Mesenchymal-epithelial transition and stemness in sarcoma

Clinical evidence reveals that upregulation of epithelial markers is prognostic for better survival in patients with sarcoma, while mesenchymal markers are prognostic for worse survival. We previously observed that forced induction of mesenchymal-epithelial transition (MET) not only decreased the cancer-like aggression of rhabdomyosarcoma (RD) cells in vitro, but also produced downregulation of key stem cell marker Zeb1. From this result, we hypothesized that MET reduces aggression in sarcoma by inhibiting stemness. If this is the case, then inducing differentiation without a concurrent MET should produce the same reduction in aggression we observe following forced MET induction. Here we attempted to address this possibility by inducing differentiation and testing for aggression and MET induction, and by isolating stem-like cells and testing for aggression, MET induction, and differentiation. Our use of all-trans retinoic acid (ATRA) to induce differentiation in RD cells caused a decrease in Zeb1 expression, but did not reliably increase expression of muscle differentiation genes. Unexpectedly, ATRA treatment also caused an increase in anchorage-independent growth. RD stem-like cells isolated via Hoechst dye exclusion exhibited increased expression of Zeb1, but did not show increased expression of muscle differentiation markers. Isolated stem-like cells were also not better at forming clones. Overall, our results do not provide evidence that a reduction in stemness is related to the reduction in aggression observed following MET. Contrary to expectation, we cannot rule out the possibility that an increase in stemness may instead be associated with reduced aggression in sarcoma.