A comparison of in vitro treatments for directing stem cells toward a sensory neural fate

Abstract

Purpose Low numbers of primary auditory neurons (ANs) may compromise the clinical performance of a cochlear implant. The focus of this research is to determine whether stem cells can be used to replace the ANs lost following deafness. To successfully replace these neurons, stem cells must be capable of directed differentiation into a sensory neural lineage in vitro and, subsequently, of survival and integration into the deafened cochlea. Materials and Methods In this study, we compared three in vitro treatments for directing the differentiation of mouse embryonic stem cells toward a sensory neural fate using neurotrophins, conditioned media from early post-natal cochlear epithelium, or media containing BMP4. Results In all treatments, stem cells were first exposed to retinoic acid, which was sufficient to induce Brn3a-positive patterning in 8-day differentiated embryoid bodies. After a further 8 days of differentiation in adherent culture conditions, BMP4 media-treated cultures produced higher proportions of cells expressing sensory neural markers in comparison to both the conditioned media and neurotrophin treatments, including significantly greater numbers of cells expressing peripherin ( P ≤ .001), tyrosine receptor kinase B ( P ≤ .001), and β-III tubulin ( P ≤ .001). Conclusions This study illustrated that combined treatment with retinoic acid and BMP4 was most effective at directing differentiation of mouse stem cells into sensory-like neurons in vitro. This finding further supports the role of bone morphogenetic proteins in the differentiation of sensory neurons from neural progenitors, and provides a basis for allotransplantation studies for auditory neuron replacement in the deaf mouse cochlea.