Tracking Cell Layer Contribution During Repair of the Tympanic Membrane

Abstract

The tympanic membrane (or ear drum) is found at the interface between the middle ear and the external ear. The membrane is composed of three layers of different embryonic origin: an outer ectodermally-derived layer, a middle neural crest-derived fibroblast layer with contribution from the mesoderm-derived vasculature, and an inner endodermally-derived mucosal layer. These layers form a thin sandwich which is often perforated as a consequence of trauma, pressure changes, or middle ear inflammation. Usually, the tympanic membrane heals with minimal scarring, but in 6% of cases the perforation fails to heal leading to hearing loss, tinnitus and pain requiring surgery. How cells bridge the gap to close the perforation is an interesting question, as this needs to happen in the absence of an initial scaffold. Here we assess the contribution, timing, and interaction of the different layers of the membrane during repair in the mouse using markers and reporter mouse lines. We show that the ectodermal layer retracts after perforation, before proliferating away from the wound edge, with Keratin 5 basal cells migrating over the hole to bridge the gap. The mesenchymal and mucosal layers then use this scaffold to complete the repair, in tandem with changes in the vasculature. Finally, differentiation of the epithelium leads to formation of a scab that falls off. Our results reveal the dynamics and interconnections between the embryonic germ layers during repair and highlight how defects in healing may occur. Unearthing the complexities of TM healing is important as chronic TMP is a common clinical issue with limited treatment options.