Latent progenitor cells as potential regulators for tympanic membrane regeneration

Abstract

Tympanic membrane (TM) perforation, in particular chronic otitis media, is one of the most common clinical problems in the world and can present with sensorineural healing loss. Here, we explored an approach for TM regeneration where the latent progenitor or stem cells within TM epithelial layers may play an important regulatory role. We showed that potential TM stem cells present highly positive staining for epithelial stem cell markers in all areas of normal TM tissue. Additionally, they are present at high levels in perforated TMs, especially in proximity to the holes, regardless of acute or chronic status, suggesting that TM stem cells may be a potential factor for TM regeneration. Our study suggests that latent TM stem cells could be potential regulators of regeneration, which provides a new insight into this clinically important process and a potential target for new therapies for chronic otitis media and other eardrum injuries.

Figure 1. Isolation of stem cells from TMs.

a) Optical microscopy (Magnification x4, x10). (b) Immunocytochemistry of epithelial stem cell markers, INGβ1, CK19, p63, and proliferation marker, Ki67. (c) Proportion of TM stem cells expressing INGβ1, CK19, p63 and Ki67.

Figure 2. Immunostaining of normal rat TM sections with INGβ1 and CK19.

Malleus handle (3), annulus (1, 5), pars tensa (2, 4).

Figure 3. Expression of INGβ1 and CK19 in acute TM perforations.

(a) TM at the first day after perforation, (b) TM at the fifth day after perforation. (c) TM at the tenth day after perforation. 1,5; annulus, 3; malleus handle, 2; pars tensa, 4; perforation site.

Figure 4. Expression of INGβ1 and CK 19 in chronic TM perforations.

a) TMs with perforations showed high expression of INGβ1 and CK 19 on the first day, (b) 1 week, (c) 3 weeks, (d) 6 weeks, (e) 9 weeks after completing the whole 7-day procedure for chronic perforations. The completely healed TMs showed low expression of INGβ1 and CK 19 at 6 weeks and 9 weeks after the chronic perforation procedure. 1, 5; annulus, 3; malleus handle, 4; pars tensa, 2; perforation site.

Figure 5. The intensity of expression on potential stem cell makers (Integrin β1 (red color) and Cytokeratin 19 (blue color) according to the anatomic areas (malleus handle; annulus (perforation site); pars tensa; annulus (unperforation site) in the TMs with acute or chronic perforations.

a) Acute TM perforation model. (b) Chronic TM perforation model.