Effect of Varying Mechanical Environment in 2D Culture on Subsequent Metastasis Process of Ovarian Cancer

Abstract

The epithelial ovarian cancer metastasizes through a unique transcoelomic routes, wherein cancer cells crumble out to body cavity, survive in an anchorage-independent manner as spheroids, and then finally stick to the outermost layer of another organ to develop a secondary tumor. The later stage mechanisms have already been elucidated in vitro and in vivo recently. Comparatively, only few clues exist to explain what happens during the early stage of metastasis. Recent studies reported marked heterogeneities in junctional proteins and cytoskeleton composition even within a single spheroid. Also, some of these intra-tumor heterogeneities are known to be sustained throughout the whole metastasis steps. Since the intra-tumor heterogeneity arises at that stage, herein, we focused on varying the mechanical environment of early-stage to induce cancer cells to have different cytoskeletal composition and so on. We cultured a non-serous high-grade ovarian cancer cell line, SK-OV-3, on polyacrylamide gels of different stiffness to establish two groups of distinct phenotypes. The difference between these two population of cells was characterized by immunostaining and migratory phenotype. To mimic the floating situation in the body cavity, we adopted the hanging drop method to generate the cellular spheroids. Afterward, the cells cultured with different stiffness were made into cellular spheroids, together or separately. The resultant spheroids were also tested with the adhesiveness and spreading ability on a physiologically relevant environment. Through these experimental setting, we examined whether different mechanical microenvironmental conditions in 2D culture, mimicking the heterogeneous conditions of the primary tumor, affect the subsequent spheroidal compaction and dissemination.