Conditional immortalization of hair cells from the inner ear

The aim of this work was to culture conditionally immortalized cells that possess the potential to differentiate into mechanosensory hair cells. Utricular epithelia at embryonic stage E16 were cultured from the vestibular system of the H2kbtsA58 transgenic mouse (Immortomouse) that carries a conditionally expressed immortalizing gene derived from the simian virus 40. Immunolabelling showed that the immortalizing transgene product, the T antigen (Tag), was expressed in utricular cells under permissive conditions and that it was inactivated under non-permissive conditions. Several morphologically distinct cell types proliferated when Tag was expressed, including those that resembled fibroblasts, nerve cells and epithelial cells. Mixed cultures of cells from the utricle, passaged up to 50 times every 3-4 days over a period of 5 months, were subsequently allowed to differentiate for 10 days by transferring them to non-permissive conditions. Monoclonal antibody markers were used to locate expression of hair cell specific antigens. One antibody that normally labels stereociliary bundles from postnatal stage P4-6 labelled cellular projections from a population of spheroid cells that were distributed across the culture surface. A second antibody that normally labels stereociliary bundles did not label the same structures. We conclude that utricular hair cell progenitors can be derived from the H2kbtsA58 transgenic mouse but that under the experimental conditions used they do not follow the normal pattern of differentiation.