtileri bazıları bir kambur duruş, vücut ağırlığı kaybı ve hareketsizlik vardır. Nörolojik bulguların bazıları nöbetler, baş eğme ve denge kaybı olabilir. Şekil 3B bir HE ölüm bir tümörün leke gösterir. Bir Kaplan-Meier arsa verileri (Şekil 4) değerlendirmek için kullanılır. % 50 hayatta kalma gün sayısı hesaplanır ve kontrol ile karşılaştırılmıştır. Bileşimi grubu bireysel radyasyon ve ilaç kollarının Ayrıca daha uzun yaşasaydınız kombinasyonu başarılı edilir. <img alt = &quot;Şekil 1&quot; src = &quot;/ files/ftp_upload/3397/3397fig1.jpg&quot; /&gt; Şekil 1. Stoelting Stereotaksik ekipmanları. <img alt="Şekil 2" src="/files/ftp_upload/3397/3397fig2.jpg" /> Şekil 2. Fare kafatasında açılan deliğin yerini şeması. <img alt="Şekil 3A" src="/files/ftp_upload/3397/3397fig3A.jpg" /> GBM implantasyonu sonrası Şekil 3A. BLI görüntüleme. <img alt="Şekil 3B," src="/files/ftp_upload/3397/3397fig3B.jpg" /> Şekil 3B. Ölüm H ve GBM tümör D. <img alt="Şekil 4" src="/files/ftp_upload/3397/3397fig4.jpg" /> Şekil 4. PARP inhibitörü 7 gün sonrası cerrahi ile enjekte U87 IC implantların Sonuçları.

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Biz ifşa hiçbir şey yok.

Gibi glioblastoma multiforme Malign gliom, en sık rastlanan primer beyin tümörleri temsil ve kasvetli bir prognoz 4 var. GBM etkilenen hastaların sağkalım cerrahi, radyasyon ve kemoterapi 5 gelişmelere rağmen (9-12 yani ay sonrası tanı) son on yılda hemen hemen hiç değişmedi. Hücrelerinin, insan gliom histolojik özellikleri mümkün olduğunca yakın benzerlik olmalıdır;; tahmin edilebilir bir büyüme olmalıdır glioma muamele çalışma için uygun bir model özellikleri tekrar...

<table border="1"><tbody><tr><td> Lab Standart Stereotaksik Instrutment </td><td> Fare ve Rat Adaptör Stoelting </td></tr><tr><td> Betadine </td><td> % 100 EtOH </td></tr><tr><td> % 70 EtOH </td><td> % 50 aseton </td></tr><tr><td> 6 &quot;Q-ipuçları </td><td> Alkol swablar </td></tr><tr><td> Hamilton enjektörü 80.008 </td><td> Ethicon 5-0 PD </td></tr><tr><td> 1701 SN 31-33G / 1 &quot;/ PT3 iğne </td><td> 1 ml şırınga </td></tr><tr><td> 25G iğne </td><td> Isıtma pedi </td></tr><tr><td> Suni gözyaşları Merhem </td><td> Çok ince uçlu kalıcı bir kalem </td></tr></tbody></table>

combination radiotherapy in an orthotopic mouse brain tumor model

Glioblastoma multiforme (GBM) 1 en yaygın ve agresif erişkin primer beyin tümörlerdir. Son yıllarda da, tümör mekaniğinin anlaşılması konusunda önemli ilerleme olmuştur ve tümörün doğrudan intraserebral inokülasyonu fizyolojik olarak uygun ortamda 2 invaziv süreci gözlemleme fırsatı sağlar etti. Bildiğim kadarıyla insan beyin tümörleri ile ilgili olarak, mevcut ortotopik modelleri karmaşık bir kraniotomi prosedürü 3 ile tümörün sağlam bir parça hücre süspansiyonları veya implantasyon stereotaksik enjeksiyon yoluyla kurulur. Bizim teknik olarak beynin doğrudan implant kullanılan bir hücre süspansiyonu oluşturmak için doku kültürü hücreleri toplamak. Cerrahi süresi yaklaşık 30 dakika ve fare sabit bir cerrahi düzlemi içinde olması gerekmektedir olarak, enjekte edilebilir bir anestezik kullanılır. Fare Stoetling (Şekil 1) tarafından yapılan bir stereotaksik jig yerleştirilir. Kıçtaer cerrahi alan temizlenir ve hazırlanır, bir kesi yapılır ve bregma kraniotomi yerini belirlemek için yer almaktadır. Kraniotomi yeri sağ 2 mm ve bregma için rostral 1 mm&#39;dir. Derinliği kafatası yüzeyinden 3 mm&#39;dir ve hücreler 2 ul her bir 2 dakika arasında bir hızla enjekte edilir. Cilt 5-0 PDS ile sütüre edilir ve farenin bir ısıtma yastığı uyanmak için izin verilir. Bizim deneyim, hücre serisine bağlı olarak, cerrahi tedavi sonrası 7-10 gün arasında yer alabilir. İlaç dağıtım ilaç bileşimi bağlıdır. Radyasyon tedavisi için fareler anestezi, ve jig yapılan özel koymak. Kurşun farenin vücudu kaplar ve farenin sadece beynin ortaya koyar. Tümörogenez ve GBM için yeni tedavilerin değerlendirilmesi çalışması doğru ve tekrarlanabilir beyin tümörü hayvan modelleri gerektirir. Böylece YÜRÜRLÜK test etmek, beynin mikro ve tümörün etkileşimi incelemek için bu ortotopik beyin modeli kullanınradyasyon olan ve olmayan farklı terapötik ajanların ess.

Watch this Scientific Journal Video about Combination Radiotherapy in an Orthotopic Mouse Brain Tumor Model at JoVE.com

Combination Radiotherapy in an Orthotopic Mouse Brain Tumor Model

Bu makalenin amacı, kemoradyoterapi çalışmaları için ortotopik glioblastoma modelinin kullanımı tanımlamaktır. Bu makale implante, hücre işleme rağmen gitmek ve bir intrakraniyal modeli kullanarak farenin radyoterapi olacaktır.

Bir Ortotopik Fare Beyin Tümörü Modelinde Kombinasyon Radyoterapi

Glioblastoma multiforme (GBM) are the most common and aggressive adult primary brain tumors1. In recent years there has been substantial progress in the ...

combination-radiotherapy-in-an-orthotopic-mouse-brain-tumor-model

Research

JoVE Journal

Medicine

16.3K Görüntüleme.  National Cancer Institute. Bu makalenin amacı, kemoradyoterapi çalışmaları için ortotopik glioblastoma modelinin kullanımı tanımlamaktır. Bu makale implante, hücre işleme rağmen gitmek ve bir intrakraniyal modeli kullanarak farenin radyoterapi olacaktır.

Video: Combination Radiotherapy in an Orthotopic Mouse Brain Tumor Model

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.

Murin Mesane Kanseri bir Ortotopik Model

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

Tıp

Multi-photon Imaging of Tumor Cell Invasion in an Orthotopic Mouse Model of Oral Squamous Cell Carcinoma

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient management strategies. Orthotopic mouse models of oral cancer have been developed to facilitate the study of factors that impact invasion and serve as model system for evaluating anti-tumor therapeutics. In these systems, visualization of disseminated tumor cells within oral cavity tissues has typically been conducted by either conventional histology or with in vivo bioluminescent methods. A primary drawback of these techniques is the inherent inability to accurately visualize and quantify early tumor cell invasion arising from the primary site in three dimensions. Here we describe a protocol that combines an established model for squamous cell carcinoma of the tongue (SCOT) with two-photon imaging to allow multi-vectorial visualization of lingual tumor spread. The OSC-19 head and neck tumor cell line was stably engineered to express the F-actin binding peptide LifeAct fused to the mCherry fluorescent protein (LifeAct-mCherry). Fox1nu/nu mice injected with these cells reliably form tumors that allow the tongue to be visualized by ex-vivo application of two-photon microscopy. This technique allows for the orthotopic visualization of the tumor mass and locally invading cells in excised tongues without disruption of the regional tumor microenvironment. In addition, this system allows for the quantification of tumor cell invasion by calculating distances that invaded cells move from the primary tumor site. Overall this procedure provides an enhanced model system for analyzing factors that contribute to SCOT invasion and therapeutic treatments tailored to prevent local invasion and distant metastatic spread. This method also has the potential to be ultimately combined with other imaging modalities in an in vivo setting.

A comprehensive overview of the techniques involved in generating a mouse model of oral cancer and quantitative monitoring of tumor invasion within the tongue through multi-photon microscopy of labeled cells is presented. This system can serve as a useful platform for the molecular assessment and drug efficacy of anti-invasive compounds.

Oral Skuamöz Hücre Karsinomu Ortotopik Fare Modeli Tümör Hücre Invasion Multi-foton Görüntüleme

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient ...

In vivo Bioluminescence Imaging of Tumor Hypoxia Dynamics of Breast Cancer Brain Metastasis in a Mouse Model

It is well recognized that tumor hypoxia plays an important role in promoting malignant progression and affecting therapeutic response negatively. There is little knowledge about in situ, in vivo, tumor hypoxia during intracranial development of malignant brain tumors because of lack of efficient means to monitor it in these deep-seated orthotopic tumors. Bioluminescence imaging (BLI), based on the detection of light emitted by living cells expressing a luciferase gene, has been rapidly adopted for cancer research, in particular, to evaluate tumor growth or tumor size changes in response to treatment in preclinical animal studies. Moreover, by expressing a reporter gene under the control of a promoter sequence, the specific gene expression can be monitored non-invasively by BLI. Under hypoxic stress, signaling responses are mediated mainly via the hypoxia inducible factor-1&alpha; (HIF-1&alpha;) to drive transcription of various genes. Therefore, we have used a HIF-1&alpha; reporter construct, 5HRE-ODD-luc, stably transfected into human breast cancer MDA-MB231 cells (MDA-MB231/5HRE-ODD-luc). In vitro HIF-1&alpha; bioluminescence assay is performed by incubating the transfected cells in a hypoxic chamber (0.1% O2) for 24 hr before BLI, while the cells in normoxia (21% O2) serve as a control. Significantly higher photon flux observed for the cells under hypoxia suggests an increased HIF-1&alpha; binding to its promoter (HRE elements), as compared to those in normoxia. Cells are injected directly into the mouse brain to establish a breast cancer brain metastasis model. In vivo bioluminescence imaging of tumor hypoxia dynamics is initiated 2 wks after implantation and repeated once a week. BLI reveals increasing light signals from the brain as the tumor progresses, indicating increased intracranial tumor hypoxia. Histological and immunohistochemical studies are used to confirm the in vivo imaging results. Here, we will introduce approaches of in vitro HIF-1&alpha; bioluminescence assay, surgical establishment of a breast cancer brain metastasis in a nude mouse and application of in vivo bioluminescence imaging to monitor intracranial tumor hypoxia.

In vivo Bioluminescence Imaging of Tumor Hypoxia Dynamics of Breast Cancer Brain Metastasis in a Mouse Model

Bioluminescence imaging of hypoxia inducible factor-1&alpha; activity is applied to monitor intracranial tumor hypoxia development in a breast cancer brain metastasis mouse model.

Meme Kanseri Beyin metastaz tümör Hipoksi Dynamics in vivo Biyoparlaklık Görüntüleme Fare Modeli

It is well recognized that tumor hypoxia plays an important role in promoting malignant progression and affecting therapeutic response negatively. There ...

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.

Bir Ortotopik Mesane Tümörü Model ve intravezikal Sarna Tedavisinin Değerlendirilmesi

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

An Orthotopic Mouse Model of Anaplastic Thyroid Carcinoma

Several types of animal models of human thyroid carcinomas have been established, including subcutaneous xenograft and orthotopic implantation of cancer cells into immunodeficient mice. Subcutaneous xenograft models have been valuable for preclinical screening and evaluation of new therapeutic treatments. There are a number of advantages to using a subcutaneous model; 1) rapid, 2) reproducible, and 3) tumor establishment, growth, and response to therapeutic agents may be monitored by visual inspection. However, substantial evidence has shed light on the short-comings of subcutaneous xenograft models1-3. For instance, medicinal treatments demonstrating curative properties in subcutaneous xenograft models often have no notable impact on the human disease. The microenvironment of the site of xenographic transplantation or injection lies at the heart of this dissimilarity.
Orthotopic tumor xenograft models provide a more biologically relevant context in which to study the disease. The advantages of implanting diseased cells or tissue into their anatomical origin equivalent within a host animal includes a suitable site for tumor-host interactions, development of disease-related metastases and the ability to examine site-specific influence on investigational therapeutic remedies. Therefore, orthotopic xenograft models harbor far more clinical value because they closely reproduce human disease. For these reasons, a number of groups have taken advantage of an orthotopic thyroid cancer model as a research tool4-7.
Here, we describe an approach that establishes an orthotopic model for the study of anaplastic thyroid carcinoma (ATC), which is highly invasive, resists treatment, and is virtually fatal in all diagnosed patients. Cultured ATC cells are prepared as a dissociated cellular suspension in a solution containing a basement membrane matrix. A small volume is slowly injected into the right thyroid gland. Overall appearance and health of the mice are monitored to ensure minimal post-operative complications and to gauge pathological penetrance of the cancer. Mice are sacrificed at 4 weeks, and tissue is collected for histological analysis. Animals may be taken at later time-points to examine more advance progression of the disease. Production of this orthotopic mouse model establishes a platform that accomplishes two objectives: 1) further our understanding of ATC pathology, and 2) screen current and future therapeutic agents for efficacy in combating ATC.

Generation of an orthotopic mouse model of anaplastic thyroid carcinoma is described here. This technique employs surgical placement of human anaplastic thyroid cancer cells into the thyroid of immunodeficient mice, thus creating a more clinically relevant setting to study disease progression as well as screen innovative therapeutic interventions.

Anaplastik Tiroid Kanseri Ortotopik Fare Modeli

Several types of animal models of human thyroid carcinomas have been established, including subcutaneous xenograft and orthotopic implantation of cancer ...

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.

Gene Teslim Çalışmaları Ortotopik Mesane Kanseri Modeli

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

A Novel High-resolution In vivo Imaging Technique to Study the Dynamic Response of Intracranial Structures to Tumor Growth and Therapeutics

We have successfully integrated previously established Intracranial window (ICW) technology 1-4 with intravital 2-photon confocal microscopy to develop a novel platform that allows for direct long-term visualization of tissue structure changes intracranially. Imaging at a single cell resolution in a real-time fashion provides supplementary dynamic information beyond that provided by standard end-point histological analysis, which looks solely at 'snap-shot' cross sections of tissue.
Establishing this intravital imaging technique in fluorescent chimeric mice, we are able to image four fluorescent channels simultaneously. By incorporating fluorescently labeled cells, such as GFP+ bone marrow, it is possible to track the fate of these cells studying their long-term migration, integration and differentiation within tissue. Further integration of a secondary reporter cell, such as an mCherry glioma tumor line, allows for characterization of cell:cell interactions. Structural changes in the tissue microenvironment can be highlighted through the addition of intra-vital dyes and antibodies, for example CD31 tagged antibodies and Dextran molecules.
Moreover, we describe the combination of our ICW imaging model with a small animal micro-irradiator that provides stereotactic irradiation, creating a platform through which the dynamic tissue changes that occur following the administration of ionizing irradiation can be assessed.
Current limitations of our model include penetrance of the microscope, which is limited to a depth of up to 900 &mu;m from the sub cortical surface, limiting imaging to the dorsal axis of the brain. The presence of the skull bone makes the ICW a more challenging technical procedure, compared to the more established and utilized chamber models currently used to study mammary tissue and fat pads 5-7. In addition, the ICW provides many challenges when optimizing the imaging.

A Novel High-resolution In vivo Imaging Technique to Study the Dynamic Response of Intracranial Structures to Tumor Growth and Therapeutics

We describe a novel in vivo imaging technique that couples fluorescent chimeric mice with intracranial windows and high-resolution 2-photon microscopy. This imaging platform aids studies of dynamic changes in brain tissue and microvasculature, at a single-cell level, following pathological insults and is adaptable to assess intracranial drug delivery and distribution.

Yeni Bir Yüksek çözünürlüklü<em&gt; In vivo</emTümör Büyüme ve Tedavi için İntrakranial Yapıların Dinamik Tepki Eğitim&gt; Görüntüleme Tekniği

We have successfully integrated previously established Intracranial window (ICW) technology 1-4 with intravital 2-photon confocal microscopy to develop a ...

An Orthotopic Murine Model of Human Prostate Cancer Metastasis

Our laboratory has developed a novel orthotopic implantation model of human prostate cancer (PCa). As PCa death is not due to the primary tumor, but rather the formation of distinct metastasis, the ability to effectively model this progression pre-clinically is of high value. In this model, cells are directly implanted into the ventral lobe of the prostate in Balb/c athymic mice, and allowed to progress for 4-6 weeks. At experiment termination, several distinct endpoints can be measured, such as size and molecular characterization of the primary tumor, the presence and quantification of circulating tumor cells in the blood and bone marrow, and formation of metastasis to the lung. In addition to a variety of endpoints, this model provides a picture of a cells ability to invade and escape the primary organ, enter and survive in the circulatory system, and implant and grow in a secondary site. This model has been used effectively to measure metastatic response to both changes in protein expression as well as to response to small molecule therapeutics, in a short turnaround time.

This orthotopic model of human prostate cancer allows for quantification of tumor size, circulating tumor cells, and formation of distinct metastasis to the lung. As cells must escape the primary organ, enter the blood stream, and implant into a secondary site, this model effectively recapitulates the scenario in humans.

İnsan Prostat Kanser metastaz Ortotopik fare modeli

Our laboratory has developed a novel orthotopic implantation model of  human prostate cancer (PCa). As PCa death is not due to the primary tumor, but  ...

An Orthotopic Glioblastoma Mouse Model Maintaining Brain Parenchymal Physical Constraints and Suitable for Intravital Two-photon Microscopy

Glioblastoma multiforme (GBM) is the most aggressive form of brain tumors with no curative treatments available to date.
Murine models of this pathology rely on the injection of a suspension of glioma cells into the brain parenchyma following incision of the dura-mater. Whereas the cells have to be injected superficially to be accessible to intravital two-photon microscopy, superficial injections fail to recapitulate the physiopathological conditions. Indeed, escaping through the injection tract most tumor cells reach the extra-dural space where they expand abnormally fast in absence of mechanical constraints from the parenchyma.
Our improvements consist not only in focally implanting a glioma spheroid rather than injecting a suspension of glioma cells in the superficial layers of the cerebral cortex but also in clogging the injection site by a cross-linked dextran gel hemi-bead that is glued to the surrounding parenchyma and sealed to dura-mater with cyanoacrylate. Altogether these measures enforce the physiological expansion and infiltration of the tumor cells inside the brain parenchyma. Craniotomy was finally closed with a glass window cemented to the skull to allow chronic imaging over weeks in absence of scar tissue development.
Taking advantage of fluorescent transgenic animals grafted with fluorescent tumor cells we have shown that the dynamics of interactions occurring between glioma cells, neurons (e.g. Thy1-CFP mice) and vasculature (highlighted by an intravenous injection of a fluorescent dye) can be visualized by intravital two-photon microscopy during the progression of the disease.
The possibility to image a tumor at microscopic resolution in a minimally compromised cerebral environment represents an improvement of current GBM animal models which should benefit the field of neuro-oncology and drug testing.

We have established a cortical orthotopic glioblastoma model in mice for intravital two-photon microscopy that recapitulates the biophysical constraints normally at play during the growth of the tumor. A chronic glass window replacing the skull above the tumor enables the follow-up of the tumor progression over time by two-photon microscopy.

Ortotopik Glioblastoma Fare Modeli Beyin parankim fiziksel kısıtlamaları bakımı ve Intravital İki foton Mikroskopi için uygundur

Glioblastoma multiforme (GBM) is the most aggressive form of brain tumors with no curative treatments available to date.
Murine models of this pathology ...

Tumor Treating Field Therapy in Combination with Bevacizumab for the Treatment of Recurrent Glioblastoma

A novel device that employs TTF therapy has recently been developed and is currently in use for the treatment of recurrent glioblastoma (rGBM). It was FDA approved in April 2011 for the treatment of patients 22 years or older with rGBM. The device delivers alternating electric fields and is programmed to ensure maximal tumor cell kill1.
Glioblastoma is the most common type of glioma and has an estimated incidence of approximately 10,000 new cases per year in the United States alone2. This tumor is particularly resistant to treatment and is uniformly fatal especially in the recurrent setting3-5. Prior to the approval of the TTF System, the only FDA approved treatment for rGBM was bevacizumab6. Bevacizumab is a humanized monoclonal antibody targeted against the vascular endothelial growth factor (VEGF) protein that drives tumor angiogenesis7. By blocking the VEGF pathway, bevacizumab can result in a significant radiographic response (pseudoresponse), improve progression free survival and reduce corticosteroid requirements in rGBM patients8,9. Bevacizumab however failed to prolong overall survival in a recent phase III trial26. A pivotal phase III trial (EF-11) demonstrated comparable overall survival between physicians&#8217; choice chemotherapy and TTF Therapy but better quality of life were observed in the TTF arm10.
There is currently an unmet need to develop novel approaches designed to prolong overall survival and/or improve quality of life in this unfortunate patient population. One appealing approach would be to combine the two currently approved treatment modalities namely bevacizumab and TTF Therapy. These two treatments are currently approved as monotherapy11,12, but their combination has never been evaluated in a clinical trial. We have developed an approach for combining those two treatment modalities and treated 2 rGBM patients. Here we describe a detailed methodology outlining this novel treatment protocol and present representative data from one of the treated patients.

A novel methodology that is employed for the treatment of recurrent glioblastomas is described. This treatment approach employs the application of alternating electric tumor treating fields (TTFields), known as TTF Therapy in combination with bevacizumab, a targeted agent that is currently FDA approved as monotherapy.

Tekrarlayan Glioblastoma&#39;nın Tedavisinde Bevacizumab ile Kombine Alan Terapi Tedavisi Tümör

A novel device that employs TTF therapy has recently been developed and is currently in use for the treatment of recurrent glioblastoma (rGBM). It was FDA ...

Bir Ortotopik Fare Beyin Tümörü Modelinde Kombinasyon Radyoterapi

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