We would like to thank Pascal DuBois for proof-reading the article. The authors would also like to thank the personnel of the Animal Facility at Hunter College of The City University of New York. This work was supported by a NIH RCMI grant to Hunter College.

윤리 선언문 모든 동물 실험은 뉴욕 시립 대학의 헌터 대학의 기관 동물 관리 및 사용위원회 (IACUC)에 의해 승인되었다. 주 : 에이트 BALB / c 마우스는이 실험 절차에서 사용 하였다. 다섯 BALB / c 마우스는 차 종양을 생성하기 위해 5 X 106 BNL 1ME A.7R.1 마우스 간암 세포를 이식 하였다. 아니오 주입 세 BALB / c 마우스는 대조군으로 사용 하였다. BNL 1ME A.7R.1은 쥐의 비 종양 간세포 세포주 BNL c12에에서 화학 변환에 의해 유도 된 쥐의 간세포 암 세포주이다. BNL 1ME A.7R.1는 아메리칸 타입 컬쳐 콜렉션 (ATCC)로부터 수득 하였다. 1. 사전 수술 <ol><li> 모든 생존 수술은 건강을 위해 그 쥐를 확인합니다. 마우스가 건강하도록하려면, 피부의 텐팅 (tenting)에 의해 탈수의 징후를 검사합니다. 질병 또는 d의 증거와 마우스를 사용하지 마십시오이 실험에서 istress. </li><li> 그들은 저체중하지 보장하기 위해 마우스의 무게를. 무게가 적어도 16-20g 있는지 확인하십시오. </li><li> 마우스 당 주입 5 × 10 6 BNL 1ME A.7R.1 마우스의 간세포 암 세포를 준비합니다 <ol><li> BNL 1ME A.7R.1 마우스 간암 세포의 70 % 융합 성 세포 배양 접시 - 10 ml의 60 %에서 대기음 매체. 5 ml의 PBS로 두 번 씻는다. 접시에 트립신 2 ML을 추가하고 5 분 동안 공기와 5 % CO 2 가습 배양기에서 37 ° C에서 품어. </li><li> 그 후, 피펫 10 μL은 10 배의 배율 현미경으로 세포 계수에 대한 수동 혈구로 세포를 트립신. 4 개의 큰 코너에 위치한 사각형 셀들의 총 수를 카운트하고 세포의 총 수에 의해 희석 계수를 곱한다. 이어서, (ML 당 셀) 카운트 제곱의 수로 나누어 세포 농도 10 (6)에 의해 곱. </li><li> 계산을 이용하여, 무균 마우스 당 500 만 세포를 수집1.5 ml의 마이크로 원심 튜브. 1.5 ml의 마이크로 원심 튜브 레이블. </li></ol></li><li> 포스페이트 완충 염수 100 ㎕ (PBS)에 5 × 106 BNL 1ME A.7R.1 마우스 HCC 세포 재현 탁. 수술에서 이식까지 얼음이 세포를 보관하십시오. </li><li> 마취의 유도 및 머리 오염을 방지하기 위해 멸균 장비와 물질을 멀리 클리핑 등의 수술에 대한 쥐의 준비에 필요한 활동을 수행합니다. </li><li> 분당 1 L (약. 2.1 %)의 농도를 일정한 유량으로 산소와 함께 기화기 이소 플루 란을 사용하여 이소 플루 란 흡입에 의해 마우스의 전신 마취를 유도한다. 주 : 시스템은 자립이며, 정밀 기화기, 산소 공급과 가스 / 마취제 소기 필요한 호흡 회로에 연결된 캐니스터 목탄을 포함한다. </li><li> 마취는 수술 중 통증을 겪고에서 동물을 방지하기 위해 적절한 지 확인하십시오. 발가락 핀치를 수행하고 비 respon 위해 동물을 관찰siveness 마취를 확인합니다. </li><li> 이 때, 코 콘에 밸브 라인, 인감 상자 열기를 전환합니다. </li><li> 즉시 마우스를 마취로, 건조 및 손상 예방 조치로 각막에 안과 연고의 작은 금액을 적용합니다. </li><li> 이어서, 노즈콘 제대로 (유지 마취)에 배치하고 연결하여, 준비 공간에 동물을 이동. 이 준비 영역에서 탈모에 의해 수술을위한 수술 절개 부위를 준비합니다. </li><li> 수술의 영역이 다른 모든 활동이없는 위치에 있는지 확인합니다. </li><li> 작업 표면이 견고하고 적절하게 소독, unsoiled과 혼란이없는 수술 전인지 확인하십시오. 리넨 또는 이상 수술 트레이의 살균 조각을 놓습니다. </li><li> 수술 필드는 수술을 수행하는 사람 만 사용할 수 있는지 확인합니다. 수술 방 적절한 소독과 청소하고 적절한 세제를 휩쓸 및 분사되어 있는지 확인ctant. </li><li> 그것은 또한 거리에 문 및 공기 흐름 등의 창에서 위치 사이트의 오염에 이르는 먼지를 확산 할 수 있는지 확인합니다. </li><li> 멸균 악기 생존 수술을 수행합니다. 어느 제조업체의 권장하거나 오토 클레이브에 따라 살균제에 악기를 흡수. 사용할 준비가 기본 멸균 공급 장치를 사용합니다. </li><li> 별도의 장비는 기능에 따라 멸균 지역의 오염을 방지하기 위해. </li><li> 동물의 간을 액세스 할 수 절개 (중간 선 절개)의 사이트 주위에 약 1 인치를 면도. 수술의 사이트에서 멀리 동물에서 머리를 제거합니다. </li><li> 면도 후, 비누 살균제 (betadine 수술 스크럽)와 영역을 문질러 멸균 PBS 용액에 과량의 소독제를 씻어. 약 5 분 동안 지속 전체 스크럽 과정이 여러 번 수행합니다. 제거하고 사용 장갑을 폐기. </li><li> 그 후, 같은 surgic로 보호 복으로 변경알 가운, 스크럽 상단, 얼굴 마스크입니다. 다음으로, 수술 장갑 착용하기 전에 적절한 스크럽 솔루션 손을 소독. 참고 :이 흘리기 및 잠재적 오염 물질과 수술 부위의 오염 방지하기 위해 필요한 단계입니다. </li></ol> 2. 수술 <ol><li> 마취의 전체 기간 동안 따뜻함을위한 외부 열원을 제공합니다. 마취는 체온 조절의 장애를 야기하기 때문에 동물은 일부 저체온증이 발생할 수 있습니다; 따라서, 체온을 유지한다. </li><li> 수술을 통해 그 뒷면에 마우스를 놓습니다. </li><li> 외과 테이프 조각과 수술 테이블에 앞 사지, 뒷다리와 마우스의 꼬리를 수정합니다. </li><li> xyphisternum에 직선 절개 즉시 열등합니다. </li><li> 간을 노출 멸균 자기 유지 외과 견인기를 사용합니다. </li><li> 주사기 차분히, 20G 바늘을 갖는 5 × 10 6 BNL 1ME A.7R.1를 포함하는 PBS 100 ㎕를 주입간 우엽에 마우스 간암 세포. </li><li> 바늘을 제거한 후, 인해 간 높은 혈관에 잠재적 인 출혈을 멈추게하기 위해 적어도 1 분 동안 멸균 면봉을 적용합니다. </li><li> 수술 클립을 사용하여 피부 폐쇄 다음 흡수성 봉합사와 피하 조직을 닫습니다. 그 후, betadine 스크럽과 함께 다시 한 번 상처를 청소하십시오. 참고 :이 수술 상처의 폐쇄를 보장합니다. </li><li> 피하 생리 식염수 100 ㎕를 제공합니다. </li></ol> 3. 수술 후 <ol><li> 수술 후 즉시, 마우스 조건 관리인에게 통지 수술 후 신분증 깨끗한 케이지에 마우스를 옮긴다. 음식과 물을 제공합니다. 참고 : 우리의 경험에 의하면, 초기 24 시간의 기간 후 수술 젖은 음식과 물을 혼자 보관하면 그냥 최선을 운임됩니다 수술을 한 마우스를. </li><li> 38 ºC을 능가하지 열을 생성하는 가열 램프와 함께 동물을 따뜻하게 유지. 유마우스가 완전히 회복 될 때까지 온도를 측정하는 온도계를 SE는. </li><li> 바람직하게는 종이 침구와 케이지 하우스 동물의 편안함을 보장합니다. </li><li> 초기 복구 한 후, 불편 함, 출혈, 통증과 고통의 징후 동물을 모니터링 할 수 있습니다. 통증을 완화하기 위해 프레 노르 핀 (0.05 -0.1 ㎎ / ㎏)을 얻었다. 밀접하게 약 2 시간 동안 활성 동작과 모양뿐만 아니라 물과 음식 소비를 모니터링. 외래 경우, 마우스의 정규 일반 주택을 다시 지역에 동물을 이동하고 24 시간 후 수술 6-12 시간 간격으로 모니터링을 계속합니다. 원하는 경우 체중 검사를 수행합니다. </li><li> 동물이 문제가 회복 데 경우, 적절한지지 요법의 조치를 취할. 마우스는 복구 할 수없는 경우, 인도적으로 CO 2 질식 자궁 경부 전위를 통해 쥐를 안락사. 안락사에 대한 승인 기관의 지침을 따르십시오. </li><li> 수술 후 보통 observab입니다 간세포 암의 임상 적 증거를 확인몸 상태 점수 르로 크게 감소. 주 :이 시점에서, 일차 종양 가능성 간에서 개발 및 전이성 예금 가능성 폐에서 개발 하였다. </li><li> 이 시점에서, 인도적인 안락사에 의해 실험에 참여 (실험 및 제어) 모든 쥐를 희생. 이산화탄소로 질식 생쥐 대상 : 매우 천천히 -10 5 분에 걸쳐 챔버 내에 이산화탄소를 방출. 이것은 호흡 정지의 원인이됩니다. </li></ol>

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The authors declare that they have no competing financial interests.

이 문서의 방법에 대한 깊이있는 설명은 (그림 2)이 최근 Ogunwobi 및 간세포 암 전이의 동소 동계 마우스 모델의 성공적인 설립의 동료에 의해보고되었다 주어진다 3. 이 절차에 대한 종양의 테이크 속도는 일반적으로 높다. 우리는 이전에 100 % 3의 종양 걸릴 속도를 관찰했다. 그러나 테이크 속도는 연구자의 능력에 따라 변수가 될 수 있습니다. 훈련하에 새로운 대학원?...

Hepatocellular carcinoma (HCC) is one of the most lethal cancers, with poor prognosis and low life expectancy. Nearly all cancer-related deaths are due to the metastatic spread of the disease from the originating organ to additional distant organs2-6.HCC progression is a complex process. Therefore, being able to model the tumor microenvironment that is naturally found in metastatic HCC in animal models can prove to be a successful and useful way to reveal relevant mechanisms in humans.7Unfortunately, the mechanisms of cancer metastasis are still largely unclear. Therefore, there is a need to establish animal models that will enable us to elucidate the underlying molecular mechanisms of metastases of cancers such as HCC3,5.
Mouse model systems are a very useful approach for delineating mechanisms and evaluating novel strategies for treatment of metastatic human cancers5,8. The various mouse model systems that presently exist are a testament to efforts of researchers to correctly depict the complexity of the disease5,8,9.
Ogunwobi and colleagues recently used a survival surgical approach to demonstrate establishment of a novel orthotopic syngeneic mouse model for the study of metastasis in HCC3. Their work established a &#8220;patient-like&#8221; mouse model that recapitulates features of aggressive and metastatic HCCs3. They further demonstrated that this mouse model system can be used to study the biology of circulating tumor cells, and that this holds potential for gaining novel insights into the mechanisms of cancer metastasis3.
The aim of this paper is to describe in detail the methodology used in establishing this &#8220;patient-like&#8221; orthotopic syngeneic mouse model of HCC metastasis3. The methodology of how to implant BNL 1ME A.7R.1 mouse HCC cells directly into the liver (the organ of origin) of the inbred wild-type immune competent laboratory mouse strain, BALB/c3 using survival surgery will be described. Unlike other mouse xenograft tumor models where human tumor cells are implanted into immune deficient mice, this system is syngeneic and is, therefore, suitable for studying the role of the immune system in tumor metastasis3,5,8. This approach will likely gain widespread use for studying the mechanisms of metastasis in solid organ cancers.

<table border="0">
	<tbody>
		<tr>
			<td>Micro Dissecting Tweezer&#160;</td>
			<td>Roboz</td>
			<td>(RS-5040)</td>
			<td>Tip .20 x .12mm</td>
		</tr>
		<tr>
			<td>Graefe Micro Dissecting Forceps, serrated curved tip</td>
			<td>Roboz</td>
			<td>(RS-5111)</td>
			<td>1 X 2 teeth, curved tip width 0.6mm</td>
		</tr>
		<tr>
			<td>Micro Dissecting Retractor-Agricola (3 by 3 prongs)</td>
			<td>Roboz</td>
			<td>(RS-6501)</td>
			<td>Blunt 3 X 3 prongs, depth 4mm, spread 25mm</td>
		</tr>
		<tr>
			<td>Micro Dissecting Retractor-Goldstein (3 by 3 prongs)</td>
			<td>Roboz</td>
			<td>(RS-6503)</td>
			<td>Blunt, 3 X 3 prongs, depth 4mm, spread 19mm</td>
		</tr>
		<tr>
			<td>Jameson Caliper (Measures tumors)</td>
			<td>Roboz</td>
			<td>(RS-6466)</td>
			<td>80mm/3 inch scale, chrome plated&#160;</td>
		</tr>
		<tr>
			<td>Micro Dissecting Scissors, large ring sissors, straight and sharp</td>
			<td>Roboz</td>
			<td>(RS-5852)</td>
			<td>23mm blades, length 4 inches, flat shanks and large rings</td>
		</tr>
		<tr>
			<td>Scalpel with blades, for delicate dissecting procedures</td>
			<td>Roboz</td>
			<td>(RS-9861-36)</td>
			<td></td>
		</tr>
		<tr>
			<td>Scalpel handle&#160;</td>
			<td>Roboz</td>
			<td>(RS-9884)</td>
			<td>Solid</td>
		</tr>
		<tr>
			<td>Littauer Stitch Sissors</td>
			<td>Roboz</td>
			<td>(RS-7074)</td>
			<td>Length 4.5 inches</td>
		</tr>
		<tr>
			<td>Brown needle holder, for easy suture tying</td>
			<td>Roboz</td>
			<td>(RS-7960)</td>
			<td>Convex jaw, fine serrations</td>
		</tr>
		<tr>
			<td>Reflex 7MM wound clips with reflex 7 clip applier</td>
			<td>Roboz</td>
			<td>(RS-9262)</td>
			<td>safe, secure alternative method of wound closure</td>
		</tr>
		<tr>
			<td>Instrument tray and lid</td>
			<td>Roboz</td>
			<td>RT-1350S</td>
			<td></td>
		</tr>
		<tr>
			<td>Mini-clipper with detachable blade</td>
			<td>Roboz</td>
			<td>(RS-5903)</td>
			<td></td>
		</tr>
		<tr>
			<td>Germinator 500 (the Germ Terminator) Dry Sterilizer</td>
			<td>Roboz</td>
			<td>Ds-400, Ds-401, DS-501</td>
			<td>For fast decontamination of micro-dissecting instruments.&#160;&#160; Instruments decontaminate within 15 seconds.&#160;</td>
		</tr>
		<tr>
			<td>Microinjection needles</td>
			<td>VWR</td>
			<td>BD305125</td>
			<td>25G Needle</td>
		</tr>
	</tbody>
</table>

a "patient-like" orthotopic syngeneic mouse model of hepatocellular carcinoma metastasis

The majority of cancer-related deaths are caused by the metastasis of the cancer rather than the primary tumor itself. Yet, the underlying mechanisms of cancer metastasis are still unclear. Animal models are essential for elucidating the mechanisms and for evaluating novel strategies for the treatment of metastatic cancers. Here, an in-depth description of a &#8220;patient-like&#8221; orthotopic syngeneic mouse model for exploring the mechanisms of metastasis of solid organ tumors is provided. The survival surgical implantation of BNL 1ME A.7R.1 mouse hepatocellular carcinoma cells directly into the liver (the organ of origin) of the inbred wild-type immune competent laboratory mouse strain, BALB/c is described. The success and reproducibility of this methodology recommends it for widespread use in elucidating the biological mechanisms of solid organ cancer metastasis.

을 Balb / c 마우스의 간은 5 × 106 BNL 1ME A.7R.1 마우스 간암 세포를 이식 하였다. HCC 개발의 임상 적 증거가 육십삼일 수술 후 관찰했다. 따라서, 마우스는 인도적으로 안락사시켰다. 검시는 안락사 쥐에 대해 수행 하였다. 폐 및 간을 절제하고, 조심 육안 검사를 육안 종양을 확인하기 위해 수행 하였다. 하나의 마우스에서 종양이 표면 복벽 (도 1a)에 일부 첨부 간 표면에서 발견되...

Watch this Scientific Journal Video about A "Patient-Like" Orthotopic Syngeneic Mouse Model of Hepatocellular Carcinoma Metastasis at JoVE.com

A &quot;Patient-Like&quot; Orthotopic Syngeneic Mouse Model of Hepatocellular Carcinoma Metastasis

암의 전이 확산 암 관련 사망의 주요 원인이다. 우리는 고형 장기의 종양에서 전이의 메커니즘을 연구하기위한 &quot;환자와 같은&quot;동 소성 동계 마우스 모델 시스템 구축을위한 우리의 생존 수술 방법에 대한 심층적 인 설명을 제공한다.

&quot;환자와 같은&quot;동소 이식 동계 마우스 모델 간세포 암의 전이

The majority of cancer-related deaths are caused by the metastasis of the cancer rather than the primary tumor itself. Yet, the underlying mechanisms of ...

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A "Patient-Like" Orthotopic Syngeneic Mouse Model of Hepatocellular Carcinoma Metastasis

12.1K 조회수.  Hunter College of the City University of New York.  암의 전이 확산 암 관련 사망의 주요 원인이다. 우리는 고형 장기의 종양에서 전이의 메커니즘을 연구하기위한 "환자와 같은"동 소성 동계 마우스 모델 시스템 구축을위한 우리의 생존 수술 방법에 대한 심층적 인 설명을 제공한다. 

비디오: A "Patient-Like" Orthotopic Syngeneic Mouse Model of Hepatocellular Carcinoma Metastasis

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.

구강 편평 세포 암종의 Orthotopic 마우스 모델에서 종양 세포 침략의 다중 광자 이미징

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

연구

의학

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.

유방암 뇌 전이의 종양 Hypoxia 역학 생체내의 Bioluminescence 이미징에서는 마우스 모델에서

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

Combination Radiotherapy in an Orthotopic Mouse Brain Tumor Model

Glioblastoma multiforme (GBM) are the most common and aggressive adult primary brain tumors1. In recent years there has been substantial progress in the understanding of the mechanics of tumor invasion, and direct intracerebral inoculation of tumor provides the opportunity of observing the invasive process in a physiologically appropriate environment2. As far as human brain tumors are concerned, the orthotopic models currently available are established either by stereotaxic injection of cell suspensions or implantation of a solid piece of tumor through a complicated craniotomy procedure3. In our technique we harvest cells from tissue culture to create a cell suspension used to implant directly into the brain. The duration of the surgery is approximately 30 minutes, and as the mouse needs to be in a constant surgical plane, an injectable anesthetic is used. The mouse is placed in a stereotaxic jig made by Stoetling (figure 1). After the surgical area is cleaned and prepared, an incision is made; and the bregma is located to determine the location of the craniotomy. The location of the craniotomy is 2 mm to the right and 1 mm rostral to the bregma. The depth is 3 mm from the surface of the skull, and cells are injected at a rate of 2 &mu;l every 2 minutes. The skin is sutured with 5-0 PDS, and the mouse is allowed to wake up on a heating pad. From our experience, depending on the cell line, treatment can take place from 7-10 days after surgery. Drug delivery is dependent on the drug composition. For radiation treatment the mice are anesthetized, and put into a custom made jig. Lead covers the mouse's body and exposes only the brain of the mouse. The study of tumorigenesis and the evaluation of new therapies for GBM require accurate and reproducible brain tumor animal models. Thus we use this orthotopic brain model to study the interaction of the microenvironment of the brain and the tumor, to test the effectiveness of different therapeutic agents with and without radiation.

The purpose of this article is to describe the use of an orthotopic glioblastoma model for chemoradiation studies. This article will go though cell processing, implanting, and radiotherapy of the mouse using an intracranial model.

Orthotopic 마우스 뇌종양 모델의 조합 방사선 치료

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

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.

역 형성 갑상선 암의 동소 마우스 모델

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

Generation of Subcutaneous and Intrahepatic Human Hepatocellular Carcinoma Xenografts in Immunodeficient Mice

In vivo experimental models of hepatocellular carcinoma (HCC) that recapitulate the human disease provide a valuable platform for research into disease pathophysiology and for the preclinical evaluation of novel therapies. We present a variety of methods to generate subcutaneous or orthotopic human HCC xenografts in immunodeficient mice that could be utilized in a variety of research applications. With a focus on the use of primary tumor tissue from patients undergoing surgical resection as a starting point, we describe the preparation of cell suspensions or tumor fragments for xenografting. We describe specific techniques to xenograft these tissues i) subcutaneously; or ii) intrahepatically, either by direct implantation of tumor cells or fragments into the liver, or indirectly by injection of cells into the mouse spleen. We also describe the use of partial resection of the native mouse liver at the time of xenografting as a strategy to induce a state of active liver regeneration in the recipient mouse that may facilitate the intrahepatic engraftment of primary human tumor cells. The expected results of these techniques are illustrated. The protocols described have been validated using primary human HCC samples and xenografts, which typically perform less robustly than the well-established human HCC cell lines that are widely used and frequently cited in the literature. In comparison with cell lines, we discuss factors which may contribute to the relatively low chance of primary HCC engraftment in xenotransplantation models and comment on technical issues that may influence the kinetics of xenograft growth. We also suggest methods that should be applied to ensure that xenografts obtained accurately resemble parent HCC tissues.

Human tumor xenografts in immunodeficient mice are valuable tools to study cancer biology. Specific protocols to generate subcutaneous and intrahepatic xenografts from human hepatocellular carcinoma cells or tumor fragments are described. Liver regeneration induced by partial hepatectomy in recipient mice is presented as a strategy to facilitate intrahepatic engraftment.

면역 결핍 마우스의 피하 및 간내 인간의 간세포 암 이종 이식의 생성

In  vivo experimental models of hepatocellular carcinoma (HCC) that recapitulate  the human disease provide a valuable platform for research into disease  ...

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.

인간의 전립선 암 전이의 동소 쥐과 모델

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  ...

Initiation of Metastatic Breast Carcinoma by Targeting of the Ductal Epithelium with Adenovirus-Cre: A Novel Transgenic Mouse Model of Breast Cancer

Breast cancer is a heterogeneous disease involving complex cellular interactions between the developing tumor and immune system, eventually resulting in exponential tumor growth and metastasis to distal tissues and the collapse of anti-tumor immunity. Many useful animal models exist to study breast cancer, but none completely recapitulate the disease progression that occurs in humans. In order to gain a better understanding of the cellular interactions that result in the formation of latent metastasis and decreased survival, we have generated an inducible transgenic mouse model of YFP-expressing ductal carcinoma that develops after sexual maturity in immune-competent mice and is driven by consistent, endocrine-independent oncogene expression. Activation of YFP, ablation of p53, and expression of an oncogenic form of K-ras was achieved by the delivery of an adenovirus expressing Cre-recombinase into the mammary duct of sexually mature, virgin female mice. Tumors begin to appear 6 weeks after the initiation of oncogenic events. After tumors become apparent, they progress slowly for approximately two weeks before they begin to grow exponentially. After 7-8 weeks post-adenovirus injection, vasculature is observed connecting the tumor mass to distal lymph nodes, with eventual lymphovascular invasion of YFP+ tumor cells to the distal axillary lymph nodes. Infiltrating leukocyte populations are similar to those found in human breast carcinomas, including the presence of &#945;&#946; and &#947;&#948; T cells, macrophages and MDSCs. This unique model will facilitate the study of cellular and immunological mechanisms involved in latent metastasis and dormancy in addition to being useful for designing novel immunotherapeutic interventions to treat invasive breast cancer.

Activation of latent mutations with adenovirus-Cre into the mammary ductal system results in a clinically relevant metastatic breast cancer. Incorporation of a YFP promoter allows tracking of distal metastatic tumor cells. This model is useful to study latent metastasis, anti-tumor immunity, and for designing novel immunotherapies to treat breast cancer.

유방암의 소설 형질 전환 마우스 모델 : 아데노 바이러스 크레와 유관 상피의 대상으로 전이성 유방암의 개시

Breast cancer is a heterogeneous disease involving complex cellular interactions between the developing tumor and immune system, eventually resulting in ...

The In Ovo Chick Chorioallantoic Membrane (CAM) Assay as an Efficient Xenograft Model of Hepatocellular Carcinoma

The chick chorioallantoic membrane (CAM) begins to develop by day 7 after fertilization and matures by day 12. The CAM is naturally immunodeficient and highly vascularized, making it an ideal system for tumor implantation. Furthermore, the CAM contains extracellular matrix proteins such as fibronectin, laminin, collagen, integrin alpha(v)beta3, and MMP-2, making it an attractive model to study tumor invasion and metastasis. Scientists have long taken advantage of the physiology of the CAM by using it as a model of angiogenesis. More recently, the CAM assay has been modified to work as an in vivo xenograft model system for various cancers that bridges the gap between basic in vitro work and more complex animal cancer models. The CAM assay allows for the study of tumor growth, anti-tumor therapies, and pro-tumor molecular pathways in a biologically relevant system that is both cost- and time-effective. Here, we describe the development of CAM xenograft model of hepatocellular carcinoma (HCC) with embryonic survival rates of up to 93% and reliable tumor take leading to growth of three-dimensional, vascularized tumors.

The In Ovo Chick Chorioallantoic Membrane CAM Assay as an Efficient Xenograft Model of Hepatocellular Carcinoma

The chick chorioallantoic membrane (CAM) is immunodeficient and highly vascularized, making it a natural in vivo model of tumor growth and angiogenesis. In this protocol, we describe a reliable method of growing three-dimensional, vascularized hepatocellular carcinoma (HCC) tumors using the CAM assay.

<em&gt; 비켜에서</em간세포 암의 효율적인 이종 이식 모델로&gt; 병아리 융모 막 (CAM) 분석

The chick chorioallantoic membrane (CAM) begins to develop by day 7 after fertilization and matures by day 12. The CAM is naturally immunodeficient and ...

Dynamic Contrast Enhanced Magnetic Resonance Imaging of an Orthotopic Pancreatic Cancer Mouse Model

Dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) has been limitedly used for orthotopic pancreatic tumor xenografts due to severe respiratory motion artifact in the abdominal area. Orthotopic tumor models offer advantages over subcutaneous ones, because those can reflect the primary tumor microenvironment affecting blood supply, neovascularization, and tumor cell invasion. We have recently established a protocol of DCE-MRI of orthotopic pancreatic tumor xenografts in mouse models by securing tumors with an orthogonally bent plastic board to prevent motion transfer from the chest region during imaging. The pressure by this board was localized on the abdominal area, and has not resulted in respiratory difficulty of the animals. This article demonstrates the detailed procedure of orthotopic pancreatic tumor modeling using small animals and DCE-MRI of the tumor xenografts. Quantification method of pharmacokinetic parameters in DCE-MRI is also introduced. The procedure described in this article will assist investigators to apply DCE-MRI for orthotopic gastrointestinal cancer mouse models.

The goal of this protocol is to apply dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) for orthotopic pancreatic tumor xenografts in mice. DCE-MRI is a non-invasive method to analyze microvasculature in a target tissue, and useful to assess vascular response in a tumor following a novel therapy.

동소 이식 췌장 암 마우스 모델의 동적 대비 강화 된 자기 공명 영상

Dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) has been limitedly used for orthotopic pancreatic tumor xenografts due to severe ...

Orthotopic Implantation and Peripheral Immune Cell Monitoring in the II-45 Syngeneic Rat Mesothelioma Model

The enormous upsurge of interest in immune-based treatments for cancer such as vaccines and immune checkpoint inhibitors, and increased understanding of the role of the tumor microenvironment in treatment response, collectively point to the need for immune-competent orthotopic models for pre-clinical testing of these new therapies. This paper demonstrates how to establish an orthotopic immune-competent rat model of pleural malignant mesothelioma. Monitoring disease progression in orthotopic models is confounded by the internal location of the tumors. To longitudinally monitor disease progression and its effect on circulating immune cells in this and other rat models of cancer, a single tube flow cytometry assay requiring only 25 &#181;l whole blood is described. This provides accurate quantification of seven immune parameters: total lymphocytes, monocytes and neutrophils, as well as the T-cell subsets CD4 and CD8, B-cells and Natural Killer cells. Different subsets of these parameters are useful in different circumstances and models, with the neutrophil to lymphocyte ratio having the greatest utility for monitoring disease progression in the mesothelioma model. Analyzing circulating immune cell levels using this single tube method may also assist in monitoring the response to immune-based treatments and understanding the underlying mechanisms leading to success or failure of treatment.

The generation of an orthotopic rat model of pleural malignant mesothelioma by implantation of II-45 mesothelioma cells into the pleural cavity of immune competent rats is presented. A flow cytometric method to analyze seven immune cell subsets in these animals from a 25 &#181;l&#160;blood sample is also described.

II-45 동계 쥐의 Mesothelioma 모델에서 소성을 주입 및 주변 면역 세포 모니터링

The enormous upsurge of interest in immune-based treatments for cancer such as vaccines and immune checkpoint inhibitors, and increased understanding of ...