Human umbilical cord mesenchymal stem cells conditioned medium exerts anti-tumor effects on KGN cells in a cell density-dependent manner through activation of the Hippo pathway

Abstract

Background: The conditioned medium from human umbilical cord mesenchymal stem cells (UCMSCs-CM) provides a new cell-free therapy for tumors due to its unique secretome. However, there are many contradictory reports about the effect of UCMSCs-CM on tumor cells. The loss of contact inhibition is a common characteristic of tumor cells. A relationship between the effect of UCMSCs-CM on tumor cells and contact inhibition in tumor cells is rarely concerned. Whether the effect of UCMSCs-CM on tumor cells is affected by cell density? Here, we explored the effect of UCMSCs-CM on granulosa tumor cell line (KGN) cells at low or high density. Methods: Growth curve and CCK8 assay were used to assess cell proliferation and viability. Scratch wound and matrigel invasion assay were implicated to detect cell motility of KGN cells. UCMSCs-CM effects on cell cycle, apoptosis and pathway-related proteins were investigated by flow cytometry, TUNEL assay, western blot and immunofluorescence analysis respectively. Results: In growth curve analysis, before KGN cells proliferated into confluence, UCMSCs-CM had no effect on cell proliferation. However, once the cells proliferate to contact each other, UCMSCs-CM significantly inhibited proliferation. Meanwhile, when KGN cells were implanted at high density, UCMSCs-CM could induce cell cycle arrest at G1 phase, inhibit cell migration, invasion and promote apoptosis. While it had no similar effect on KGN cells implanted at low density. In mechanism, the UCMSCs-CM treatment activated the Hippo pathway when KGN cells were implanted at high density. Consistently, the MST1/2 inhibitor, XMU-MP-1, inhibited the activation of the Hippo pathway induced by UCMSCs-CM treatment and accordingly declined the anti-tumor effect of UCMSCs-CM on KGN cells. Conclusions: The effect of UCMSCs-CM on tumor cells is affected by cell density. UCMSCs-CM exerted anti-tumor effect on KGN cells by activating Hippo pathway to restore contact inhibition. Our results suggest that UCMSCs-CM is a promising therapeutic candidate for GCT treatment.