Abstract
Cancer-associated fibroblasts (CAFs) play a critical role in tumor progression and are associated with high-grade malignancy and hindrance of anti-cancer pharmacotherapy. While it is observed that CAFs exhibit myofibroblast-like phenotypes and are highly contractile, the process by which normal fibroblasts (NFs) are transformed into CAFs is yet to be elucidated. To gain insight about this process, we constructed a coculture system where GFP-labeled NIH3T3 fibroblasts were cocultured with cancer or normal mammary gland epithelial cells, respectively. The dynamic interactions between fibroblasts and epithelial cells were recorded by timelapse microscopy for 20 hours upon seeding and tracked using image correlation analysis. The results showed that fibroblasts cocultured with cancer epithelial cells exhibited different cell mobility profiles compared to normal controls. Furthermore, pattern recognition-based image analysis revealed that fibroblasts/cancer epithelial cell coculture produced lesion-like structures whereas the control coculture produced gland-like structures. Fibroblasts were then microdissected from the cocultures to examine the expression level of various CAF-related proteins in Western blot. The results showed that myosin II phosphorylation is significantly upregulated in fibroblasts cocultured with cancer epithelial cells, indicating NF-CAF transformation can occur within 20 hours after NFs are in contact with cancer cells.View Large Image | View Hi-Res Image | Download PowerPoint Slide
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