作者感谢妇产科系的成员和癌症生物学为他们有益的讨论和技术援助。 （;到H梶山15K15604）这项研究是由日本学术振兴会（JSPS）赠款在资助科学研究的支持。

该治疗方案遵循由名古屋大学通过动物实验的指导方针。 1.悬浮细胞的制备<ol><li>人卵巢透明细胞癌的细胞系。 <ol><li>维持ES-2细胞12中，用10％胎牛血清（FBS）和青霉素/链霉素的RPMI-1640培养基（青霉素:100单位/ ml，链霉素:0.1毫克/毫升）。在一个5％CO 2的加湿培养箱中培养细胞在37℃。 </li><li>收集在对数生长期的细胞在0.05％胰蛋白酶/ 0.02％EDTA溶液（胰蛋白酶/ EDTA）。 <ol><li>首先，从细胞培养容器中删除旧的介质，然后用PBS洗涤细胞一次（PBS不含Ca 2+ / Mg 2+离子 ，3 -第4毫升，每为25cm 2）。 </li><li>接下来，添加每25厘米胰蛋白酶/ EDTA的2 1毫升旋转细胞培养容器以覆盖与胰蛋白酶/ EDTA的细胞单层。拆除和丢弃胰蛋白酶/ EDTA后，返回小区培养容器的孵化器3 - 5分钟或直至细胞被分离。 </li><li>在显微镜下检查细胞以确保将细胞从细胞培养容器中，浮动的表面脱离。 </li><li>添加含有生长培养基以灭活胰蛋白酶（5毫升，每为25cm 2），并重新悬浮细胞新鲜血清。传送再悬浮细胞至15ml锥形管中。 </li><li>离心在300×g离心5分钟。在4℃或室温下进行离心任。小心取出上清，重新暂停与生长培养基细胞（每为25cm 2 5ml）中。 </li></ol></li><li>计数细胞（ 例如 ，通过台盼蓝排除），然后在生长培养基中以2×10 8个细胞/ ml的密度重新悬浮。把悬浮细胞冰上。 </li></ol></li><li>解冻冷冻的细胞外基质（ECM）为基础的水凝胶的等分试样在冰浴中。 </li><li>添加细胞悬浮液，以基于ECM的水凝胶的等分试样在1:1的比例（终小区concentration:1×10 5细胞/基于中等的ECM的水凝胶的微升）并充分混合。放置制备的细胞悬浮液在冰浴中直至使用。 </li></ol> 2.原位接种悬浮细胞注意:手术不需要专用的套件。手术可以在实验室台式在是单独的，并具有最小的活动房的区域来执行。层流罩也可被使用。无菌手术器械必须使用剪刀不应该被用来使皮肤切口。 <ol><li>使用8周龄的雌性裸鼠（BALB / c小鼠NU / NU）此原位接种模型。 </li><li>首先，麻醉用吸入异氟烷鼠标（2 - 4％）。通过验证后，缺乏坚定的脚趾捏撤回检查麻醉深度。鼠标麻醉后，将一个平淡无菌眼药膏的眼睛，以防止干燥是必要的。 </li><li>放置鼠标麻醉上OPERATING阶段腹侧下来。消毒手术区数次酒精和一个基于洗必泰碘基或擦洗。 </li><li>使靠近垂直右肋弓与股骨之间的脊椎1〜2厘米的切口。使用切口周围部位的无菌手术盖布。用夹子固定修复皮肤切表面，以保持开放。 注意:拉钩可被用来代替夹具。 </li><li>接着，使一第二切口（大约1厘米）在壁层腹膜第一切口下，打开腹腔。夹紧阴刻腹部表面。夹与夹子围绕患侧卵巢和输卵管从腹腔取出卵巢的脂肪组织。小心地放在纱布卵巢和输卵管。然后，将鼠标腹侧下解剖显微镜下。 </li><li>将细胞悬液中胰岛素注射器（1/2毫升，29克）刚刚注射前。小心注入所制备的细胞悬浮液（1- 2微升）进卵巢。几秒，保留卵巢内的针，以确保基于ECM的水凝胶的凝胶化。凝胶化后，注入肿瘤细胞保持卵巢内。 </li><li>仔细返回卵巢到体内。缝合无菌医疗丝绸第二切口，然后用密封夹伤皮肤。 </li><li>手术后，将小鼠施用初始24小时的时间内的镇痛（ 例如 ，美洛昔康，肌内（IM）或皮下（sc），0.2毫克/千克）。 </li></ol> 3.手术后的治疗<ol><li>将在干净的纸单独的笼子每只小鼠用热敷，以保持温暖鼠标，直到它已经恢复了足够的意识，以保持胸骨斜卧。然后，返回的动物笼子里饲养和监控。每天监控小鼠3 - 7日之后进行手术治疗。 </li><li>术后小鼠监控<ol><li>在疼痛的迹象的情况下，管理analgeSICS每天（ 如美洛昔康，IM或SC，0.2毫克/千克）。 </li><li>如果小鼠有感染（ 如发红，肿胀，放电）的迹象，每天给药注射抗感染剂（ 如青霉素，即时通讯，10 IU /公斤）。 </li></ol></li><li>手术后的治疗后14天 - 取下皮肤缝合材料10。 </li></ol> 4.肿瘤和转移的分析注意:肿瘤会在注射子房2形成 - 3周接种后。 <ol><li>牺牲由二氧化碳的小鼠，然后收集样品（ 例如 ，肿瘤，转移瘤，器官）13-15作进一步的分析14,16-20。 注:如果在体内成像系统是可用的，轴承产生生物发光（ 如萤光素酶）或荧光蛋白（ 例如 ，GFP）的酶的细胞可以在这个协议中使用。这个修改后的协议将提供有用的信息癌细胞从原发肿瘤扩散的动力学。 </li></ol>

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作者什么都没有透露。

动物模型是分析肿瘤的发展，进展，转移，和药物疗效的机理对癌细胞是必不可少的。荷人卵巢癌细胞也已用作动物模型。这里，我们描述在原位的网站卵巢肿瘤细胞的给药的微创过程。通过从背侧面腹膜后方法使用这一步骤，接种到卵巢优于其它常用的技术提供显著优点。首先，原位接种可以提供反映在人类13,21肿瘤环境更有效和准确的结果。其次，从背侧此腹膜后方法只需要一个小的...

上皮性卵巢癌（EOC）占癌症相关死亡率的妇科恶性肿瘤1中的最高速度。 EOC与预后较差，主要是由于其后期症状，并经常与多个浸染腹腔及远处转移有关2-4。腹膜传播是一个多步骤的过程。首先，肿瘤细胞从原发病灶脱落，并迁移进入腹腔。当肿瘤细胞附着到腹膜间皮后，便开始通过间皮5,6-侵入组织。为了更好地理解肿瘤生物学（ 如癌症进展和治疗反应），小鼠模型提供了丰富的信息。与人癌细胞的异种移植物被广泛用于其中将人癌细胞皮下接种，腹膜内，静脉内，以及原位小鼠模型。与原位克动物模型筏肿瘤可以更有效地和精确地生成反映在人类肿瘤环境中与一个异位接枝肿瘤的动物模型中比较结果。因此，对许多人类肿瘤，原位移植模型已经建立7-11。 这里，我们描述通过从背侧面，这限制了侵袭性相比腹侧接种腹膜后途径的人类EOC细胞的原位接种。这种技术可提供各种对EOC有用的信息，特别是腹膜内传播的机制。

<table border="0" cellpadding="0" cellspacing="0" width="744">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:268px;">Matrigel matrix&#160;</td>
			<td style="width:151px;">BD</td>
			<td style="width:163px;">354234</td>
			<td style="width:163px;">ECM-based hydrogel</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Insulin syringe</td>
			<td>TERUMO</td>
			<td>SS-05M2913</td>
			<td>1/2cc, 29Gx1/2&#34;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Suturing needle with suture</td>
			<td></td>
			<td></td>
			<td>11mm, 3/8, Nylon6-0</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Reflex 7mm Wound Clip Applier</td>
			<td>CellPoint Scientific</td>
			<td>204-1000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">&#160;Reflex 7mm wound clips</td>
			<td>CellPoint Scientific</td>
			<td>203-1000</td>
			<td></td>
		</tr>
	</tbody>
</table>

murine experimental model of original tumor development and peritoneal metastasis via orthotopic inoculation with ovarian carcinoma cells

上皮性卵巢癌（EOC）与预后不良，因为它表明腹膜转移有关。改善预后，以控制腹膜是重要的。然而，目前还不清楚的肿瘤细胞是如何从原发病灶分离并附着到间皮。适当的动物模型的建立需要获得体内腹膜的机制的理解。在目前的研究中，我们介绍从局部注射EOC细胞进入鼠卵巢表面向转移的发展，包括腹膜及远处器官的过程。在8周龄雌性裸鼠（BALB / c小鼠NU / NU）被使用。下的视显微镜视野，EOC细胞（1×10 5细胞/中细胞外基质（ECM）的微升为基础的水凝胶/单侧卵巢/小鼠）通过从背侧面腹膜后途径注射到鼠卵巢。这种方法是一种乐为鼠标SS侵入性的程序，最大限度地减少对卵巢的伤害。在这里，我们描述了在原始和转移性肿瘤的形成的EOC的发展的方法步骤。

<html>六裸鼠接种了与人类透明细胞癌细胞系ES-2自己的卵巢，如本协议所述。如在图1A中所示，2周后，将6只小鼠的5例中具有ES-2细胞接种卵巢肿瘤，但有在对侧卵巢肿瘤无无接种（用作对照）。此外，5只小鼠的2显示腹水多腹膜浸染。细胞转移到肝脏（ 图1B）。卵巢，都与在接种部位和在控制侧上的肿瘤块，进行解剖，切片，用苏木精-曙红（HE）染...

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Murine Experimental Model of Original Tumor Development and Peritoneal Metastasis via Orthotopic Inoculation with Ovarian Carcinoma Cells

澄清卵巢癌的腹膜传播的多步骤的过程，目前的研究提出了原发肿瘤的发展和腹膜转移通过原位接种这些肿瘤细胞的鼠实验模型。

原发肿瘤发展小鼠实验模型和腹膜转移<em&gt;通过</em&gt;原位接种卵巢癌细胞

Epithelial ovarian carcinoma (EOC) is associated with a poor prognosis because it shows peritoneal dissemination. To improve the prognosis, it is ...

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Murine Experimental Model of Original Tumor Development and Peritoneal Metastasis via Orthotopic Inoculation with Ovarian Carcinoma Cells

10.4K 次观看.  Bell Research Center for Reproductive Health and Cancer. 澄清卵巢癌的腹膜传播的多步骤的过程，目前的研究提出了原发肿瘤的发展和腹膜转移通过原位接种这些肿瘤细胞的鼠实验模型。 

视频: Murine Experimental Model of Original Tumor Development and Peritoneal Metastasis via Orthotopic Inoculation with Ovarian Carcinoma Cells

In Vivo Model for Testing Effect of Hypoxia on Tumor Metastasis

Hypoxia has been implicated in the metastasis of Ewing sarcoma (ES) by clinical observations and in vitro data, yet direct evidence for its pro-metastatic effect is lacking and the exact mechanisms of its action are unclear. Here, we report an animal model that allows for direct testing of the effects of tumor hypoxia on ES dissemination and investigation into the underlying pathways involved. This approach combines two well-established experimental strategies, orthotopic xenografting of ES cells and femoral artery ligation (FAL), which induces hindlimb ischemia. Human ES cells were injected into the gastrocnemius muscles of SCID/beige mice and the primary tumors were allowed to grow to a size of 250 mm3. At this stage either the tumors were excised (control group) or the animals were subjected to FAL to create tumor hypoxia, followed by tumor excision 3 days later. The efficiency of FAL was confirmed by a significant increase in binding of hypoxyprobe-1 in the tumor tissue, severe tumor necrosis and complete inhibition of primary tumor growth. Importantly, despite these direct effects of ischemia, an enhanced dissemination of tumor cells from the hypoxic tumors was observed. This experimental strategy enables comparative analysis of the metastatic properties of primary tumors of the same size, yet significantly different levels of hypoxia. It also provides a new platform to further assess the mechanistic basis for the hypoxia-induced alterations that occur during metastatic tumor progression in vivo. In addition, while this model was established using ES cells, we anticipate that this experimental strategy can be used to test the effect of hypoxia in other sarcomas, as well as tumors orthotopically implanted in sites with a well-defined blood supply route.

In Vivo Model for Testing Effect of Hypoxia on Tumor Metastasis

This manuscript describes the development of an animal model that allows for the direct testing of the effects of tumor hypoxia on metastasis and the deciphering the mechanisms of its action. Although the experiments described here focus on Ewing sarcoma, a similar approach can be applied to other tumor types.

体内模型对缺氧对肿瘤转移的影响测试

Hypoxia has been implicated in the metastasis of Ewing sarcoma (ES) by clinical observations and in vitro data, yet direct evidence for its pro-metastatic ...

科研

癌症研究

A Murine Orthotopic Bladder Tumor Model and Tumor Detection System

This protocol describes the generation of bladder tumors in female C57BL/6J mice using the murine bladder cancer cell line MB49, which has been modified to secrete human Prostate Specific Antigen (PSA), and the procedure for the confirmation of tumor implantation. In brief, mice are anesthetized using injectable drugs and are made to lay in the dorsal position. Urine is vacated from the bladder and 50 &#181;L of poly-L-lysine (PLL) is slowly instilled at a rate of 10 &#181;L/20 s using a 24 G IV catheter. It is left in the bladder for 20 min by stoppering the catheter. The catheter is removed and PLL is vacated by gentle pressure on the bladder. This is followed by instillation of the murine bladder cancer cell line (1 x 105 cells/50 &#181;L) at a rate of 10 &#181;L/20 s. The catheter is stoppered to prevent premature evacuation. After 1 h, the mice are revived with a reversal drug, and the bladder is vacated. The slow instillation rate is important, as it reduces vesico-ureteral reflux, which can cause tumors to occur in the upper urinary tract and in the kidneys. The cell line should be well re-suspended to reduce clumping of cells, as this can lead to uneven tumor sizes after implantation.
This technique induces tumors with high efficiency. Tumor growth is monitored by urinary PSA secretion. PSA marker monitoring is more reliable than ultrasound or fluorescence imaging for the detection of the presence of tumors in the bladder. Tumors in mice generally reach a maximum size that negatively impacts health by about 3 - 4 weeks if left untreated. By monitoring tumor growth, it is possible to differentiate mice that were cured from those that were not successfully implanted with tumors. With only end-point analysis, the latter may be mistakenly assumed to have been cured by therapy.

This protocol describes the generation of murine orthotopic bladder tumors in female C57BL/6J mice and the monitoring of tumor growth.

小鼠原位膀胱肿瘤模型和肿瘤检测系统

This protocol describes the generation of bladder tumors in female C57BL/6J mice using the murine bladder cancer cell line MB49, which has been modified ...

Isolation of Circulating Tumor Cells in an Orthotopic Mouse Model of Colorectal Cancer

Despite the advantages of easy applicability and cost-effectiveness, subcutaneous mouse models have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Orthotopic mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in hollow organs such as the large bowel. In order to produce uniform tumors which reliably grow and metastasize, standardized techniques of tumor cell preparation and injection are critical. 
We have developed an orthotopic mouse model of colorectal cancer (CRC) which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor cells from either primary tumors, 2-dimensional (2D) cell lines or 3-dimensional (3D) organoids are injected into the cecum and, depending on the metastatic potential of the injected tumor cells, form highly metastatic tumors. In addition, CTCs can be found regularly. We here describe the technique of tumor cell preparation from both 2D cell lines and 3D organoids as well as primary tumor tissue, the surgical and injection techniques as well as the isolation of CTCs from the tumor-bearing mice, and present tips for troubleshooting.

We describe the establishment of orthotopic colorectal tumors via injection of tumor cells or organoids into the cecum of mice and the subsequent isolation of circulating tumor cells (CTCs) from this model.

在直肠癌原位小鼠模型中循环肿瘤细胞的分离

Despite the advantages of easy applicability and cost-effectiveness, subcutaneous mouse models have severe limitations and do not accurately simulate ...

Acellular and Cellular Lung Model to Study Tumor Metastasis

It is difficult to isolate tumor cells at different points of tumor progression. We created an ex vivo lung model that can show the interaction of tumor cells with a natural matrix and continual flow of nutrients, as well as a model that shows the interaction of tumor cells with normal cellular components and a natural matrix. The acellular ex vivo lung model is created by isolating a rat heart-lung block and removing all the cells using the decellularization process. The right main bronchus is tied off and tumor cells are placed in the trachea by a syringe. The cells move and populate the left lung. The lung is then placed in a bioreactor where the pulmonary artery receives a continual flow of media in a closed circuit. The tumor grown on the left lung is the primary tumor. The tumor cells that are isolated in the circulating media are circulating tumor cells and the tumor cells in the right lung are metastatic lesions. The cellular ex vivo lung model is created by skipping the decellularization process. Each model can be used to answer different research questions.

Here, we present a protocol for an ex vivo lung cancer model that mimics the steps of tumor progression and helps to isolate a primary tumor, circulating tumor cells, and metastatic lesions.

脱细胞和细胞肺模型研究肿瘤转移

It is difficult to isolate tumor cells at different points of tumor progression. We created an ex vivo lung model that can show the interaction of tumor ...

Intramucosal Inoculation of Squamous Cell Carcinoma Cells in Mice for Tumor Immune Profiling and Treatment Response Assessment

Head and neck squamous cell carcinoma (HNSCC) is a debilitating and deadly disease with a high prevalence of recurrence and treatment failure. To develop better therapeutic strategies, understanding tumor microenvironmental factors that contribute to the treatment resistance is important. A major impediment to understanding disease mechanisms and improving therapy has been a lack of murine cell lines that resemble the aggressive and metastatic nature of human HNSCCs. Furthermore, a majority of murine models employ subcutaneous implantations of tumors which lack important physiological features of the head and neck region, including high vascular density, extensive lymphatic vasculature, and resident mucosal flora. The purpose of this study is to develop and characterize an orthotopic model of HNSCC. We employ two genetically distinct murine cell lines and established tumors in the buccal mucosa of mice. We optimize collagenase-based tumor digestion methods for the optimal recovery of single cells from established tumors. The data presented here show that mice develop highly vascularized tumors that metastasize to regional lymph nodes. Single-cell multiparametric mass cytometry analysis shows the presence of diverse immune populations with myeloid cells representing the majority of all immune cells. The model proposed in this study has applications in cancer biology, tumor immunology, and preclinical development of novel therapeutics. The resemblance of the orthotopic model to clinical features of human disease will provide a tool for enhanced translation and improved patient outcomes.

Here we present a reproducible method for developing an orthotopic murine model of head and neck squamous cell carcinoma. Tumors demonstrate clinically relevant histopathological features of the disease, including necrosis, poor differentiation, nodal metastases, and immune infiltration. Tumor-bearing mice develop clinically relevant symptoms including dysphagia, jaw displacement, and weight loss.

小鼠鳞状细胞细胞的肿瘤免疫分析及治疗反应评价

Head and neck squamous cell carcinoma (HNSCC) is a debilitating and deadly disease with a high prevalence of recurrence and treatment failure. To develop ...

Patient-derived Orthotopic Xenograft Models for Human Urothelial Cell Carcinoma and Colorectal Cancer Tumor Growth and Spontaneous Metastasis

Cancer patients have poor prognoses when lymph node (LN) involvement is present in both high-grade urothelial cell carcinoma (HG-UCC) of the bladder and colorectal cancer (CRC). More than 50% of patients with muscle-invasive UCC, despite curative therapy for clinically-localized disease, will develop metastases and die within 5 years, and metastatic CRC is a leading cause of cancer-related deaths in the US. Xenograft models that consistently mimic UCC and CRC metastasis seen in patients are needed. This study aims to generate patient-derived orthotopic xenograft (PDOX) models of UCC and CRC for primary tumor growth and spontaneous metastases under the influence of LN stromal cells mimicking the progression of human metastatic diseases for drug screening. Fresh UCC and CRC tumors were obtained from consented patients undergoing resection for HG-UCC and colorectal adenocarcinoma, respectively. Co-inoculated with LN stromal cell (LNSC) analog HK cells, luciferase-tagged UCC cells were intra-vesically (IB) instilled into female non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, and CRC cells were intra-rectally (IR) injected into male NOD/SCID mice. Tumor growth and metastasis were monitored weekly using bioluminescence imaging (BLI). Upon sacrifice, primary tumors and mouse organs were harvested, weighed, and formalin-fixed for Hematoxylin and Eosin and immunohistochemistry staining. In our unique PDOX models, xenograft tumors resemble patient pre-implantation tumors. In the presence of HK cells, both models have high tumor implantation rates measured by BLI and tumor weights, 83.3% for UCC and 96.9% for CRC, and high distant organ metastasis rates (33.3% detected liver or lung metastasis for UCC and 53.1% for CRC). In addition, both models have zero mortality from the procedure. We have established unique, reproducible PDOX models for human HG-UCC and CRC, which allow for tumor formation, growth, and metastasis studies. With these models, testing of novel therapeutic drugs can be performed efficiently and in a clinically-mimetic manner.

This protocol describes the generation of patient-derived orthotopic xenograft models by intra-vesically instilling high-grade urothelial cell carcinoma cells or intra-rectally injecting colorectal cancer cells into non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice for primary tumor growth and spontaneous metastases under the influence of lymph node stromal cells, which mimics the progression of human metastatic diseases.

患者衍生的人类泌尿细胞癌和结肠直肠癌肿瘤生长和自发转移的正交异种移植模型

Cancer patients have poor prognoses when lymph node (LN) involvement is present in both high-grade urothelial cell carcinoma (HG-UCC) of the bladder and ...

Micromanipulation of Circulating Tumor Cells for Downstream Molecular Analysis and Metastatic Potential Assessment

Blood-borne metastasis accounts for&#160;most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new tumors at distant sites. CTCs are found in the bloodstream of patients as single cells (single CTCs) or as multicellular aggregates (CTC clusters and CTC-white blood cell clusters), with the latter displaying a higher metastatic ability. Beyond enumeration, phenotypic and molecular analysis is extraordinarily important to dissect CTC biology and to identify actionable vulnerabilities. Here, we provide a detailed description of a workflow that includes CTC immunostaining and micromanipulation, ex vivo culture to assess&#160;proliferative and survival capabilities of individual cells, and in vivo metastasis-formation assays. Additionally, we provide a protocol to achieve the dissociation of CTC clusters into individual cells and the investigation of intra-cluster heterogeneity. With these approaches, for instance, we precisely quantify survival and proliferative potential of single CTCs and individual cells within CTC clusters, leading us to the observation that cells within clusters display better survival and proliferation in ex vivo cultures compared to single CTCs. Overall, our workflow offers a platform to dissect the characteristics of CTCs at the single cell level, aiming towards the identification of metastasis-relevant pathways and a better understanding of CTC biology.

Here, we present an integrated workflow to identify phenotypic and molecular features that characterize circulating tumor cells (CTCs). We combine live immunostaining and robotic micromanipulation of single and clustered CTCs with single cell-based techniques for downstream analysis and assessment of metastasis-seeding ability.

循环肿瘤细胞的微操作, 用于下游分子分析和转移电位评估

Blood-borne metastasis accounts for&#160;most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new ...

Multidimensional Coculture System to Model Lung Squamous Carcinoma Progression

Tumor-stroma interactions play a critical role in the development of lung squamous carcinoma (LUSC). However, understanding how these dynamic interactions contribute to tissue architectural changes observed during tumorigenesis remains challenging due to the lack of appropriate models. In this protocol, we describe the generation of a 3D coculture model using a LUSC primary cell culture known as TUM622. TUM622 cells were established from a LUSC patient-derived xenograft (PDX) and have the unique property to form acinar-like structures when seeded in a basement membrane matrix. We demonstrate that TUM622 acini in 3D coculture recapitulate key features of tissue architecture during LUSC progression as well as the dynamic interactions between LUSC cells and components of the tumor microenvironment (TME), including the extracellular matrix (ECM) and cancer-associated fibroblasts (CAFs). We further adapt our principal 3D culturing protocol to demonstrate how this system could be utilized for various downstream analyses. Overall, this organoid model creates a biologically rich and adaptable platform that enables one to gain insight into the cell-intrinsic and extrinsic mechanisms that promote the disruption of epithelial architectures during carcinoma progression and will aid the search for new therapeutic targets and diagnostic markers.

An in vitro model system was developed to capture tissue architectural changes during lung squamous carcinoma (LUSC) progression in a 3-dimensional (3D) co-culture with cancer-associated fibroblasts (CAFs). This organoid system provides a unique platform to investigate the roles of diverse tumor cell-intrinsic and extrinsic changes that modulate the tumor phenotype.

多维共同培养系统模拟肺鳞癌进展

Tumor-stroma interactions play a critical role in the development of lung squamous carcinoma (LUSC). However, understanding how these dynamic interactions ...

Pathological Analysis of Lung Metastasis Following Lateral Tail-Vein Injection of Tumor Cells

Metastasis, the primary cause of morbidity and mortality for most cancer patients, can be challenging to model preclinically in mice. Few spontaneous metastasis models are available. Thus, the experimental metastasis model involving tail-vein injection of suitable cell lines is a mainstay of metastasis research. When cancer cells are injected into the lateral tail-vein, the lung is their preferred site of colonization. A potential limitation of this technique is the accurate quantification of the metastatic lung tumor burden. While some investigators count macrometastases of a pre-defined size and/or include micrometastases following sectioning of tissue, others determine the area of metastatic lesions relative to normal tissue area. Both of these quantification methods can be exceedingly difficult when the metastatic burden is high. Herein, we demonstrate an intravenous injection model of lung metastasis followed by an advanced method for quantifying metastatic tumor burden using image analysis software. This process allows for investigation of multiple end-point parameters, including average metastasis size, total number of metastases, and total metastasis area, to provide a comprehensive analysis. Furthermore, this method has been reviewed by a veterinary pathologist board-certified by the American College of Veterinary Pathologists (SEK) to ensure accuracy.

Intravenous injection of cancer cells is often used in metastasis research, but the metastatic tumor burden can be difficult to analyze. Herein, we demonstrate a tail-vein injection model of metastasis and include a novel approach to analyze the resulting metastatic lung tumor burden.

肿瘤细胞外侧尾静脉注射后肺转移的病理分析

Metastasis, the primary cause of morbidity and mortality for most cancer patients, can be challenging to model preclinically in mice. Few spontaneous ...

Modeling Primary Bone Tumors and Bone Metastasis with Solid Tumor Graft Implantation into Bone

Primary bone tumors or bone metastasis from solid tumors result in painful osteolytic, osteoblastic, or mixed osteolytic/osteoblastic lesions. These lesions compromise bone structure, increase the risk of pathologic fracture, and leave patients with limited treatment options. Primary bone tumors metastasize to distant organs, with some types capable of spreading to other skeletal sites. However, recent evidence suggests that with many solid tumors, cancer cells that have spread to bone may be the primary source of cells that ultimately metastasize to other organ systems. Most syngeneic or xenograft mouse models of primary bone tumors involve intra-osseous (orthotopic) injection of tumor cell suspensions. Some animal models of skeletal metastasis from solid tumors also depend on direct bone injection, while others attempt to recapitulate additional steps of the bone metastatic cascade by injecting cells intravascularly or into the organ of the primary tumor. However, none of these models develop bone metastasis reliably or with an incidence of 100%. In addition, direct intra-osseous injection of tumor cells has been shown to be associated with potential tumor embolization of the lung. These embolic tumor cells engraft but do not recapitulate the metastatic cascade. We reported a mouse model of osteosarcoma in which fresh or cryopreserved tumor fragments (consisting of tumor cells plus stroma) are implanted directly into the proximal tibia using a minimally invasive surgical technique. These animals developed reproducible engraftment, growth, and, over time, osteolysis and lung metastasis. This technique has the versatility to be used to model solid tumor bone metastasis and can readily employ grafts consisting of one or multiple cell types, genetically-modified cells, patient-derived xenografts, and/or labeled cells that can be tracked by optical or advanced imaging. Here, we demonstrate this technique, modeling primary bone tumors and bone metastasis using solid tumor graft implantation into bone.

Bone metastasis models do not develop metastasis uniformly or with a 100% incidence. Direct intra-osseous tumor cell injection can result in embolization of the lung. We present our technique modeling primary bone tumors and bone metastasis using solid tumor graft implantation into bone, leading to reproducible engraftment and growth.

通过实体瘤移植植入骨模拟原发性骨肿瘤和骨转移

Primary bone tumors or bone metastasis from solid tumors result in painful osteolytic, osteoblastic, or mixed osteolytic/osteoblastic lesions. These ...

原发肿瘤发展小鼠实验模型和腹膜转移

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