p27Kip1 Knockdown Induces Proliferation in the Organ of Corti in Culture after Efficient shRNA Lentiviral Transduction

• Published in: Journal of the Association for Research in Otolaryngology 2013-08, Vol.14 (4), p.495-508
• Main Authors: Maass, Juan C., Berndt, F. Andrés, Cánovas, José, Kukuljan, Manuel
• Format: Article
• Language: English
• Subjects: Animals; Cell Cycle - drug effects; Cell Differentiation - drug effects; Cell Proliferation - drug effects; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p27 - deficiency; Cyclin-Dependent Kinase Inhibitor p27 - genetics; Gene Expression Regulation - drug effects; Gene Expression Regulation - genetics; Gene Knockdown Techniques; Genetic Vectors - genetics; Lentivirus - genetics; Medicine; Medicine & Public Health; Models, Animal; Neurobiology; Neurosciences; Organ of Corti - cytology; Organ of Corti - physiology; Otorhinolaryngology; Rats; Rats, Sprague-Dawley; Regeneration; Research Article; RNA, Small Interfering - pharmacology; Transduction, Genetic - methods
• ISSN: 1525-3961; 1438-7573
• DOI: 10.1007/s10162-013-0383-2

The cells in the organ of Corti do not exhibit spontaneous cell regeneration; hair cells that die after damage are not replaced. Supporting cells can be induced to transdifferentiate into hair cells, but that would deplete their numbers, therefore impairing epithelium physiology. The loss of p27Kip1 function induces proliferation in the organ of Corti, which raises the possibility to integrate it to the strategies to achieve regeneration. Nevertheless, it is not known if the extent of this proliferative potential, as well as its maintenance in postnatal stages, is compatible with providing a basis for eventual therapeutic manipulation. This is due in part to the limited success of approaches to deliver tools to modify gene expression in the auditory epithelium. We tested the hypothesis that the organ of Corti can undergo significant proliferation when efficient manipulation of the expression of regulators of the cell cycle is achieved. Lentiviral vectors were used to transduce all cochlear cell types, with efficiencies around 4 % for hair cells, 43 % in the overall supporting cell population, and 74 % within lesser epithelial ridge (LER) cells. Expression of short hairpin RNA targeting p27Kip1 encoded by the lentiviral vectors led to measurable proliferation in the organ of Corti and increase in LER cells number but not hair cell regeneration. Our results revalidate the use of lentiviral vectors in the study and in the potential therapeutic approaches for inner ear diseases, as well as demonstrate that efficient manipulation of p27Kip1 is sufficient to induce significant proliferation in the postnatal cochlea.