Deafness models have been made by a variety of approaches but problems with models in the mouse have hampered their use for studies of auditory physiology. Many methods rely on drugs that cause damage to cells in organs other than the ear and therefore lead to unacceptably high rates of mortality. The available methods also result in damage to multiple structures in the cochlea. Our objective is to develop a genetic model in which we can induce the death of specific cell types in the cochlea including hair cells and spiral ganglion neurons, two critical cell types for hearing loss. Such a model would expedite studies on cell function and development as well as inner ear regeneration. A model that was highly reproducible and that resulted in apoptotic cell death without damage to tissues that surround the dead cells would expedite procedures for cell replacement by cell transplantation or by use of genetic manipulation or therapeutic means of stimulating endogenous cells to replace the lost cell types. Our objective in Aim 1 is to make mice that contain a gene for cell targeting placed behind a CMV promoter for strong expression of the transgene. The construct contains a STOP sequence that prevents expression of a death-inducing caspase gene cloned between lox sites to permit its removal by a recombinase. In this construct the human caspase-3 gene is fused with FK506 binding sites, adding a further layer of control. Activity of the transgene will be tested in vitro and in vivo in Aim 1a. Our objective is to have lines of mice that provide partial or complete ablation, which is to be achieved by a transgenic approach in Aim 1b and a knock-in approach in Aim 1c. Our objective in Aim 2 is to test two lines of mice for targeted cell ablation in the cochlea. In the first line of mice we will target inner ear hair cells by crossing the transgenic to a Cre mouse specific for hair cells, while in the second line of mice we will cross the caspase transgenic with a Cre mouse that will allow targeting of spiral ganglion neurons. The approach, using a cross to a Cre mouse to target the cell type of choice, and treatment with an FK-506 analogue to induce cell death by dimerization of caspase-3, permits us to control both specification of tissue type and timing of cell death.
In this grant we will develop a mouse model to be used for the study of deafness. This model will be critical for the development of treatments for inner ear disorders in which replacement of hair cells or neurons will be evaluated. Because of the clear correlation between loss of cochlear cells and sensorineural hearing loss, improvements in our ability to regenerate these cells will lead directly to new treatments for loss of hearing.
| Fujioka, Masato; Okano, Hideyuki; Edge, Albert S B (2015) Manipulating cell fate in the cochlea: a feasible therapy for hearing loss. Trends Neurosci 38:139-44 |
| Bramhall, Naomi F; Shi, Fuxin; Arnold, Katrin et al. (2014) Lgr5-positive supporting cells generate new hair cells in the postnatal cochlea. Stem Cell Reports 2:311-22 |
| Mizutari, Kunio; Fujioka, Masato; Hosoya, Makoto et al. (2013) Notch inhibition induces cochlear hair cell regeneration and recovery of hearing after acoustic trauma. Neuron 77:58-69 |
| Fujioka, Masato; Tokano, Hisashi; Fujioka, Keiko Shiina et al. (2011) Generating mouse models of degenerative diseases using Cre/lox-mediated in vivo mosaic cell ablation. J Clin Invest 121:2462-9 |
