Bilandzic Maree, Stenvers Kaye L
Reproductive Development and Cancer Laboratory, MIMR-PHI Institute of Medical Research;
Reproductive Development and Cancer Laboratory, MIMR-PHI Institute of Medical Research; Department of Developmental Biology and Anatomy, Monash University.
J Vis Exp. 2014 May 20(87):51655. doi: 10.3791/51655.
Ovarian cancers metastasize by shedding into the peritoneal fluid and dispersing to distal sites within the peritoneum. Monolayer cultures do not accurately model the behaviors of cancer cells within a nonadherent environment, as cancer cells inherently aggregate into multicellular structures which contribute to the metastatic process by attaching to and invading the peritoneal lining to form secondary tumors. To model this important stage of ovarian cancer metastasis, multicellular aggregates, or spheroids, can be generated from established ovarian cancer cell lines maintained under nonadherent conditions. To mimic the peritoneal microenvironment encountered by tumor cells in vivo, a spheroid-mesothelial co-culture model was established in which preformed spheroids are plated on top of a human mesothelial cell monolayer, formed over an extracellular matrix barrier. Methods were then developed using a real-time cell analyzer to conduct quantitative real time measurements of the invasive capacity of different ovarian cancer cell lines grown as spheroids. This approach allows for the continuous measurement of invasion over long periods of time, which has several advantages over traditional endpoint assays and more laborious real time microscopy image analyses. In short, this method enables a rapid, determination of factors which regulate the interactions between ovarian cancer spheroid cells invading through mesothelial and matrix barriers over time.
卵巢癌通过脱落至腹腔积液并扩散至腹膜内的远端部位发生转移。单层培养不能准确模拟癌细胞在非粘附环境中的行为，因为癌细胞会自然聚集形成多细胞结构，这些结构通过附着并侵入腹膜内衬形成继发性肿瘤，从而促进转移过程。为了模拟卵巢癌转移的这一重要阶段，可以从在非粘附条件下培养的既定卵巢癌细胞系中生成多细胞聚集体或球体。为了模拟肿瘤细胞在体内遇到的腹膜微环境，建立了一种球体-间皮细胞共培养模型，其中预先形成的球体接种在人腹膜间皮细胞单层上，该单层细胞在细胞外基质屏障上形成。然后开发了使用实时细胞分析仪对作为球体生长的不同卵巢癌细胞系的侵袭能力进行定量实时测量的方法。这种方法允许长时间连续测量侵袭情况，与传统的终点分析以及更费力的实时显微镜图像分析相比具有多个优点。简而言之，该方法能够快速确定随着时间推移调节卵巢癌球体细胞通过间皮和基质屏障进行侵袭的相互作用的因素。
