Effects Of Human Papillomavirus Type 16 E5 Gene On The Expression Of EGFR、p21、 P53 And Rb In The Human Oral Epithelial Celland The Human Immortalized Oral Epithelial Cell

Objective Human papillomavirus type 16(HPV16)is one of the most pathogenic of HPV virus,and this virus can be detected in the major cervical cancer epithelial cells.recently,numerous epidemiological reports have shown that HPV16 is also detected in a portion of oral squamous cell carcinoma(OSCC),considered to be highly correlated with the occurrence of the disease.HPV16 mainly encodes three oncogenes: E5,E6 and E7.At present,The carcinogenic function of E6 and E7 protein has been studied very deeply,carcinogenic mechanism for everyone’s consensus.However,the oncogenic function of E5 gene is not clear.The E5 gene is often deleted during the whole genome of the virus is integrated into the host DNA,so E5 is thought to play a role in the early stage of the tumor.E5 also has a very weak transformation in the host cell.In this study,we are trying to investigate the effects of HPV16 E5 on the expression of EGFR、p21、p53 and Rb in the human oral epithelial cell(HOEC)and the human immortalized oral epithelial cell(HIOEC),and to discuss the potential mechanism of HPV16 E5 in the occurrence and development of oral squamous cell carcinoma.Methods(1)Open reading frame of HPV16 E5 was cloned into the lentiviral vector pLOV-N to prepare recombinant plasmid pLOV-E5,which was stably transfected into HOEC and HIOEC,respectively.(2)EGFR、p21、p53 and Rb were detected by real-time quantitative PCR(RT-qPCR)and Western blot on the mRNA level and the protein level,respectively.(3)Cell proliferation after stable trasfection was evaluated by MTT assay n groups of HOEC and groups of HIOEC,respectively.Results(1)The lentiviral vector pLOV-E5 and the control vector pLOV-N were successfully transfected into HOEC and HIOEC,respectively,and stable transfected clones were obtained.(2)After stable trasfection of HPV16 E5,the HOEC cell wasn’t transformed into immortalized cell.The protein levels of EGFRin HOEC were markedly overexpressed,while p21 and p53 protein levels were downregulated,as well as the mRNA levels of p21 and p53.However,the mRNA levels of EGFR had no obvious change after trasfection.There were also no significant changes of Rb on the mRNA and protein levels.Compared with the control cells,the proliferation activity of HOEC/pLOV-E5 cells was significantly increased(p<0.05).(3)The protein levels of EGFR 、 p21 and p53 in HPV16 E5-expressing HIOEC cell were obviously overexpressed,as well as the mRNA levels of p21 and p53.However,the mRNA levels of EGFR had no apparent different between groups.There were also no significant changes of Rb on the mRNA and protein levels.The proliferation activity of HIOEC/pLOV-E5 cell was significantly higher than that of the control groupsby MTT assay.(p<0.05).Conclusions(1)HPV16 E5 can promote the proliferation of the HOEC cell and upregulate the expression of EGFR,while down-regulate the expression of p21 and p53.(2)HPV16 E5 can stimulate the expression of EGFR、p21 and p53 as well as the proliferation of HIOEC.