OBJECTIVETo investigate the role of HPV16E6 and E7 during the transformation of oral epithelial cells.

METHODSAn human immortalized oral epithelial cell line (HIOEC) was established by transfecting HPV16E6, E7 open reading frames using recombinant retroviral system plxsn to human normal oral epithelial cells. Expression of HPV16E6, E7, Rb, P16 and Cycin D1 were analyzed by Western blot in HIOEC and human normal oral epithelial cells. Formation of complex of HPV16E7 and Rb were analyzed by Immunoprecipitation-western blot. Human normal oral epithelial cells and the oral epithelial cells transfected with plxsn were used as control groups.

RESULTSHIOEC expressed HPV16 E6 and E7; HIOEC expressed both hyperphosphorylated and underphosphorylated Rb while oral epithelial cells in two control groups only expressed hyperphosphorylated Rb. HPV16 E7 formed complex with underphosphorylated Rb; the level of P16 and Cyclin D1 had no remarkable change.

CONCLUSIONSHPV16E7 plays an important role in the immortalization of oral epithelial cells induced by HPV16.

Blotting, Western ; Cell Line ; Cell Transformation, Neoplastic ; Cyclin D1 ; analysis ; Cyclin-Dependent Kinase Inhibitor p16 ; analysis ; Humans ; Mouth Mucosa ; metabolism ; pathology ; virology ; Oncogene Proteins, Viral ; physiology ; Papillomavirus E7 Proteins ; Phosphorylation ; Repressor Proteins ; Retinoblastoma Protein ; analysis

OBJECTIVETo establish an immortalized oral epithelial cell line.

METHODSNormal human oral epithelial cells were transfected with HPV16E6/E7 open reading frames using recombinant retroviral system pLXSN. Expression of HPV16E6 and E7 protein were tested by Western blot in three kinds of cells. To define cellular biological characterization of HPV16E6/E7 transfected cells, a series analysis were performed, including protraction of growth curve, HE staining, immunocytochemical staining and scanning electron microscope observation. The tumorigenicity was assessed by colony formation and transplanting the cells into nude mice.

RESULTSHuman oral epithelial cells transfected with HPVE6/E7 has been in culture for over 18 months. The cell line was named HIO615. Western blot analysis showed HIO615 expressed HPV16 E6 and E7 protein. HIOC were positive for cytokeratin, tonofibril and desmosome as observed by scanning electron microscope. The number of large colonies of dense multilayer cells was low (0.77%). No tumor developed in nude mice injected subcutaneously with HIOEC.

CONCLUSIONA human immortalized oral epithelial cell line induced by HPV16E6 and E7 has been successfully established.

Animals ; Cell Line, Transformed ; Cell Transformation, Viral ; Humans ; Mice ; Mice, Inbred BALB C ; Mouth Mucosa ; cytology ; ultrastructure ; virology ; Oncogene Proteins, Viral ; genetics ; Papillomavirus E7 Proteins ; Repressor Proteins