Immunohistochemical Study of Keratocystic Odontogenic Tumors and Orthokeratinized Odontogenic Cysts

Objectives: To clarify the differences between keratocystic odontogenic tumors (KCOT) and orthokeratinized odontogenic cysts (OOC), we investigated the localization of CK10, CK13, Bcl-2, and Bax proteins in these lesions by immunohistochemical techniques. Method: We used formalin-fixed, paraffin-embedded sections of 10 KCOT specimens and 10 OOC specimens that we obtained from Osaka Dental University Hospital, Osaka, Japan. Deparaffinized sections were autoclaved at 121 °C for 15 min in 10 mM citrate buffer solution (pH 6.0), for retrieval of the CK10, CK13, and Bax antigens, or alternatively, in 10 mM citrate buffer at pH 9.0, for the retrieval of the Bcl-2 antigen. Endogenous peroxidase activity was blocked by immersing the sections in 3% H₂O₂ solution. The sections were incubated with mouse monoclonal antibodies to CK10, CK13 or Bcl-2, or polyclonal antibody to Bax, each for 1 h at room temperature. This step was followed by incubation of the sections with peroxidase-conjugated dextran polymer reagent for 30 min. The sites of antibody binding were visualized by 3,3'-diaminobenzidine-tetrahydrochroride (DAB) precipitation. Counterstaining was performed with hematoxylin. To evaluate the immunohistochemical staining patterns, we categorized the lining epithelium of KCOT and OOC into three layers: the surface layer (SUL), the spinous layer (SL) and the basal layer (BL). Results: CK10 was localized to the SUL in 3 KCOT cases and to the SUL and SL in 10 OOC cases. By comparison, CK13 was localized to the SUL and SL in 8 KCOT cases, but was not observed in OOC. Bcl-2 protein was localized to the BL in 10 KCOT cases, but was not observed in OOC. Bax was localized to the SUL and SL in 5 KCOT cases and was also localized to the SL and BL in 10 OOC cases. Conclusion: CK10, CK13, Bcl-2, and Bax proteins show differential expression patterns in KCOT and OOC.