title: HBD-2 is downregulated in oral carcinoma cells by DNA hypermethylation, and increased expression of hBD-2 by DNA demethylation and gene transfection inhibits cell proliferation and invasion.
abstract: Human β-defensin-2 (hBD-2) is a type of epithelial antimicrobial peptide. The expression level of hBD-2 mRNA is lower in oral carcinoma cells (OCCs) than in healthy oral epithelium. Yet, it is still unknown how hBD-2 expression is downregulated in OCCs. The present study investigated DNA hypermethylation of hBD-2 in OCCs and the effect of the demethylation and increased expression of hBD-2 on cell proliferation and invasion. Six different types of oral carcinoma cell lines (OSC-19, BSC-OF, SAS, HSC-2, HSC-4 and HSY) and normal oral keratinocytes (NOKs) were used. The expression levels of hBD-2 in all OCCs were significantly lower than that in the NOKs. Treatment with DNA methyltransferase inhibitor, 5-aza-dC, at the concentration of 50 µM significantly induced upregulation of expression of hBD-2 in the OCCs. Using methylation-specific PCR, DNA hypermethylation was observed in all OCCs. These results suggest that DNA hypermethylation is, at least in part, involved in the decreased expression of hBD-2 in OCCs. We examined the effect of 5-aza-dC on the cell proliferation and invasive ability of OCCs. The cell invasion assays showed that the number of OCCs treated with 5-aza-dC on the filters was significantly lower than that of the controls. We examined whether increased expression of hBD-2 generated by gene transfection inhibited the proliferation and invasion of SAS cells. The number of SAS cells exhibiting increased expression of hBD-2 on the filters in the invasion assay were significantly lower on day 7 when compared with the control. hBD-2 may function as a tumor suppressor. Increased expression of hBD-2 induced by demethylation or increased expression generated by gene transfection may be useful therapeutic methods for oral carcinoma.
authors.affiliation: Division of Oral Medicine and Pathology, Department of Human Biology and Pathophysiology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Kamino
authors.affiliation: Division of Pediatric Dentistry, Department of Oral Growth and Development, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Yoshihito
authors.initials: Y
authors.lastname: Kurashige
authors.affiliation: Division of Disease Control and Molecular Epidemiology, Department of Oral Growth and Development, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Osamu
authors.initials: O
authors.lastname: Uehara
authors.affiliation: Division of Oral Medicine and Pathology, Department of Human Biology and Pathophysiology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Sato
authors.affiliation: Division of Oral Medicine and Pathology, Department of Human Biology and Pathophysiology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Michiko
authors.initials: M
authors.lastname: Nishimura
authors.affiliation: Division of Oral Medicine and Pathology, Department of Human Biology and Pathophysiology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Koki
authors.initials: K
authors.lastname: Yoshida
authors.affiliation: Division of Biochemistry, Department of Oral Biology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Toshiya
authors.initials: T
authors.lastname: Arakawa
authors.affiliation: Division of Oral and Maxillofacial Surgery, Department of Human Biology and Pathophysiology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Nagayasu
authors.affiliation: Division of Pediatric Dentistry, Department of Oral Growth and Development, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Saitoh
authors.affiliation: Division of Oral Medicine and Pathology, Department of Human Biology and Pathophysiology, School of Dentistry, Health Sciences University of Hokkaido, Hokkaido 061-0293, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Abiko
mesh_terms: D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D019175:DNA Methylation / Q000187:drug effects*; D000077209:Decitabine; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism; D009055:Mouth / Q000166:cytology; D009062:Mouth Neoplasms / Q000473:pathology*; D023083:beta-Defensins / Q000235:genetics* / Q000378:metabolism*
chemical_list: C106641:DEFB4A protein, human; D023083:beta-Defensins; D000077209:Decitabine; D001374:Azacitidine