The inflammatory cytokines TNFa and IL1b lead to poor prognosis in breast cancer. Here, we determined the possibility that TNFa and IL1b, that are highly expressed by breast tumor cells, induce Epithelial-to-Mesenchymal Transition (EMT) and invasive properties in non-transformed breast epithelial cells adjacent to the tumor cells. Our previous studies have shown that 3 days of stimulation by TNFa has led to EMT in breast cancer cells. In the present study, we found that EMT of non-transformed breast cells required 3 weeks stimulation by TNFa+IL1b, and was evidenced by formation of cellular protrusions, E-cadherin reduction and vimentin up-regulation. The EMT-regulating transcription factors Zeb1 and Snail were up-regulated only after long-term stimulation with TNFa+IL1b, while Twist was down-regulated upon the long-term stimulation. The delayed activation of Zeb1 and Snail and inhibition of Twist caused the delayed EMT process. In the absence of Twist activation, the cytokines TNFa+IL1b took over, and have potentiated the EMT-inducing abilities of Zeb1 and Snail, by that recapitulating the activities of Twist.
Importantly, the EMT process induced by long-term stimulation with TNFa+IL1b has indeed led to increased invasion-related activities in the non-transformed cells. This has been evidenced by increased release of MMPs, augmented migration and invasion properties of these cells.
Our findings identify novel pro-malignancy roles for TNFa and IL1b, showing that they induce EMT and invasive properties in non-transformed cells, mediated by Zeb1 and Snail, and by recapitulating the activities of Twist. In patients, such processes may cause re-seeding of the non-transformed cells at the primary tumor site, where they may undergo transformation, thus contributing to formation of a secondary tumor at the primary site. This secondary tumor is expected to carry a different genetic setup than the primary tumor, as was observed in approximately 25% of breast cancer patients with recurrent disease.
