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Updated: Jun 15, 2026
In Vitro Evaluation of Oncogenic Transformation in Human Mammary Epithelial Cells
Published on: September 24, 2020
Novel in vitro assays for the characterization of EMT in tumourigenesis
Vincent Koo1, Amgad El Mekabaty, Peter Hamilton
1Uro-oncology Research Group, Centre for Cancer Research & Cell Biology, Queen's University Belfast, Belfast, Northern Ireland, UK.
Background:
Two novel assays quantifying Epithelial to Mesenchymal Transition (EMT) were compared to traditional motility and migration assays. TGF-beta1 treatment of AY-27 rat bladder cancer cells acted as a model of EMT in tumourigenesis.
Methods:
AY-27 rat bladder cancer cells incubated with 3 ng/ml TGF-beta1 or control media for 24 or 48 h were assessed using novel and traditional assays. The Spindle Index, a novel measure of spindle phenotype, was derived from the ratio of maximum length to maximum width of cells. The area covered by cells which migrated from a fixed coverslip towards supplemented agarose was measured in a novel chemoattractant assay. Motility, migration and immunoreactivity for E-cadherin, Vimentin and cytokeratin were assessed.
Results:
TGF-beta1 treated cells had increased "spindle" phenotype together with decreased E-cadherin, decreased Cytokeratin-18 and increased Vimentin immunoreactivity. After 48 h, the mean Spindle Index of TGF-beta1 treated cells was significantly higher than Mock (p=0.02, Bonferroni test) and there were significant differences in migration across treatment groups measured using the novel chemoattractant assay (p=0.02, Chi-square). TGF-beta1 significantly increased matrigel invasion.
Conclusion:
The Spindle Index and the novel chemoattractant assay are valuable adjunctive assays for objective characterization of EMT changes during tumourigenesis.
