原发肿瘤发展小鼠实验模型和腹膜转移<em>通过</em>原位接种卵巢癌细胞
Summary
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。
Abstract
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。
Introduction
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。
Protocol
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.悬浮细胞的制备人卵巢透明细胞癌的细胞系。 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℃。 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…
Representative Results
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…
Discussion
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…
Declarações
The authors have nothing to disclose.
Acknowledgements
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。
Materials
| Matrigel matrix | BD | 354234 | ECM-based hydrogel |
| Insulin syringe | TERUMO | SS-05M2913 | 1/2cc, 29Gx1/2" |
| Suturing needle with suture | 11mm, 3/8, Nylon6-0 | ||
| Reflex 7mm Wound Clip Applier | CellPoint Scientific | 204-1000 | |
| Reflex 7mm wound clips | CellPoint Scientific | 203-1000 |
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Citar este artigo
Koya, Y., Kajiyama, H., Liu, W., Shibata, K., Senga, T., Kikkawa, F. Murine Experimental Model of Original Tumor Development and Peritoneal Metastasis via Orthotopic Inoculation with Ovarian Carcinoma Cells. J. Vis. Exp. (118), e54353, doi:10.3791/54353 (2016).