  Tumour spheroids have the potential to be used as preclinical
chemosensitivity assays. However, the production of three dimensional (3D)
tumour spheroids remains challenging as not all tumour cell lines form
spheroids with regular morphologies and spheroid transfer often induces
disaggregation. In the field of pancreatic cancer, the MiaPaCa-2 cell line is
an interesting model for research but it is known for its difficulty to form
stable spheroids; also, when formed, spheroids from this cell line are weak and
arduous to manage and to harvest for further analyses such as multiple staining
and imaging. In this work, we compared different methods (i.e. hanging drop,
round bottom wells and Matrigel embedding, each of them with or without
methylcellulose in the media) to evaluate which one allowed to better overpass
these limitations. Morphometric analysis indicated that hanging drop in
presence of methylcellulose leaded to well-organized spheroids; interestingly,
quantitative PCR (qPCR) analysis reflected the morphometric characterization,
indicating that same spheroids expressed the highest values of CD44, VIMENTIN,
TGF beta1 and Ki67. In addition, we investigated the generation of MiaPaCa-2
spheroids when cultured on substrates of different hydrophobicity, in order to
minimize the area in contact with the culture media and to further improve
spheroid formation.
