Biz Harold Varmus, Brian C. Lewis, Douglas Hanahan, Danny Huang, Sharon Pang, Megan Wong ve Manasi M. Godbole teşekkür ederim. Y.C.N.D. DOD grant W81XWH-16-1-0619 ve NIH grant 1R01CA204916 tarafından desteklenir.

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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+crest-directed+gene+transfer+demonstrates+Wnt1+role+in+melanocyte+expansion+and+differentiation+during+mouse+development.">Neural crest-directed gene transfer demonstrates Wnt1 role in melanocyte expansion and differentiation during mouse development.</a> Proc Natl Acad Sci USA. 97 (18), 10050-10055 (2000).</li><li>Loftus, S. K., Larson, D. M., Watkins-Chow, D., Church, D. M., Pavan, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+RCAS+vectors+useful+for+functional+genomic+analyses.">Generation of RCAS vectors useful for functional genomic analyses.</a> DNA Res. 8 (5), 221-226 (2001).</li><li>Bromberg-White, J. 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T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+PTEN-regulating+microRNA+miR-26a+is+amplified+in+high-grade+glioma+and+facilitates+gliomagenesis+in+vivo.">The PTEN-regulating microRNA miR-26a is amplified in high-grade glioma and facilitates gliomagenesis in vivo.</a> Genes Dev. 23 (11), 1327-1337 (2009).</li><li>Ahronian, L. G., Lewis, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+high-titer+RCAS+virus+from+DF1+chicken+fibroblasts.">Generation of high-titer RCAS virus from DF1 chicken fibroblasts.</a> Cold Spring Harb Protoc. 2014 (11), 1161-1166 (2014).</li><li>Green, M. 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Yazarlar ifşa gerek yok.

Bu çalışmada, bir güçlü fare modeli, RIP-Tag açıkladığımız; RIP-tva, yoluyla kimlik ve metastatik faktörler karakterizasyonu için kuş Retrovirüsler somatik gen teslim elde etmek için. Her ne kadar RIP-etiket; RIP-tva fareler geliştirmek pankreas nöroendokrin tümör, bu fare modelde tanımlanan metastatik faktörler de metastaz diğer kanser türleri teşvik.
Bizim yaklaşım somatik genetik değişiklikler özellikle premalign lezyonlar pankreas β hücrel...

En kanserleri somatik mutasyon 1ile ortaya çıkmaktadır. Geleneksel genetiği fare modelleri (et) tumorigenesis 2için belirli genetik değişiklik katkısını önemli anlayışlar hazırladık. Ancak, bazı sınırlamaları vardır. Bu modellerin en büyük dezavantajı sadece bir doku bazı hücrelerde genetik değişiklikler elde insanlarda tümör oluşumu sporadik doğası çoğaltılmaz var. Transgenik ve nakavt fareler mutasyonların da germline geliştirme etkiler için potansiyeli vardır. Ayrıca, bu fare modelleri üreten pahalı ve zaman alıcı olduğunu.
Metastaz kanser alanında önemli bir konudur. Metastaz modelleme et içinde zor olmuştur. Spontan metastaz fare nadirdir. Alellerle değişken ve gecikme süresi uzun metastaz 3et içinde. Metastatik cascade erken adımda ortadan kalkar böylece deneysel metastaz modelleri fareler, dolaşımı içine direkt enjeksiyon hücre istihdam.
Bazı fare modelleri metastatik faktörler eğitim yukarıdaki sınırlamaları aşmak için bir bitransgenic fare modeli, RIP-Tag geliştirdik; RIP-tva4. Strateji alt grubu-A kuş lökozu virüs, tva 6,7için bir son derece senkronize tümör ilerleme fare modeli, RIP-etiket 5ve reseptör kullanımını birleştirerek üzerinde temel alır. Bu RIP-etiket; RIP-tvafare modeli gen somatically bir tek bitransgenic fare yük sunulmasına olanak sağlar. Rb ve p53 Tümör baskılayıcı fonksiyonlarını baskılayarak SV40 T antijen ile fareler Hiperplazi, anjiogenez, adenoma ve invaziv karsinom gibi aşamaları ile pankreas nöroendokrin tümör insan tumorigenesis benzer bir şekilde geliştirin. Bu RIP-Tag model kanser, bunlarla sınırlı olmamak için pankreas nöroendokrin tümör işaretlerinden anlayışımız için çok öğretici oldu. Preklinik denemeler 8' de kullanılmaktadır.
Biz kuş Retrovirüsler intracardiacally enjeksiyonuyla RIP-etiket içine somatik gen transferi için bir iletişim kuralı mevcut; RIP-tva fareler. Kuş Retrovirüsler RCASBP elde edilen başarılı enfeksiyonu aktif Proliferasyona hedef hücrelerin gerekli. Bu nedenle, RIP-Tag seçti; RIP-tva fareler, 7 hafta-in yaş, ne zaman Hiperplazi pankreas adacıkları yaklaşık % 50 geliştirir. Sol ventrikül İntrakardiyak enjeksiyon yüksek titresi virüslerin % 10-20 4bir enfeksiyon verimlilik elde etmek için gereklidir. Virüs pankreas adacıkları ulaşmadan bu teslim yöntemi dolaşım içinde viral parçacıkların önemli seyreltme azaltır.
Bu yaklaşımı kullanarak, daha önce Bcl-xL kanser metastaz onun anti-apoptotik işlev 4,9/ bağımsız teşvik göstermiştir. BCL-xL RIP-etiket tumorigenic Ontojeni boyunca tüm pankreas β hücrelerindeki bir transgene ile ifade edildi zaman bu anti-apoptotik-bağımsız metastatik işlev gözlenen değil; RIP-Bcl-xL fare modeli 10. Bu nedenle, bizim fare modeli tanımlamak ve tumorigenesis daha sonraki bir aşamada ifade genler işlevleri tanımlamak için eşsiz bir fırsat sunmaktadır. Çünkü adacıkları % 2-4'ü geliştirmek tümörler RIP-Tag; içinde içine RIP-tva bitransgenic fareler viral enfeksiyon ve tüm premalign lezyonlar olmadan RCASBP elde edilen kuş Retrovirüsler ile enfekte, sadece tumorigenesis doğal seyri üzerinde seçici bir avantaj görüşmek faktörler belirlenebilir. Özellikle, pankreas lenf düğümleri veya diğer organlara metastaz normalde RIP-Tag oluşmaz çünkü metastatik faktörler en kolay bu yöntem tarafından tanınır; RIP-tva fareler.

<table border="0" cellpadding="0" cellspacing="0" width="747">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">RCASBP-Y DV plasmid</td>
			<td style="width:123px;">Addgene</td>
			<td style="width:119px;">11478</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">RCAS-RNAi plasmid</td>
			<td style="width:123px;">Addgene</td>
			<td style="width:119px;">15182</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">DMEM</td>
			<td style="width:123px;">Corning</td>
			<td>10-013-CV</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">fetal bovine serum</td>
			<td>Atlanta Biologicals</td>
			<td>25-005-CI</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">L-glutamine, 100x</td>
			<td style="width:123px;">Corning</td>
			<td>25-005-CI</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Penicillin-Streptomycin solution, 100x</td>
			<td style="width:123px;">Corning</td>
			<td>30-002-CI</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">PBS-/-, 1X</td>
			<td style="width:123px;">Corning</td>
			<td>21-040-CV</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Superfect</td>
			<td>Qiagen</td>
			<td>301305</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Polyallomer centrifuge tube</td>
			<td>Beckman Coulter</td>
			<td>326823</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">0.45 mm Nalgene 
			Syringe Filters with PES Membrane</td>
			<td style="width:123px;">Thermo Scientific</td>
			<td style="width:119px;">194-2545</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Insulin Syringes&#160;</td>
			<td style="width:123px;">BD</td>
			<td>329461</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">synaptophysin</td>
			<td style="width:123px;">Vector Laboratories</td>
			<td>VP-S284</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">VECTASTAIN Elite ABC HRP Kit (Peroxidase, Rabbit IgG)&#160;</td>
			<td style="width:123px;">Vector Laboratories</td>
			<td>PK-6101</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">AmpliTaq DNA Polymerase with Buffer II</td>
			<td>Life Technologies</td>
			<td>N8080153</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MyTaq DNA Polymerase</td>
			<td>Bioline</td>
			<td>BIO-21106</td>
			<td></td>
		</tr>
	</tbody>
</table>

identification and characterization of metastatic factors by gene transfer into the novel rip-tag; rip-tva murine model

Metastatik kanser ölümleri solid tümör olan hastalarda yüzde 90'ını hesapları. Kanser metastaz sürücüleri daha iyi anlamak için ve yeni tedavi hedefleri belirlemek için acil bir ihtiyaç vardır. İlerleme birincil kanser metastaz için sürücü moleküler olayları araştırmak için bir bitransgenic fare modeli, RIP-Tag geliştirdik; RIP-tva. Bu fare modelinde, sıçan insülin organizatörü (RIP) pankreas beta hücrelerinde SV40 T antijen (etiket) ve reseptör alt grubu bir kuş lökozu virüs (tva) için ifade kullanıyor. Fareler pankreas nöroendokrin tümörlerin % 100 alellerle Hiperplazi, anjiogenez, adenoma ve invaziv karsinom gibi aşamaları ile insan tumorigenesis için benzer iyi tanımlanmış aşamalarla ile geliştirmek. Çünkü RIP-etiket; RIP-tva fareler metastatik hastalık geliştirmek değil, metastaz teşvik genetik değişikliklere kolayca belirlenebilir. Somatik gen aktarım tva ifade, pankreas β Proliferasyona premalign lezyonlar elde edilir kuş Retrovirüsler istenen genetik değişikliği barındıran İntrakardiyak sızdırma yoluyla. Bir titresi &#62; ml başına 1 x 108 bulaşıcı birim vivo içinde enfeksiyon için uygun kabul edilir. Buna ek olarak, kuş Retrovirüsler tümörler RIP-etiket içinde türetilmiş hücre hatları bulaşabilir; RIP-tva fareler yüksek verimlilik ile. Hücre hatları metastatik faktörler karakterize etmek için de kullanılabilir. Burada nasıl bu fare modeli kullanmak ve tümör metastazı aday genler işlevleri değerlendirmek için satırları hücre göstermektedir.

Vivo ve in vitro enfeksiyon oranı RIP-etiket; RIP-tva tümör hücreleri RCASBP tabanlı virüs tarafından 20~ % 20 ve % 80 ~ sırasıyla vardır. RIP-etiket; RIP-tva fare modeli, her fare 400 pankreas adacıkları yaklaşık % 4'ü doğal olarak geliştirmek tümörler 20; Bu nedenle tümör hücreleri her fare virüsler tarafından teslim genlerin potansiyel etkisinin histolojik ve fenotipik analiz iç...

Watch this Scientific Journal Video about Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel RIP-Tag; RIP-tva Murine Model at JoVE.com

Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel RIP-Tag; RIP-tva Murine Model

Biz bir roman somatik gen aktarım sistem RIP-etiket kullanmak göstermek için bir iletişim kuralı mevcut; RIP-tva metastaz genlerinde fonksiyonu çalışmaya fare modeli. Kuş Retrovirüsler gen transferi önceden Malign, invaziv olmayan lezyonlar yetişkin farelerde pankreas β hücrelerinin içine sağlamak için intracardiacally teslim edilir.

Kimlik ve metastatik faktörler gen transferi ile karakterizasyonu roman RIP-etiketinin içine; RIP-tva fare modeli

Metastatic cancer accounts for 90% of deaths in patients with solid tumors. There is an urgent need to better understand the drivers of cancer metastasis ...

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Cancer Research

Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel RIP-Tag; RIP-tva Murine Model

8.7K Görüntüleme.  Weill Cornell Medicine. Biz bir roman somatik gen aktarım sistem RIP-etiket kullanmak göstermek için bir iletişim kuralı mevcut; RIP-tva metastaz genlerinde fonksiyonu çalışmaya fare modeli. Kuş Retrovirüsler gen transferi önceden Malign, invaziv olmayan lezyonlar yetişkin farelerde pankreas β hücrelerinin içine sağlamak için intracardiacally teslim edilir.

Video: Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel RIP-Tag; RIP-tva Murine Model

Advanced Animal Model of Colorectal Metastasis in Liver: Imaging Techniques and Properties of Metastatic Clones

Patients with a limited number of hepatic metastases and slow rates of progression can be successfully treated with local treatment approaches1,2. However, little is known about the heterogeneity of liver metastases, and animal models capable of evaluating the development of individual metastatic colonies are needed. Here, we present an advanced model of hepatic metastases that provides the ability to quantitatively visualize the development of individual tumor clones in the liver and estimate their growth kinetics and colonization efficiency. We generated a panel of monoclonal derivatives of HCT116 human colorectal cancer cells stably labeled with luciferase and tdTomato and possessing different growth properties. With a splenic injection followed by a splenectomy, the majority of these clones are able to generate hepatic metastases, but with different frequencies of colonization and varying growth rates. Using the In Vivo&#160;Imaging System (IVIS), it is possible to visualize and quantify metastasis development with in vivo luminescent and ex vivo fluorescent imaging. In addition, Diffuse Luminescent Imaging Tomography (DLIT) provides a 3D distribution of liver metastases in vivo. Ex vivo fluorescent imaging of harvested livers provides quantitative measurements of individual hepatic metastatic colonies, allowing for the evaluation of the frequency of liver colonization and the growth kinetics of metastases. Since the model is similar to clinically observed liver metastases, it can serve as a modality for detecting genes associated with liver metastasis and for testing potential ablative or adjuvant treatments for liver metastatic disease.

The ability of metastatic clones to colonize distant sites depends on their proliferation capacity and/or their ability to survive in the host microenvironment without significant proliferation. Here, we present an animal model that allows quantitative visualization of both types of liver colonization by metastatic clones.

İleri Hayvan Karaciğer Kolorektal Metastaz Model: Görüntüleme Teknikleri ve Metastatik Klonların Özellikleri

Patients with a limited number of hepatic metastases and slow rates of progression can be successfully treated with local treatment approaches1,2. ...

Kanser Araştırmaları

A Syngeneic Mouse Model of Metastatic Renal Cell Carcinoma for Quantitative and Longitudinal Assessment of Preclinical Therapies

Renal cell carcinoma (RCC) affects &#62; 60,000 people in the United States annually, and ~ 30% of RCC patients have multiple metastases at the time of diagnosis. Metastatic RCC (mRCC) is incurable, with a median survival time of only 18 months. Immune-based interventions (e.g., interferon (IFN) and interleukin (IL)-2) induce durable responses in a fraction of mRCC patients, and multikinase inhibitors (e.g., sunitinib or sorafenib) or anti-VEGF receptor monoclonal antibodies (mAb) are largely palliative, as complete remissions are rare. Such shortcomings in current therapies for mRCC patients provide the rationale for the development of novel treatment protocols. A key component in the preclinical testing of new therapies for mRCC is a suitable animal model. Beneficial features that recapitulate the human condition include a primary renal tumor, renal tumor metastases, and an intact immune system to investigate any therapy-driven immune effector responses and the formation of tumor-induced immunosuppressive factors. This report describes an orthotopic mRCC mouse model that has all of these features. We describe an intrarenal implantation technique using the mouse renal adenocarcinoma cell line Renca, followed by the assessment of tumor growth in the kidney (primary site) and lungs (metastatic site).

Implementation of an orthotopic model of renal cell carcinoma in immunocompetent mice affords the investigator a clinically-relevant system defined by the presence of a primary renal tumor and lung metastases in the same animal. This system can be used to preclinically test a variety of treatments in vivo.

Preklinik Tedaviler Nicel ve Boyuna Değerlendirme A sinjeneik Fare Modeli Yapmış Bir Renal Hücreli Karsinom

Renal cell carcinoma (RCC) affects &#62; 60,000 people in the United States annually, and ~ 30% of RCC patients have multiple metastases at the time of ...

A Mouse Model of Fatigue Induced by Peripheral Irradiation

Cancer-related fatigue (CRF) is a distressing and costly condition that often affects patients receiving cancer treatments, including radiation therapy. Here we describe a method using targeted peripheral irradiation to induce fatigue-like behavior in mice. With appropriate shielding, the irradiation targets the lower abdominal/pelvic region of the mouse, sparing the brain, in an effort to model radiation treatment received by individuals with pelvic cancers. We deliver an irradiation dose that is sufficient to induce fatigue-like behavior in mice, measured by voluntary wheel-running activity (VWRA), without causing obvious morbidity. Since wheel running is a normal, voluntary behavior in mice, its use should have little confounding effect on other behavioral tests or biological measures. Hence, wheel running can be used as a feasible outcome measure in understanding the behavioral and biological correlates of fatigue. CRF is a complex condition with frequent comorbidities, and likely has causes related both to cancer and its various treatments. The methods described in this paper are useful for investigating radiation-induced changes that contribute to the development of CRF and, more generally, to explore the biological networks that can explain the development and persistence of a peripherally-triggered but centrally-driven behavior like fatigue.

We describe a method using targeted peripheral irradiation to induce fatigue-like behavior in mice. The selected non-lethal irradiation dose leads to a week-long reduction in voluntary wheel-running activity.

Periferik Radyasyonla Bağlı Yorgunluk A Fare Modeli

Cancer-related fatigue (CRF) is a distressing and costly condition that often affects patients receiving cancer treatments, including radiation therapy. ...

Fabrication of Tongue Extracellular Matrix and Reconstitution of Tongue Squamous Cell Carcinoma In Vitro

In order to construct an effective and realistic model for tongue squamous cell carcinoma (TSCC) in vitro, the methods were created to produce decellularized tongue extracellular matrix (TEM) which provides functional scaffolds for TSCC construction. TEM provides an in vitro niche for cell growth, differentiation, and cell migration. The microstructures of native extracellular matrix (ECM) and biochemical compositions retained in the decellularized matrix provide tissue-specific niches for anchoring cells. The fabrication of TEM can be realized by deoxyribonuclease (DNase) digestion accompanied with a serious of organic or inorganic pretreatment. This protocol is easy to operate and ensures high efficiency for the decellularization. The TEM showed favorable cytocompatibility for TSCC cells under static or stirred culture conditions, which enables the construction of the TSCC model. A self-made bioreactor was also used for the persistent stirred condition for cell culture. Reconstructed TSCC using TEM showed the characteristics and properties resembling clinical TSCC histopathology, suggesting the potential in TSCC research.

Fabrication of Tongue Extracellular Matrix and Reconstitution of Tongue Squamous Cell Carcinoma In Vitro

A method is shown here for the preparation of the tongue extracellular matrix (TEM) with efficient decellularization. The TEM can be used as functional scaffolds for the reconstruction of a tongue squamous cell carcinoma (TSCC) model under static or stirred culture conditions.

Dil hücre dışı matriks ve dil Skuamöz Hücre Karsinomu Vitro sulandırma imalatı

In order to construct an effective and realistic model for tongue squamous cell carcinoma (TSCC) in vitro, the methods were created to produce ...

A Bioluminescent and Fluorescent Orthotopic Syngeneic Murine Model of Androgen-dependent and Castration-resistant Prostate Cancer

Orthotopic tumor modeling is a valuable tool for pre-clinical prostate cancer research, as it has multiple advantages over both subcutaneous and transgenic genetically engineered mouse models. Unlike subcutaneous tumors, orthotopic tumors contain more clinically accurate vasculature, tumor microenvironment, and responses to multiple therapies. In contrast to genetically engineered mouse models, orthotopic models can be performed with lower cost and in less time, involve the use of highly complex and heterogeneous mouse or human cancer cell lines, rather that single genetic alterations, and these cell lines can be genetically modified, such as to express imaging agents. Here, we present a protocol to surgically injecting a luciferase- and mCherry-expressing murine prostate cancer cell line into the anterior prostate lobe of mice. These mice developed orthotopic tumors that were non-invasively monitored in vivo and further analyzed for tumor volume, weight, mouse survival, and immune infiltration. Further, orthotopic tumor-bearing mice were surgically castrated, leading to immediate tumor regression and subsequent recurrence, representing castration-resistant prostate cancer. Although technical skill is required to carry out this procedure, this syngeneic orthotopic model of both androgen-dependent and castration-resistant prostate cancer is of great use to all investigators in the field.

The goal of this protocol is to demonstrate the intra-prostatic injection of prostate cancer cells, with subsequent castration. Orthotopic pre-clinical models of androgen-dependent and castration-resistant prostate cancer are critical to study the disease in the context of a clinically relevant tumor microenvironment and an immunocompetent host.

Kollarındaki ve floresan Orthotopic Syngeneic fare modeli androjen bağlı ve kısırlaştırma dayanıklı prostat kanseri

Orthotopic tumor modeling is a valuable tool for pre-clinical prostate cancer research, as it has multiple advantages over both subcutaneous and ...

Virus Delivery of CRISPR Guides to the Murine Prostate for Gene Alteration

With an increasing incidence of prostate cancer, identification of new tumor drivers or modulators is crucial. Genetically engineered mouse models (GEMM) for prostate cancer are hampered by tumor heterogeneity and its complex microevolution dynamics. Traditional prostate cancer mouse models include, amongst others, germline and conditional knockouts, transgenic expression of oncogenes, and xenograft models. Generation of de novo mutations in these models is complex, time-consuming, and costly. In addition, most of traditional models target the majority of the prostate epithelium, whereas human prostate cancer is well known to evolve as an isolated event in only a small subset of cells. Valuable models need to simulate not only prostate cancer initiation, but also progression to advanced disease.
Here we describe a method to target a few cells in the prostate epithelium by transducing cells by viral particles. The delivery of an engineered virus to the murine prostate allows alteration of gene expression in the prostate epithelia. Virus type and quantity will hereby define the number of targeted cells for gene alteration by transducing a few cells for cancer initiation and many cells for gene therapy. Through surgery-based injection in the anterior lobe, distal from the urinary track, the tumor in this model can expand without impairing the urinary function of the animal. Furthermore, by targeting only a subset of prostate epithelial cells the technique enables clonal expansion of the tumor, and therefore mimics human tumor initiation, progression, as well as invasion through the basal membrane.
This novel technique provides a powerful prostate cancer model with improved physiological relevance. Animal suffering is limited, and since no additional breeding is required, overall animal count is reduced. At the same time, analysis of new candidate genes and pathways is accelerated, which in turn is more cost efficient.

This protocol describes a newly established method for virus delivery to the murine prostate. Using either CRISPR/Cas9 technology, gene overexpression, or Cre recombinase delivery, the technique allows orthotopic alteration of gene expression and implements a novel mouse model for prostate cancer.

Virüs gen değiştirme için fare prostat CRISPR kılavuzları teslimini

With an increasing incidence of prostate cancer, identification of new tumor drivers or modulators is crucial. Genetically engineered mouse models (GEMM) ...

Enrichment and Characterization of the Tumor Immune and Non-immune Microenvironments in Established Subcutaneous Murine Tumors

The tumor immune microenvironment (TIME) has recently been recognized as a critical mediator of treatment response in solid tumors, especially for immunotherapies. Recent clinical advances in immunotherapy highlight the need for reproducible methods to accurately and thoroughly characterize the tumor and its associated immune infiltrate. Tumor enzymatic digestion and flow cytometric analysis allow broad characterization of numerous immune cell subsets and phenotypes; however, depth of analysis is often limited by fluorophore restrictions on panel design and the need to acquire large tumor samples to observe rare immune populations of interest. Thus, we have developed an effective and high throughput method for separating and enriching the tumor immune infiltrate from the non-immune tumor components. The described tumor digestion and centrifugal density-based separation technique allows separate characterization of tumor and tumor immune infiltrate fractions and preserves cellular viability, and thus, provides a broad characterization of the tumor immunologic state. This method was used to characterize the extensive spatial immune heterogeneity in solid tumors, which further demonstrates the need for consistent whole tumor immunologic profiling techniques. Overall, this method provides an effective and adaptable technique for the immunologic characterization of subcutaneous solid murine tumors; as such, this tool can be used to better characterize the tumoral immunologic features and in the preclinical evaluation of novel immunotherapeutic strategies.

Here we describe a method to separate and enrich components of the tumor immune and non-immune microenvironment in established subcutaneous tumors. This technique allows for the separate analysis of tumor immune infiltrate and non-immune tumor fractions which can permit comprehensive characterization of the tumor immune microenvironment.

Zenginleştirme ve kurulan subkutan fare tümör tümör bağışıklık ve Non-immün Microenvironments karakterizasyonu

The tumor immune microenvironment (TIME) has recently been recognized as a critical mediator of treatment response in solid tumors, especially for ...

Identification, Histological Characterization, and Dissection of Mouse Prostate Lobes for In Vitro 3D Spheroid Culture Models

Genetically engineered mouse models (GEMMs) serve as effective pre-clinical models for investigating most types of human cancers, including prostate cancer (PCa). Understanding the anatomy and histology of the mouse prostate is important for the efficient use and proper characterization of such animal models. The mouse prostate has four distinct pairs of lobes, each with their own characteristics. This article demonstrates the proper method of dissection and identification of mouse prostate lobes for disease analysis. Post-dissection, the prostate cells can be further cultured in vitro for mechanistic understanding. Since mouse prostate primary cells tend to lose their normal characteristics when cultured in vitro, we outline here a method for isolating the cells and growing them as 3D spheroid cultures, which is effective for preserving the physiological characteristics of the cells. These 3D cultures can be used for analyzing cell morphology and behavior in near-physiological conditions, investigating altered levels and localizations of key proteins and pathways involved in the development and progression of a disease, and looking at responses to drug treatments.

Genetically engineered mice are useful models for investigating prostate cancer mechanisms. Here we present a protocol to identify and dissect prostate lobes from a mouse urogenital system, differentiate them based on histology, and isolate and culture the primary prostate cells in vitro as spheroids for downstream analyses.

Kimlik, histolojik karakterizasyonu ve için fare Vitro 3D küresel kültür modelleri prostat lob diseksiyonu

Genetically engineered mouse models (GEMMs) serve as effective pre-clinical models for investigating most types of human cancers, including prostate ...

Three-Dimensional Bone Extracellular Matrix Model for Osteosarcoma

Osteosarcoma (OS) is the most common and a highly aggressive primary bone tumor. It is characterized with anatomic and histologic variations along with diagnostic or prognostic difficulties. OS comprises genotypically and phenotypically heterogeneous cancer cells. Bone microenvironment elements are proved to account for tumor heterogeneity and disease progression. Bone extracellular matrix (BEM) retains the microstructural matrices and biochemical components of native extracellular matrix. This tissue-specific niche provides a favorable and long-term scaffold for OS cell seeding and proliferation. This article provides a protocol for the preparation of BEM model and its further experimental application. OS cells can grow and differentiate into multiple phenotypes consistent with the histopathological complexity of OS clinical specimens. The model also allows visualization of diverse morphologies and their association with genetic alterations and underlying regulatory mechanisms. As homologous to human OS, this BEM-OS model can be developed and applied to the pathology and clinical research of OS.

The bone extracellular matrix (BEM) model for osteosarcoma (OS) is well established and shown here. It can be used as a suitable scaffold for mimicking primary tumor growth in vitro and providing an ideal model for studying the histologic and cytogenic heterogeneity of OS.

Üç boyutlu kemik hücre dışı matriks modeli osteosarkom için

Osteosarcoma (OS) is the most common and a highly aggressive primary bone tumor. It is characterized with anatomic and histologic variations along with ...

Murine Model of Metastatic Liver Tumors in the Setting of Ischemia Reperfusion Injury

Liver ischemia and reperfusion (I/R) injury, a common clinical challenge, remains an inevitable pathophysiological process that has been shown to induce multiple tissue and organ damage. Despite recent advances and therapeutic approaches, the overall morbidity has remained unsatisfactory especially in patients with underlying parenchymal abnormalities. In the context of aggressive cancer growth and metastasis, surgical I/R is suspected to be the promoter regulating tumor recurrence. This article aims to describe a clinically relevant murine model of liver I/R and colorectal liver metastasis. In doing so, we aim to assist other investigators in establishing and perfecting this model for their routine research practice to better understand the effects of liver I/R on promoting liver metastases.

We describe in detail a clinically relevant colorectal cancer liver metastases (CRLM) tumor model and the influence of liver ischemia reperfusion (I/R) in tumor growth and metastasis. This model can help to better understand the mechanisms underlying surgery-induced promotion of liver metastatic growth.

İskemi Reperfüzyon Hasarı Ayarında Metastatik Karaciğer Tümörlerinin Murine Modeli

Liver ischemia and reperfusion (I/R) injury, a common clinical challenge, remains an inevitable pathophysiological process that has been shown to induce ...