It is known that histological changes occur in the tympanic membrane (TM) and the middle ear mucosa in COM. Normally, the tympanic membrane has no cellular structure other than mast and Langerhans cells. A previous study found that mast and Langerhans cells in the TM increased in COM . Another study reported an increase in connective tissue and cell proliferation in the TM in a cholesteatoma model . In our study, a few fibroblasts, inflammatory cells and vascular structures forming the stromal connective tissue were detected in the avivation group.
Although many studies have investigated the mechanisms of cholesteatoma formation, its pathogenesis remains unclear . Many molecules and proteins have been investigated [13,14,15,16]. Studies on cholesteatoma tissue have shown that epidermal growth factor receptor (EGFR) disorder plays a role in the development of cholesteatoma. Bujia et al. reported EGFR upregulation in cholesteatoma tissue . Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a protein that binds to the EGFR and participates in cell proliferation. It has been observed that HB-EGF is more expressed in cholesteatoma tissue than in normal skin, and its expression level correlates with bone degradation . Although high keratinocyte growth factor (KGF) levels have been found in cholesteatoma tissue, its receptor levels have been shown to be the same as those in normal skin tissue . A study suggested that heat shock proteins 60 and 70 may play a role in the aetiopathogenesis of cholesteatoma and that they produce their effects through IL-1 and TNF-α . These findings are promising for cholesteatoma treatment options. In a case report, Chao et al.  reported that the patient completely recovered from cholesteatoma after long-term anti-TNF-a antibody (infliximab) treatment. Our study found high periostin and fibronectin levels in cholesteatoma tissue This suggests that periostin and fibronectin may play a role in the mechanism of cholesteatoma formation and that agents targeting periostin and fibronectin may be used in the treatment of cholesteatoma in the future.
In a study comparing cholesteatoma and normal skin, type 4 collagen continuity was observed in the basement membrane but not in cholesteatoma tissue. Moreover, increased immune reactivity of fibronectin was found in the basement membrane and subepithelial connective tissue . Another study found that tenascin and fibronectin expressed along the epidermal–stromal junction were in the form of a band and extended to the deep stroma. Furthermore, tenascin and fibronectin expression was suggested to disturb cell-matrix interactions during the proliferation phase of cholesteatoma tissue .
Periostin expression has been shown to play a role in the pathogenesis of otolaryngologic diseases such as allergic rhinitis, chronic rhinosinusitis with nasal polyps, aspirin-induced asthma, eosinophilic otitis media and IgG4-related diseases [20,21,22,23,24,25]. Our results suggest that COM can be added to these diseases. A study on ankylosing spondylitis patients found an inverse relationship between disease activity and blood periostin levels. Serum periostin levels were considerably low in patients with high disease activity. The authors also suggested that serum periostin levels in patients with nasal polyps can be used to predict the presence, extent and outcome of the disease . In our study, we detected high periostin levels in cholesteatoma tissue samples but found no significant difference between serum periostin, fibronectin and tenascin-C levels. We speculate that this may be because cholesteatoma is a local disease.
A study on cholesteatoma reported that fibronectin was significantly upregulated in cholesteatoma and granulation tissue. It is known that fibronectin mediates in vitro adhesion of fibroblasts and endothelial cells and promotes in vitro keratinocyte migration and proliferation. Therefore, fibronectin was assumed to play an important role in the pathogenesis of cholesteatoma . In our study, stromal staining of fibronectin was markedly more intense in cholesteatoma tissue than in other tissues. This suggests that fibronectin may play a role in the development of cholesteatoma.
Tenascin-C promotes cell migration and adhesion . It also plays a role in regulating cell functions, such as tenascin-C epithelial proliferation and migration . It has been reported that tissue tenascin-C levels can be used as predictors of disease severity in fibrosis, myocarditis and inflammatory bowel disease . In our study, stromal tenascin-C staining was significantly less intense in the skin samples. However, no correlation was found between tenascin-C and cholesteatoma.
Certain limitations of this study should be mentioned. Firstly, the small sample size of our groups. Secondly, we could not include normal middle ear mucosa in the study due to ethical barriers. Finally, samples cannot be evaluated by a quantitative reverse transcription-polymerase chain reaction and gene expression because we did not have the appropriate equipment and equipment to perform gene analysis. As a continuation of this work, we will continue to work on this issue as well.