Role of Transcription Factors in the Motoneuron Differentiation of Adult Human Olfactory Neuroepithelial-Derived Progenitors

¹ University of Louisville, Department of Anatomical Sciences & Neurobiology, Louisville, Kentucky
² University of Louisville, Department of Surgery-Otolaryngology, Louisville, Kentucky

^* To whom correspondence should be addressed. E-mail: fjrois01{at}gwise.louisville.edu.

	Abstract

Neurosphere forming cell (NSFC) lines have been established from cultures of human adult olfactory neuroepithelium. Few of these cells ever express mature neuronal or glial markers in MEM supplemented with 10% FBS or defined medium. However these neural progenitors have the potential to differentiate along glial or neuronal lineages. To evaluate the potential of NSFCs to form motoneurons, transcription factors, Olig2, Ngn2 and HB9, were introduced into NSFCs to determine if their expression is sufficient for motoneuron specification and differentiation as has been shown in the early development of the avian and murine central nervous systems in vivo. NSFCs transfected with Olig2, Ngn2, and HB9 alone exhibited no phenotypic lineage-restriction. In contrast, simultaneous transfection of Ngn2 and HB9 cDNA increased the expression of Isl1/2 a motoneuron marker when the cells were maintained in medium supplemented with retinoic acid, forskolin, and sonic hedgehog (RFS). Furthermore, a population of Olig2-expressing NSFCs also expressed Ngn2. Co-transfection of NSFCs with Olig2 and HB9, but not Olig2 and Ngn2, increased Isl1/2 expression. Co-culture of NSFCs transfected with Ngn2-HB92, or Olig2 and HB9 with purified chicken skeletal muscle demonstrated frequent contacts that resembled neuromuscular junctions. These studies demonstrate that transcription factors governing the early development of chick and mouse motoneuron formation are able to drive human adult olfactory neuroepithelial progenitors to differentiate into motoneurons in vitro. Our long-term goal is to develop cell populations for future studies of the therapeutic utility of these olfactory-derived NSFCs for autologous cell replacement strategies for CNS trauma and neurodegenerative diseases.

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