Name: spheroid assay to measure tgf-&#946;-induced invasion
Uploaded: 2011-11-16T13:00:00
Description: Watch this Scientific Journal Video about Spheroid Assay to Measure TGF-ÃŽÂ²-induced Invasion at JoVE.com

我们感谢肯岩田（OSI制药，纽约，美国）试剂和弗雷德米勒（芭芭拉安Karmanos癌症Intitute，底特律，美国）细胞株。

1。 METHOCEL解决方案的准备（500毫升） <ol><li>高压灭菌6克甲基纤维素在500毫升瓶含有磁力搅拌器。 </li><li>甲基为20分钟，在250毫升预热的DMEM/F12无血清（60℃）溶解。 </li><li>加入250毫升的DMEM/F12含10％马血清（RT）的。混合过夜（4℃）。 </li><li>清除离心（5000G，2H，4℃）的解决方案。 </li><li>以明确的高粘度的解决方案到一个新的管（约90-95％的原液）。 </li><li>储存在4℃，直到使用C。 </li></ol><p cla...

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我们建立了一个球体的模型，其中MCF10A1细胞和其恶性衍生物成胶原基质中的TGF -β-依赖性入侵。首先，球体形成96井圆底暂停存在甲基板。当然，也U形聚甲基丙烯酸羟乙酯涂层板，可用于这一目的。甲基纤维素是绝对必要的，在这一步，，因为细胞就会死亡中的甲基纤维素的情况下。 TGF -β是直接添加到胶原METHOCEL混合，因为我们发现，细胞的反应较差的TGF -β时，这个因素是添加胶原蛋白的中型?...

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论（可选） </td></tr><tr><td>甲基纤维素</td><td> Sigma - Aldrich公司</td><td> M0512 </td><td></td></tr><tr><td> PureCol </td><td>高级Biomatrix </td><td> 5005 - B </td><td></td></tr><tr><td> pH指示剂条</td><td>默克公司</td><td> 9533 </td><td></td></tr><tr><td> 96，以及悬挂 板</td><td>格雷纳生物一</td><td> 650 185 </td><td></td></tr><tr><td> 96孔粘连训练班 板</td><td>格雷纳生物一</td><td> 655 180 </td><td></td></tr></tbody></table>

spheroid assay to measure tgf-&#946;-induced invasion

TGF -β有反对乳腺癌进展中的角色，作为一个在初始阶段的抑癌， 但在稍后阶段1,2刺激的侵袭和转移。此外，TGF -β是经常过度表达在乳腺癌中，其表达与预后差， 转移 3,4有关。其中TGF -β诱导的入侵是不能很好地理解的机制。 TGF -β诱发细胞通过TGF -β型II（TβRII）和I型（TβRI）受体反应。当TGF -β诱导的异聚复合物的形成，TβRII磷酸化TβRI。激活TβRI启动Smads的磷酸化受体（R - Smads的），即Smad2的和Smad3的细胞内的规范信号转导通路。这些活化的R - Smads的形式与Smad4的异聚复合物，积聚在细胞核内调节靶基因的转录 5 。除了以前ð旁切Smad通路，在其他几个非Smad信号通路的激活受体激活的结果，例如丝裂原活化蛋白激酶（MAPK）途径6。 要研究的TGF -β在乳腺癌的不同​​阶段中的作用，我们使用的MCF10A电池系统。该系统由自发永生MCF10A1供应量（M1）乳房上皮细胞的H - RAS转化M1衍生MCF10AneoT供应量（M2），在小鼠8癌前病变，生产和M2衍生MCF10CA1a（M4），成立M2的异种移植和形式与能力转移到肺9高年级癌。这MCF10A系列提供了可能性，研究与恶性肿瘤的不同等级是由不同的遗传背景不偏颇细胞的反应。 对于TGF -β诱导的入侵的分析，我们产生的同型MCF10A球体细胞培养EMBEdded在三维胶原基质在体外 （图1）。这种模型非常类似于人类肿瘤在体内建立一个氧气和营养物质的梯度， 造成 10球体内部的外部和静态的，甚至坏死细胞活跃，侵入细胞。球体为基础的检测也已证明，以更好地复述比单层培养 11的耐药性。这MCF10三维模型系统允许我们调查的乳腺细胞的侵入特性，在不同阶段的恶性肿瘤的TGF -β信号的影响。

Watch this Scientific Journal Video about Spheroid Assay to Measure TGF-ÃŽÂ²-induced Invasion at JoVE.com

Spheroid Assay to Measure TGF-ÃƒÅ½Ã‚Â²-induced Invasion

描述一个分析，定量分析转化生长因子（TGF）-β诱导的三维胶原凝胶侵袭。这个实验需要的细胞株MCF10A系列，代表不同阶段乳腺癌的发展优势。通过这种方法可以用于其他细胞系，并可能被用来调查其他潜在的激活或入侵抑制剂。

球体的检测，以测量TGF -β诱导的入侵

TGF-&beta; has opposing roles in breast cancer progression by acting as a tumor suppressor in the initial phase, but stimulating invasion and metastasis ...

Research

JoVE Journal

Medicine

Spheroid Assay to Measure TGF-&#946;-induced Invasion

A Real-time Electrical Impedance Based Technique to Measure Invasion of Endothelial Cell Monolayer by Cancer Cells

一个实时的基于电阻抗技术测量癌细胞入侵内皮细胞单层

Metastatic dissemination of malignant cells requires degradation of basement membrane, attachment of tumor cells to vascular endothelium, retraction of ...

科研

医学

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Modeling and Imaging 3-Dimensional Collective Cell Invasion

3维建模和成像集体细胞的侵袭

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A Modified In vitro Invasion Assay to Determine the Potential Role of Hormones, Cytokines and/or Growth Factors in Mediating Cancer Cell Invasion

A Modified In vitro Invasion Assay to Determine the Potential Role of Hormones, Cytokines and/or Growth Factors in Mediating Cancer Cell Invasion

一种改进<em&gt;在体外</em&gt;入侵检测，以确定激素，在调节肿瘤细胞侵袭的细胞因子和/或生长因子的潜在作用

Blood serum serves as a chemoattractant towards which cancer cells migrate and invade, facilitating their intravasation into microvessels. However, the ...

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在实时的间皮细胞卵巢癌球体附件和入侵评估

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A complex integration of molecular and electrical signals is needed to transform a quiescent uterus into a contractile organ at the end of pregnancy. ...

Three-Dimensional (3D) Tumor Spheroid Invasion Assay

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三维（3D）肿瘤球体侵袭试验

Invasion of surrounding normal tissues is generally considered to be a key hallmark of malignant (as opposed to benign) tumors. For some cancers in ...

Chick Heart Invasion Assay for Testing the Invasiveness of Cancer Cells and the Activity of Potentially Anti-invasive Compounds

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The goal of the chick heart assay is to offer a relevant organ culture method to study tumor invasion in three dimensions. The assay can distinguish ...