HPV E7 Regulates The Tumor Microenvironment Through SIRT1 And Contributes To The Proliferation Of Cervical Cancer Cells

Objective: It is well known that human papilloma virus(HPV)infection is closely related to cervical cancer,HPV E7 plays an important role in cell malignant transformation,but the singaling pathway influenced by E7 has not been well known.In recent years,studies have shown that the depend on the NAD+ histone acetylation enzyme SIRT1 which is closely related to tumor microenvironment,is a potential target of tumor therapy.The purpose of this study was to detecte the expressions of E7 and SIRT1 in the cervical tissues and their relationships and the realted protein with tumor microenvironment in human cervical cancer Si Ha cells,to explore the mechanisms of SIRT1 in the development of cervical cancer.Methods: A total of 103 clinical specimens were collected from the department of pathology in the affiliated hospital of Qingdao university between Jan 2014 to Mar 2014,including CIN ? grade,CIN ?-? grade,cervical squamous carcinoma and normal cervical tissue.Immunohistochemical staining was used to detect the SIRT1 and E7 protein expressions in the clinical specimens,the correlation analyses was made by Fisher-c2 method,chemical synthesis of small interfering RNA targeting E7 gene was transfected into cervical cancer Si Ha cells,the expression level of E7 m RNA was detected by RT-PCR,and the expression of E7,SIRT1,Notch-1,Hes-1,VEGF and HIF-1 a protein were detected by Western blot,the cell proliferation was assessed by CCK-8 method.Results: Immunohistochemical staining indicated that the expressions of SIRT1 in normal cervical tissues and less degree of cervical intraepithelial neoplasia(CIN?grade)were negative,while the positive rate was 32% in CIN?-? grade cervical tissues(8/25)and 60% in the cervical cancer tissues(12/20).The expressions of E7 in normal cervical tissues were not found,while the positive rate was 21% in CIN ? grade(8/38),the positive rate was 48% in CIN?-?(12/25)and 60% in cervical cancer tissues(12/20).The Fisher-?2 analysis of the relationships between E7 and SIRT1 showed that P=0.019.RT-PCR results showed that the m RNA expression of si RNA-E7 group was significantly down-regulated(P<0.05)compared to the other control groups.Western blot showed the expressions of E7 and SIRT1 protein in the si RNA-E7 group was significantly down-regulated(P<0.05),while the expression of Notch-1,Hes-1 and HIF-1? were up-regulated,the expression of VEGF was down-regulated(P<0.05).CCK-8proliferation assay demonstrated that the cell inhibition rates in si RNA-E7 group,the negative control group and the transfection regent control group was 38%,9% and 7.4%separately.Conclusion: In cervical cancer tissues,there was a positive correlation between E7 and SIRT1.Si Ha cell proliferation could be inhibited by the si RNA targeting E7 gene,the expression of SIRT1 was down-regulated,the expression of Notch-1?Hes-1and HIF-1?proteins related to tumor microenment was up-regulated,the expression of VEGF protein was down-regulated,indicating that E7 could regulates the tumor microenvironment through SIRT1 and contributes to the development of cervical cancer.