Mouse vestibular supporting cells induce directional differentiation of human amniotic fluid stem cells into functional neurons: a pilot study

Abstract

Objective: To investigate the feasibility of inducing human amniotic fluid stem cells into functional neurons by supporting contact co-culture depended on feeder layer from mouse vestibular supporting cells. Methods: Human amniotic fluid stem cells were isolated to culture. The vestibular tissues were obtained from the newborn C57BL/6J mouse by enzymatic digestion and cell culture, the hollow spheres were selected to prepare a monolayer feeder layer. The nGFP-labeled amniotic fluid stem cells were planted on the surface of the feeder layer to form the supporting contact co-culture without adding any exogenous nerve growth factor and neuronal signal inducing factor, and detected the expression of Tuj1 and PSD95, and investigated whether there were ion channels in neurons by FM1-43. Human amniotic fluid stem cells and mouse vestibular supporting cells, which were differentiated separately, and Transwell coculture was used as the control group. Results: The feeder layer expressed the special marker P27(kip1)of the inner ear supporting cell. The nGFP-labeled amniotic fluid stem cells were inoculated on the feeder layer, and (52.0±3.0)% of the nGFP cells expressed Tuj1,which had typical neurons morphological characteristics[protrusion length (110.7±6.2) μm]; the feeder layer cells were differentiated separately, of which (1.1±0.6) % expressed Tuj1 in the control group; the amniotic fluid stem cells were differentiated independently without typical neuron morphological features [protrusion length (16±4.1) μm], of which (92.0±1.0) % expressed Tuj1. The amniotic fluid stem cells and the feeder layer were co-cultured in Transwell: although (92.0±1.0)% of amniotic fluid stem cells had the expression of Tuj1, which had no typical neurons morphological feature[protrusion length (17±4.5) μm], only (1.2±0.9) % of Tuj1 were observed in the feeder layer. Conclusion: By supporting contact co-culture, the feeder layer from the vestibular supporting cells can successfully differentiate human amniotic fluid stem cells into neurons.