Differential Expression of Human b-defensins in Epithelium of Oral Cancer

Human β-defensins (hBDs) are small cationic antimicrobial and immunomodulatory peptides expressed in oral epithelium. Objectives: To determine in vivo peptide expression of hBDs in the evolution of oral squamous cell carcinomas. Methods: Immunofluorescence microscopy was performed on formalin-fixed, paraffin-embedded oral tissue biopsy sections of normal, moderate dysplasia, carcinoma in situ, and oral squamous cell carcinoma. Antigen-retrieved and serum-blocked slides were probed with primary antibodies followed by staining with secondary antibodies conjugated with Alexa Fluor488 or Cy3. The localization and expression of hBDs were detected with a Zeiss Fluorescence Microscope. Cells cultured in DMEM-FBS media were treated with TNFα and LPS for 16 hr, followed by RT-PCR analysis for hBD transcripts. Results: In normal oral epithelium and moderate dysplasia, hBD-3 peptide is expressed primarily in the basal layer, whereas hBD-1 and -2 are expressed largely in the lucidum and granulosum layers. In carcinoma in situ, only hBD-3 is expressed, similar to the basal cell layer of normal epithelium. In terminally-differentiated oral squamous cell carcinoma, the expression of hBD-1 and -2 is limited to cells surrounding the “keratin pearl”, whereas hBD-3 expression is located mainly in the basal layers. β-catenin, a cellular protein whose activation is linked to tumorigenesis, is translocated into the nucleus of cancer cells in carcinoma in situ. TNFα and LPS stimulation induce the expression of hBD-2, but not hBD-3, in oral cancer cell lines SasL1, Sas2, SasH1, and Hsc3. Conclusion: HBD-3 expression is predominantly expressed in proliferating basal layer cells of oral epithelium and in undifferentiated oral cancer cells, suggesting that the peptide is involved in the initiation and progression of oral cancer. Oral cancer cell lines fail to express hBD-3 under various conditions, indicating that these cancer cells have undergone differentiation. Supported by American Heart Association Scientist Development Grant 0535088N/Case Western Reserve University School of Dental Medicine.