Time-lapse analyses of epithelial and mesenchymal pancreatic ductal adenocarcinoma sphere formation and spread on plates

Abstract

Abstract Background The most pancreatic cancers are pancreatic ductal adenocarcinomas (PDAC), which have been classified as cells with epithelial or mesenchymal features. We recently found that the morphology of spheres formed in three-dimensional cultures and the effect of anticancer drugs differed between epithelial and mesenchymal PDAC cell lines. Cancer forms a mass and metastasizes in the human body; however, there are differences in the ability of epithelial and mesenchymal PDAC to invade the surrounding area. Methods Sphere formation and the ability of spheres to adhere and spread on culture plates were examined by time-lapse imaging using five epithelial and three mesenchymal PDAC cell lines. Results Immunocytochemical staining confirmed that E-cadherin was highly expressed in epithelial PDAC spheres, and vimentin was highly expressed in mesenchymal PDAC spheres. In the process of forming spheres using ultra-low attachment plates, the majority of epithelial PDAC cell lines showed an early decrease in sphere area. When this decrease ceased, the covering cells on the sphere surface fused to form a smooth surface. Mesenchymal PANC-1 and MIA PaCa-2 cells showed decreased reduction in the spherical area and few fused surface areas. When the formed PDAC spheres were seeded onto normal plates, the spheres of PK-8 cells with the highest E-cadherin expression did not adhere to the normal plates, even after 60 h, and epithelial PK45-P and T3M-4 spheres hardly adhered. In contrast, the area of adhesion and spread of mesenchymal PANC-1 and KP4 cell spheres on normal plates markedly increased from the early period. Conclusions Three-dimensional cultures of epithelial or mesenchymal PDAC spheres play an important role in examining the diversity of tumor formation and invasion in pancreatic cancer.