The Notch pathway is a cell signaling pathway determining initial specification and subsequent cell fate
in the inner ear. Previous studies have suggested that new hair cells (HCs) can be regenerated in the inner
ear by manipulating the Notch pathway. In the present study, delivery of siRNA to Hes1 and Hes5 using a
transfection reagent or siRNA to Hes1 encapsulated within poly(lactide-co-glycolide acid) (PLGA) nanoparticles
increased HC numbers in non-toxin treated organotypic cultures of cochleae and maculae of
postnatal day 3 mouse pups. An increase in HCs was also observed in cultured cochleae and maculae of
mouse pups pre-conditioned with a HC toxin (4-hydroxy-2-nonenal or neomycin) and then treated with
the various siRNA formulations. Treating cochleae with siRNA to Hes1 associated with a transfection
reagent or siRNA to Hes1 delivered by PLGA nanoparticles decreased Hes1 mRNA and up-regulated Atoh1
mRNA expression allowing supporting cells (SCs) to acquire a HC fate. Experiments using cochleae and
maculae of p27kip1/-GFP transgenic mouse pups demonstrated that newly generated HCs transdifferentiated
from SCs. Furthermore, PLGA nanoparticles are non-toxic to inner ear tissue, readily
taken up by cells within the tissue of interest, and present a synthetic delivery system that is a safe
alternative to viral vectors. These results indicate that when delivered using a suitable vehicle, Hes
siRNAs are potential therapeutic molecules that may have the capacity to regenerate new HCs in the
inner ear and possibly restore human hearing and balance function.