一个<em在体外系统来研究肿瘤休眠和转移性增长
Summary
修改后的3 – D在体外培养系统是其中的几个重组基底膜的肿瘤细胞株的生长特性与休眠或增生的肿瘤细胞转移的辅助站点的行为相关<em>在体内</em>。
Abstract
乳腺癌复发往往遵循一个潜伏期长，其中有没有癌症的迹象，并转移可能不会成为多年后切除原发肿瘤的辅助治疗，直到临床明显。这种现象的一个可能的解释是，肿瘤细胞有种子的转移部位，耐常规疗法，并保持时间 1-4长时间处于休眠状态。
以前在辅助站点休眠癌细胞的存在已被描述为静态，既不增殖也不是孤立发生凋亡 5-7的细胞。此外，这些孤立的细胞已被证明传播从原发肿瘤的早期疾病进展8-10的阶段，并驻留在病人的骨髓，血液和淋巴结1,4,11中生长被捕。因此，了解的机制，规范增殖状态的休眠或交换机是发现新的目标和干预措施，以防止疾病的复发的关键。然而，揭开肿瘤休眠开关调节转移性增长的机制已经阻碍了缺乏可用的模型系统。
在体内和体外模型系统研究进展肿瘤细胞的转移已被描述以前 1,12-14 。然而，这些模型系统并没有提供实时和高通量的方式进入孤处于休眠状态的肿瘤细胞增殖转移性疾病的出现触发什么机械的见解。最近，我们在体外系统开发的一个三维模型 ， 在细胞体内生长的特点，表现出休眠（D2.OR MCF7，K7M2 – AS.46）或增生（D2A1，MDA – MB – 231，K7M2）转移性行为在体内 。我们证明了，表现在体内肿瘤细胞在转移部位的休眠保持静态时，在一个3维（3D）基底膜提取物（BME）培养，而高度在体内转移的细胞很容易在三维培养增殖后，变量，但相对较短时期的平静。重要的是利用体外模型系统中的三维，我们首次展示了ECM的成分，起着重要的调节是否处于休眠状态的肿瘤细胞会切换到增殖状态，并确认 在体内研究15-17作用。因此，本报告中所描述的模型系统提供了在体外培养的方法，以模型的肿瘤休眠研究的微环境诱导的增殖生长的过渡。
Protocol
Discussion
的基本机制，保持传播的肿瘤细胞处于休眠状态或在其过渡到转移性增长的结果仍是未知。这种现象已经非常困难的研究在人类患者4,12和少数临床前模型已开发来解决这个问题。然而， 在体内和体外模型系统，为肿瘤休眠的 一些特点 （1,12审查）。然而， 在体内肿瘤休眠模式主要用来验证规范肿瘤休眠的 潜在机制，但不适合进行实时探索一个单一?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项研究是由国家癌症研究所的院内研究计划中的一部分支持。
Materials
| Name of the reagent | Company | Catalogue number | Comments |
|---|---|---|---|
| DMEM high glucose | Invitrogen | 11965-118 | |
| DMEM low glucose | Invitrogen | 11885-092 | |
| Fetal bovine serum (FBS) | Invitrogen | 10091-148 | |
| Growth factor-reduced 3-D Cultrex Basement Membrane Extract | Trevigen Inc. | Protein concentration between 14-15mg/ml | |
| D2.0R and D2A1 cell lines | 5,19 | ||
| K7M2 and K7M2AS1.46 cells | 20 | ||
| MCF-7 and MDA-MB-231 breast cancer cells | ATCC | ||
| An 8 chamber glass slide system | (Lab -TEK, Thermo scientific) | 177402 | |
| Cell Titer 96 AQueous One Solution cell proliferation assay kit | Promega | G3580 | |
| VECTASHIELD mounting medium with DAPI | Vector Laboratories Inc. | H-1200 | |
| Normal donkey serum | Jackson ImmunoResearch | 017-000-121 | |
| Elisa Plate Reader | Bio-Tec | Record 490nm | |
| Confocal microscope | Zeiss-LSM-510 | Magnification x63 |
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