The Effect of Melatonın on Hıppo Sıgnallıng Pathway in Dental Pulp Stem Cells

Abstract

Dental pulp stem cells (DPSCs) are heterogeneous multipotent adult stem cells with high capacity to differentiate to both neuronal and non-neuronal cell lineage. Hippo pathway regulates diverse cellular processes, including cell survival, proliferation, differentiation, and organ size. Yes-associated protein (YAP) is an important downstream effector of the Hippo signaling pathway involved in neuronal differentiation of neural progenitor cells. Melatonin has a regulatory role for the differentiation of neuronal lineage. Therefore, melatonin may have modulatory role in neurogenic differentiation by interacting with Hippo signaling pathway. In regard with this, DPSCs were incubated with growth and dopaminergic neuronal differentiation medium with or without melatonin (10 µM) for 21 days. The morphological changes were followed by phase contrast microscopy and differentiation of DPSCs was evaluated by immunofluorescence labelling with Neun, GFAP, TH and DMP1. Furthermore, we evaluated the presence of neural progenitor cells by nestin immunoreactivity. Hippo signaling pathway was investigated by evaluating the immunoreactivity of YAP and p-YAPY357. Our results were also supported by western-blot analysis and SOX2, PCNA and caspase-3 were also evaluated. The positive immunoreactivity for Neun, TH and negative immunoreactivity for GFAP showed the successful differentiation of DPSCs to neurons, not glial cells. Melatonin (10µM) addition to dopaminergic media induced TH and decreased significantly nestin expression. The expressions of PCNA and caspase-3 were also decreased significantly with melatonin addition into growth media. Melatonin treatment induced phosphorylation of YAPY357 and reduced YAP expression. In conclusion, melatonin has potential to induce the neuronal differentiation and reduce proliferation of DPSC by increasing phosphorylation of YAPY357 and eliminating the activity of YAP, which indicates the active state of Hippo signaling pathway.