Establishment and characterization of an in vitro model for cholesteatoma.
OBJECTIVE
Experimental models are of importance to study the pathogenesis of middle ear cholesteatoma, however, they were not established until now. We aimed to develop in vitro model of middle ear cholesteatoma using primary keratinocytes and fibroblasts isolated from cholesteatoma tissue. HaCaT cell line was used as a "skin equivalent" and to compare the grade of homogeneity between cholesteatoma keratinocytes and HaCaT cells.


METHODS
Primary keratinocytes were isolated from cholesteatoma tissue, co-cultured with preliminary prepared feeder layer from cholesteatoma fibroblasts and subsequently air-exposed. The protein profile of cholesteatoma keratinocytes and HaCaT cells was evaluated by means of immunoblot using monoclonal antibody against cytokeratin (CK) 13 and 16. Tissue localization of CK 13 and 16 was accomplished with immunohistochemistry.


RESULTS
Different protein profile and stronger expression of CK 13 and 16 were demonstrated in cholesteatoma keratinocytes in comparison with HaCaT cells. Bigger stratification was observed in the 3D-in vitro systems when both cholesteatoma keratinocytes and HaCaT cells were respectively co-cultured with fibroblasts in comparison with the corresponding control groups without fibroblasts.


CONCLUSIONS
3D-model demonstrates the significance of intercellular interaction between components of cholesteatoma tissue.
Raynov
Alexander M
AM
Department of Otorhinolaryngology, University Hospital Tzaritza Joanna, Medical University-Sofia, Sofia, Bulgaria.
Choung
Yun-Hoon
YH
Park
Hun Yi
HY
Choi
Seong Jun
SJ
Park
Keehyun
K