title: MYC gene delivery to adult mouse utricles stimulates proliferation of postmitotic supporting cells in vitro.
abstract: The inner ears of adult humans and other mammals possess a limited capacity for regenerating sensory hair cells, which can lead to permanent auditory and vestibular deficits. During development and regeneration, undifferentiated supporting cells within inner ear sensory epithelia can self-renew and give rise to new hair cells; however, these otic progenitors become depleted postnatally. Therefore, reprogramming differentiated supporting cells into otic progenitors is a potential strategy for restoring regenerative potential to the ear. Transient expression of the induced pluripotency transcription factors, Oct3/4, Klf4, Sox2, and c-Myc reprograms fibroblasts into neural progenitors under neural-promoting culture conditions, so as a first step, we explored whether ectopic expression of these factors can reverse supporting cell quiescence in whole organ cultures of adult mouse utricles. Co-infection of utricles with adenoviral vectors separately encoding Oct3/4, Klf4, Sox2, and the degradation-resistant T58A mutant of c-Myc (c-MycT58A) triggered significant levels of supporting cell S-phase entry as assessed by continuous BrdU labeling. Of the four factors, c-MycT58A alone was both necessary and sufficient for the proliferative response. The number of BrdU-labeled cells plateaued between 5-7 days after infection, and then decreased ~60% by 3 weeks, as many cycling cells appeared to enter apoptosis. Switching to differentiation-promoting culture medium at 5 days after ectopic expression of c-MycT58A temporarily attenuated the loss of BrdU-labeled cells and accompanied a very modest but significant expansion of the sensory epithelium. A small number of the proliferating cells in these cultures labeled for the hair cell marker, myosin VIIA, suggesting they had begun differentiating towards a hair cell fate. The results indicate that ectopic expression of c-MycT58A in combination with methods for promoting cell survival and differentiation may restore regenerative potential to supporting cells within the adult mammalian inner ear.
authors.affiliation: Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, United States of America. joburns@wakehealth.edu
authors.forename: Joseph C
authors.initials: JC
authors.lastname: Burns
authors.forename: James J
authors.initials: JJ
authors.lastname: Yoo
authors.forename: Anthony
authors.initials: A
authors.lastname: Atala
authors.forename: John D
authors.initials: JD
authors.lastname: Jackson
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics; D018014:Gene Transfer Techniques; D006198:Hair Cells, Auditory / Q000166:cytology / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D008938:Mitosis / Q000235:genetics; D009154:Mutation; D000081412:Myosin VIIa; D009218:Myosins / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D009924:Organ Culture Techniques; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics* / Q000378:metabolism; D016196:S Phase / Q000235:genetics; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism; D012444:Saccule and Utricle / Q000166:cytology* / Q000378:metabolism*; D013997:Time Factors
chemical_list: C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; C000631455:MYO7A protein, human; C489428:Myc protein, mouse; C000631456:Myo7a protein, mouse; D000081412:Myosin VIIa; D050814:Octamer Transcription Factor-3; C494174:Pou5f1 protein, mouse; D016271:Proto-Oncogene Proteins c-myc; D055748:SOXB1 Transcription Factors; C527783:Sox2 protein, mouse; D009218:Myosins