The Expression Of ?Np63 In Cervical Cancer And Its Relative Mechanisms Study

BackgroundCervical cancer is the most common gynecologic cancer in women.The incidence and mortality of cervical cancer is second only to breast cancer.Compared with other gynecologic malignancies,cervical cancer develops in a younger population of women.Cervical cancer is the only disease with a definite cause of all cancerous lesions in humans.In recent years with the successful development and clinical application of HPV vaccine,to some extent,promote the prevention and treatment of cervical cancer,but he core problems such as the pathogenesis and the risk of cancer infection are still inconclusive.p63,a member of the p53 gene family,which can be divided into two categories: TA isotypes and ?N isotypes.Whereas the TA isoforms show a p53-like funcations,can induce cell cycle arrest and apoptosis,?N isoforms act toward p53 function and promote the growth of transformed cells.p63 mainly plays an important role in the development,differentiation and morphogenesis of epithelial,and exert an oncogenic effect.Under normal physiological conditions,p63 hyperplasia exists in various epithelial tissues.The study found that p63 has important significance for the development of embryonic ectoderm,epithelial stem cells,epithelial tissue.It has been reported that in the cervical,the p63 protein is mainly expressed in the basal and parietal cells,also expressed in the transitional area of the cervical reserve cells and cervix immature and atrophic squamous epithelium and cervical intraepithelial neoplasia,and ?Np63 as the main expression.However,the role of p63 protein in the development of cervical cancer mechanism is not very clear.So we study the expression of ?Np63 in cervical cancer and its related mechanisms,and provide new ideas for the diagnosis and treatment of cervical cancer.Methods1.Immunohistochemistry technique were used to detect the expression of ?Np63 in 23 normal tissues,20 cervical intraepithelial neoplasi and 59 cervical cancer tissues.2.The expression of ?Np63 in cervical cancer cell lines were detected by RT-PCR and Western blot.3.Small interference RNA were used to knockdown the expression of ?Np63 in cervical cancer cell lines Caski and Siha.The proliferation of cervical cancer ceells were detected by CCK8.The apoptosis was of cervical cancer ceells were detected by Flow cytometry instrument.4.The genes related to ?Np63 were screened by gene chip.The differentially expressed genes were verified by RT-PCR and Western blot.Results1.Immunohistochemistry technique showed that ?Np63 in cervical squamous cell carcinoma was mainly localised in the nucleus.The expression of ?Np63 was different in different tissues(p?0.001).The results showed that the expression of ?Np63 was associated with the histological grade and differentiation(p = 0.005),but not related to age and lymph nodes metastasis.2.RT-PCR and Western blot analysis showed that ?Np63 expression in cervical cancer cells were different.The expression in Caski and Siha were high,and the expression in Hela and C33 A was low.Small interference RNA were used to knockdown the expression of ?Np63 in Caski and Siha.CCK8 showed that down-rugulated of ?Np63 can inhibit proliferation in ?Np63-siRNA transfected cells compared with negative control cells(p?0.05).Flow cytometry instrument showed that down-rugulated of ?Np63 can increase the percentage of cells in ?Np63-siRNA transfected cells compared with negative control cells(p?0.05).3.Gene chip were used to detect the differentially expressed genes related to ?Np63 in cervical cancer cell line Caski.RT-PCR and western blot were used to detect the expression of differentially expressed genes.Conclusions1.Immunohistochemistry technique showed that the expression of ?Np63 was high in cervical cancer tisues,which is mainly located in the nucleus.And our results indicated that high expression of ?Np63 were associated with cervical cancer clinical stage and differentiation,but not related to age and lymph nodes metastasis.2.The expression of ?Np63 in Siha and Caski cells was higher than that in Hela and C33 A cells.The expression of ?Np63 in Caski and Siha cells was inhibited by small interference RNA.The result of experiment showed that ?Np63 can increase cervical cancer cell proliferation,and inhibit cervical cancer cell apoptosis.3.The results of gene chip analysis showed that there were 425 differentially expressed genes.260 genes were up-regulated expression and 165 genes were down-regulated expression.Through analysis,we found that ?Np63 was closely related to EGFR and DKK1.The expression of EGFR and DKK1 was down-regulated when ?Np63 was inhibited by small interference RNA.The results were consistent with the results of the chip analysis.