Fig. 1: The TM displays a rapid and robust proliferative response to injury macroscopically.

a Schematic image of a murine TM dissected en bloc to include the pars flaccida (PF) and pars tensa (PT) (left panel). Unwounded (UW) and Wounded (WO) tympanic membranes depicted. Perforations were made in the anterior PT (right panel). b Representative wild-type murine TMs harvested from different mice at the indicated timepoints post-perforation. Red arrowheads indicate the site of perforation and gross resolution over time. c To characterize the proliferative response to injury, perforations were created in the left TMs of mice on day 0. On days 1, 2, 3, 7, and 14, EdU was injected IP 2 h prior to TM harvest. d Edu-labeled whole-mount TMs demonstrate a peak proliferative response 3 days post-injury, with resolution by day 14. White dashed circles indicate the perforation. e Graph of the number of EdU+ cells in a 400 × 1200 μm area over the malleus in response to injury. Results of t-tests for WO vs. UW TMs at a single time-point are indicated with the black bars with the outer limits representing standard deviation; *p < 0.05. f A cross-sectional view of the TM demonstrates the 3 major layers: inner mucosa, middle mesenchyme, and outer epidermis. When wounded, the injury crosses all 3 layers. g The mesenchymal layer contains two layers of inner circular and outer radial collagen fibers. h Hematoxylin and Eosin (H&E) stained sections of the TM at the level of the perforation mid pars tensa at multiple timepoints following perforation. Tissue stratification peaks at Day 7 and resolves by Day 21. The epidermal layer is oriented downward, and the black circle indicates the site of the perforation. Scale bars: (b) 200 μm (d): 100 μm (h) 100 μm.