Assessing estrogen-induced proliferative response in an endometrial cancer cell line using a universally applicable methodological guide

Abstract

Objective: Translational endometrial cancer (EC) research benefits from an in vitro experimental approach using EC cell lines. We demonstrated the steps that are required to examine estrogen induced proliferative response, a simple yet important research question pertinent to EC and devised a pragmatic methodological workflow for utilising EC cell lines in experimental models. Methods/materials: Comprehensive review of all commercially available EC cell lines was carried out, and Ishikawa cell line was selected to study the oestrogen responsiveness with HEC1A, RL95-2 and MFE280 cell lines as comparators where appropriate, examining relevant differential molecular (steroid receptors) and functional (phenotype, anchorage-independent growth, hormone responsiveness, migration, invasion and chemosensitivity) characteristics in 2D and 3D cultures in vitro using immunocytochemistry, immunofluorescence, qPCR and western blotting. In vivo tumour, formation and chemosensitivity were also assessed in a chick chorioallantoic membrane (CAM) model. Results: Short Tandem Repeat (STR) analysis authenticated the purchased cell lines while gifted cells deviated significantly from the published profile. We demonstrate the importance of prior assessment of the suitability of each cell line for the chosen in vitro experimental technique. Prior establishment of baseline, non-enriched conditions was required to induce a proliferative response to estrogen. The CAM model was a suitable in vivo multi-cellular animal model for EC, for producing rapid and reproducible data. Conclusions: We have developed a methodological guide for EC researchers when using endometrial cell lines to answer important translational research questions (exemplified by estrogen responsive cell proliferation), to facilitate robust data, while saving time and resources.