Gatesman Ammer Amanda, Hayes Karen E, Martin Karen H, Zhang Lingqing, Spirou George A, Weed Scott A
Department of Neurobiology and Anatomy, Program in Cancer Cell Biology, Mary Babb Randolph Cancer Center, West Virginia University, USA.
J Vis Exp. 2011 Jul 25(53):2941. doi: 10.3791/2941.
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). Fox1(nu/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.
头颈部癌症的局部区域侵犯与转移风险相关，在设计和实施患者管理策略方面构成了一项艰巨挑战。已开发出口腔癌原位小鼠模型，以促进对影响侵犯因素的研究，并作为评估抗肿瘤治疗方法的模型系统。在这些系统中，口腔组织内播散肿瘤细胞的可视化通常通过传统组织学或体内生物发光方法进行。这些技术的一个主要缺点是，固有地无法在三维空间中准确可视化和量化源自原发部位的早期肿瘤细胞侵犯。在此，我们描述了一种方案，该方案将已建立的舌鳞状细胞癌（SCOT）模型与双光子成像相结合，以实现舌部肿瘤扩散的多向量可视化。对OSC-19头颈部肿瘤细胞系进行稳定改造，使其表达与mCherry荧光蛋白融合的F-肌动蛋白结合肽LifeAct（LifeAct-mCherry）。注射这些细胞的Fox1(nu/nu)小鼠可靠地形成肿瘤，通过离体应用双光子显微镜可观察到舌头。该技术可在切除的舌头中对肿瘤块和局部侵犯细胞进行原位可视化，而不会破坏局部肿瘤微环境。此外，该系统可通过计算侵犯细胞从原发肿瘤部位移动的距离来量化肿瘤细胞侵犯。总体而言，该程序提供了一个增强的模型系统，用于分析促成SCOT侵犯的因素以及为预防局部侵犯和远处转移扩散量身定制的治疗方法。该方法最终还有可能在体内环境中与其他成像方式相结合。
