Role of Sonic Hedgehog Pathway in the Proliferation and Stemness of Murine Skin-derived Precursor Cells

Objectives: Skin-derived precursor cells (SKPs) are able to be differentiated to mesenchymal lineage cells such as adipogenic, osetogenic and chondrogenic cells in addition to neuronal cells including catecholaminergic neuron. Hedgehog (Hh)-Gli signaling is involved in brain development, self-renewal of neural stem cells and proliferation of various precursor cells. Here we presented data demonstrating the effects of sHh signaling by sHh agonist (Purmorphamine; Pur) in murine SKPs.
Methods: The cells from embryonic murine back skin (E17.5) were propagated in DMEM-F12 (3:1) containing 2% B27 supplement, 20 ng/ml EGF and 40 ng/ml bFGF. At passage 1 or 2, sHh agonist (Pur) or antagonist (Cylclopamine; CP) was treated for 72 h in SKP medium. Gene expression level of each treatment group was analyzed by quantitative PCR. Proliferation of SKPs was analyzed by Standardization of the MTS assay.
Results: SKPs formed spheres and sHh signaling activation promoted this event. The expression level of stem cell markers (Oct4, Nanog, Sox2, Klf4, c-Myc, Rex1 and Sall4), neural progenitor markers (Bmi1, Gli1, c-Kit and Nestin) and cell-lineage specific markers (Patched1, p63 and Sox10) were changed by the treatment of Pur, a sHh agonist. In particular, the expression level of c-Myc and Klf4 (pluripotency marker), and Bmi1, Gli1 and Nestin (neural progenitor marker) was significantly increased by Pur. Inhibition of sHh signaling by CP decreased cell proliferation and the expression level of Bmi1 and Gli1.
Conclusions: Our finding shows that sHh activation by Pur increases cell proliferation and SKP sphere formation with enhancing stemness of the cells.