Abstract
The Notch signaling pathway plays an important role in otic neurogenesis by regulating the differentiation of inner ear hair cells and supporting cells. Notch-regulated differentiation is required for the regeneration of hair cells in the inner ear. The temporal expression pattern of Notch ligands and receptors during in vitro hair cell-like cell differentiation from human embryonic stem cells (hESCs) was detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Subsequently, pAJ-U6-shRNA-CMV-Puro/GFP recombinant lentiviral vectors encoding short hairpin RNAs were used to silence JAG-1, JAG-2, and DLL-1, according to the temporal expression pattern of Notch ligands. Then, the effect of each ligand on the in vitro differentiation of hair cells was examined by RT-PCR, immunofluorescence, and scanning electron microscopy (SEM). The results showed that the individual deletion of JAG-2 or DLL-1 had no significant effect on the differentiation of hair cell-like cells. However, the simultaneous inhibition of both DLL-1 and JAG-2 increased the number of hair cell-like cells and decreased the number of supporting cells. JAG-2 and DLL-1 may have a synergistic role in in vitro hair cell differentiation.