Palmer Trenis D, Lewis John, Zijlstra Andries
Department of Pathology, Vanderbilt University.
J Vis Exp. 2011 May 30(51):2815. doi: 10.3791/2815.
During metastasis cancer cells disseminate from the primary tumor, invade into surrounding tissues, and spread to distant organs. Metastasis is a complex process that can involve many tissue types, span variable time periods, and often occur deep within organs, making it difficult to investigate and quantify. In addition, the efficacy of the metastatic process is influenced by multiple steps in the metastatic cascade making it difficult to evaluate the contribution of a single aspect of tumor cell behavior. As a consequence, metastasis assays are frequently performed in experimental animals to provide a necessarily realistic context in which to study metastasis. Unfortunately, these models are further complicated by their complex physiology. The chick embryo is a unique in vivo model that overcomes many limitations to studying metastasis, due to the accessibility of the chorioallantoic membrane (CAM), a well-vascularized extra-embryonic tissue located underneath the eggshell that is receptive to the xenografting of tumor cells (figure 1). Moreover, since the chick embryo is naturally immunodeficient, the CAM readily supports the engraftment of both normal and tumor tissues. Most importantly, the avian CAM successfully supports most cancer cell characteristics including growth, invasion, angiogenesis, and remodeling of the microenvironment. This makes the model exceptionally useful for the investigation of the pathways that lead to cancer metastasis and to predict the response of metastatic cancer to new potential therapeutics. The detection of disseminated cells by species-specific Alu PCR makes it possible to quantitatively assess metastasis in organs that are colonized by as few as 25 cells. Using the Human Epidermoid Carcinoma cell line (HEp3) we use this model to analyze spontaneous metastasis of cancer cells to distant organs, including the chick liver and lung. Furthermore, using the Alu-PCR protocol we demonstrate the sensitivity and reproducibility of the assay as a tool to analyze and quantitate intravasation, arrest, extravasation, and colonization as individual elements of metastasis.
在转移过程中，癌细胞从原发肿瘤扩散，侵入周围组织，并扩散到远处器官。转移是一个复杂的过程，可能涉及多种组织类型，跨越不同的时间段，且常常发生在器官深处，这使得研究和量化变得困难。此外，转移过程的有效性受到转移级联反应中多个步骤的影响，因此难以评估肿瘤细胞行为的单个方面的作用。因此，转移实验经常在实验动物中进行，以提供一个必要的现实环境来研究转移。不幸的是，这些模型因其复杂的生理学特性而进一步复杂化。鸡胚是一种独特的体内模型，由于绒毛尿囊膜（CAM）易于获取，它克服了研究转移的许多局限性。绒毛尿囊膜是位于蛋壳下方的一种血管丰富的胚外组织，易于接受肿瘤细胞的异种移植（图1）。此外，由于鸡胚天然免疫缺陷，绒毛尿囊膜很容易支持正常组织和肿瘤组织的植入。最重要的是，禽类绒毛尿囊膜成功支持大多数癌细胞特征，包括生长、侵袭、血管生成和微环境重塑。这使得该模型对于研究导致癌症转移的途径以及预测转移性癌症对新的潜在治疗方法的反应特别有用。通过物种特异性Alu PCR检测播散细胞，使得定量评估少至25个细胞定植的器官中的转移成为可能。我们使用人表皮样癌细胞系（HEp3），利用这个模型分析癌细胞向远处器官（包括鸡的肝脏和肺）的自发转移。此外，使用Alu-PCR方案，我们证明了该检测方法作为分析和定量转移的各个要素（包括内渗、滞留、外渗和定植）的工具的敏感性和可重复性。
