Abstract #5604: Bmi-1 protects human oral epithelial cells from ionizing radiation

Oral mucositis is a debilitating side effect of anti-cancer therapy that affects almost all patients exposed to therapeutic radiation. Pathogenesis of oral mucositis involves the direct genotoxic insult to the epithelial cells and the inflammatory response triggered by ionizing radiation (IR) or pro-inflammatory cytokines, such as TNF-\#945;. Epithelial tissue damage by IR is attributed to accumulation of DNA damage that leads to premature senescence. We previously showed that Bmi-1 enhances replication of normal human oral keratinocytes (NHOK). In the current study, we tested whether Bmi-1 could rescue NHOK from IR-induced premature senescence. Primary NHOK was infected with retroviral vector expressing Bmi-1or the empty vector (B0) to establish NHOK/Bmi-1 and NHOK/B0 cells, respectively. Three dimensional (3D) oral mucosal cultures were generated from these cells. We compared the phenotypic and molecular alternations of the cells and the reconstructs after exposure to IR or TNF-\#945;. The vast majority of NHOK/B0 cells underwent senescence within 5 days post-IR. These cells stopped proliferation and expressed senescence-associated \#946;-galactosidase (SA \#946;-Gal) activity. However, NHOK/Bmi-1 cells continued to proliferate, maintained the clonogenic properties, and exhibited reduced staining for SA \#946;-Gal even after 10 Gy IR. We then determined the role of Bmi-1 in DNA repair. NHOK/Bmi-1 demonstrated enhanced in vitro DNA end joining activity and in vivo double-strand break repair activity compared to those of NHOK/B0. Since oral mucositis is an inflammatory disorder, we determined the inflammatory signaling in NHOK/B0 and NHOK/Bmi-1. The pro-inflammatory signaling mediated by NF-\#954;B was significantly inhibited by Bmi-1. We determined the phenotypic responses of the 3D cultures exposed to IR or TNF-\#945; by histological examination and immunohistochemistry for proliferating cell nuclear antigen (PCNA), a cell proliferation marker. Upon exposure to either IR or TNF-\#945;, NHOK/B0 cells cultured in 3D models revealed disorganized epithelial structure and lost the expression of PCNA. However, NHOK/Bmi-1 cells retained the organized epithelial stratification and the proliferating basal cell layer. These results indicate that Bmi-1 protects oral epithelial cells from IR-induced mucositis by mitigating the genotoxic stress and pro-inflammatory signaling. This work was supported in part by grants (R01DE18295, K02DE18959, U19AI67769) from NIDCR/NIH. Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 5604.