title: Trophic effects of keratinocytes on the axonal development of sensory neurons in a coculture model.
abstract: The epidermis, the outermost structure of the skin, fulfils important roles as a physical barrier between the organism and its environment and as a neuroendocrine, immune and sensory organ. It is innervated by unmyelinated sensory fibres conveying nociceptive and thermoceptive information. Little is known concerning the functional interactions between these sensory fibres and the keratinocytes, which constitute 95% of the epidermal cells. We have developed a coculture model of primary rat sensory neurons and keratinocytes, as well as of equivalent cell-lines: ND7-23 neurons and A431 keratinocytes. We show that primary dorsal root ganglion neurons survive well in a standard keratinocyte reference medium containing a low concentration of calcium, but fail to extend axons. However, when neurons are cocultured with keratinocytes, axonal outgrowth is strongly stimulated. The use of a Transwell culture system indicated that the stimulation of axonal growth depends on a soluble factor secreted by keratinocytes. Axon outgrowth was also induced by nerve growth factor or brain-derived neurotrophic factor, but not by neurotrophin 3 or glial cell-derived neurotrophic factor. Neurons cocultured with keratinocytes did not change their responses to ATP, capsaicin or high potassium solution, as measured by calcium imaging. The trophic effect of keratinocytes concerned essentially a population of medium-sized (17-25 microm) neurons, some of which expressed substance P-like immunoreactivity and responded to capsaicin. Our preparation, in which cells are maintained at low external calcium concentration, could represent a useful in vitro model for characterizing the effect of skin-derived guidance and trophic factors.
authors.affiliation: Institut des Neurosciences Cellulaires et Intégratives (INCI), UMR7168 Centre National de la Recherche Scientifique, Université Louis Pasteur, F-67084 Strasbourg, France.
authors.forename: Lauriane
authors.initials: L
authors.lastname: Ulmann
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Rodeau
authors.forename: Louis
authors.initials: L
authors.lastname: Danoux
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Contet-Audonneau
authors.forename: Gilles
authors.initials: G
authors.lastname: Pauly
authors.forename: Rémy
authors.initials: R
authors.lastname: Schlichter
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001369:Axons / Q000502:physiology* / Q000648:ultrastructure; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology; D002118:Calcium / Q000502:physiology; D002211:Capsaicin / Q000494:pharmacology; D002460:Cell Line; D018920:Coculture Techniques; D003470:Culture Media; D005727:Ganglia, Spinal / Q000166:cytology; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D015603:Keratinocytes / Q000502:physiology* / Q000648:ultrastructure; D009414:Nerve Growth Factors / Q000494:pharmacology; D009475:Neurons, Afferent / Q000502:physiology* / Q000648:ultrastructure; D051381:Rats; D017208:Rats, Wistar; D013373:Substance P / Q000378:metabolism
chemical_list: D019208:Brain-Derived Neurotrophic Factor; D003470:Culture Media; D009414:Nerve Growth Factors; D013373:Substance P; D000255:Adenosine Triphosphate; D002211:Capsaicin; D002118:Calcium