| Background:Cervical cancer is one of the most common malignant tumors in gynecological oncology,ranking fourth in female tumors,seriously threatening women’s health.Therefore,it is great significance to continuously study the molecular mechanism of cervical cancer,discover the tumor markers of early diagnosis and prognosis,and explore its potential therapeutic targets for the diagnosis and treatment of cervical cancer.It is now known that S100A16 is up-regulated in many tumors,which can promote the invasiveness and metastasis of cancer cells.Studies have shown that the expression of tumor S100A16 is an independent factor affecting the prognosis of certain tumors;the poor prognosis of tumor patients is significantly related to abnormal expression of S100A16.However,whether S100A16 is also involved in the pathogenesis of cervical cancer and its possible molecular mechanism has not been reported yet,so it is of great clinical significance to study S100A16 as a new diagnostic and prognostic indicator and potential target for cervical cancerObjective:To explore and clarify the role of S100A16 in cervical cancer and its potential biological mechanisms,and to determine whether S100A16 can serve as a new diagnostic and prognostic marker and potential target for cervical cancer.Methods:1.Investigation of expression and biological function of S100A16 in cervical cancer,normal cervical tissue and adjacent nontumor cervical tissues(1)Analysis of S100A16 mRNA expression in normal cervical tissues and cervical cancer tissues by bioinformatics and investigation of its biological function in cervical cancer.(2)Collection of clinical pathological specimens and detection of S100A16 mRNA and protein expression in cervical cancer and adjacent nontumor cervical tissues by RT-qPCR,tissue microarray technology,and tissue immunohistochemistry,as well as correlation with clinical pathological indicators and prognosis.2.Study on the Role of S100A16 in Promoting Cervical Cancer Cell Proliferation,Metastasis,EMT and Apoptosis(1)Establishment of S100A16 low expression cervical cancer cell lines;detection of S100A16 gene expression in each group of cells by RT-PCR;counting of each cell proliferation with CCK-8;Transwell invasion experiment to detect the migration ability of each group of cells;cell scratch experiment to detect the in vitro movement and migration of each group of cells;flow cytometry to detect the effect of S100A16 knockdown on apoptosis and cycle of each group of cells,and WB detection of the expression of proliferation and apoptosis related factors of each group of cells.(2)Investigation of the role of S100A16 in the proliferation,metastasis,EMT and apoptosis of cervical cancer in vivo by subcutaneous tumor transplantation in nude mice.3.Study on the Mechanism of S100A16 Promoting the Biological Behavior of Cervical Cancer(1)WB detection of changes in PI3K,Akt,mTOR,p-PI3K,p-Akt and p-mTOR pathway related proteins after S100A16 knockdown.(2)Study the effect of blocking PI3K on PI3K/AKT in cervical cancer cells and transplanted tumors;Through subcutaneous tumor formation test in nude mice.(3)investigate the effect of S100A16 knockdown and overexpression on the proliferation and apoptosis of cervical cancer cells,and at the same time,detect the changes of PI3K,Akt,mTOR,p-PI3K,p-Akt and p-mTOR pathway related proteins.Results:(1)Analysis of TCGA database showed that the expression of S100A16 mRNA was significantly up-regulated in tumor samples,which was statistically significant(P<0.05).Meanwhile,the mutation frequency of patients with high and low expression of S100A16 was different.Univariate and multivariate Cox analysis showed that S100A16 could be an independent prognostic factor for cervical cancer.Gene set difference analysis(GSVA)showed that S100A16 was related to P53 pathway,PI3K/AKT/mTOR,oxidative stress,Wnt β-Catenin,EMT and other signaling pathways;Drug sensitivity data from GDSC database showed that S10016 was related to the sensitivity of Gemcitabine,ABT.263,ABT.888,AP.24534,AS601245 and Axitinib.CIBERSORT algorithm analysis showed that S100A16 was significantly positively correlated with adipocytes and dendritic cells and significantly negatively correlated with B cells and T cells.GO analysis results showed that S100A16 was enriched in chromosome segregation,nuclear division,catalytic activity for DNA,and cell fission pathways.KEGG results showed that the gene was mainly enriched in cell cycle,DNA replication,meiotic reduction division and other pathways.(2)By analyzing clinical specimens and comparing to adjacent tissues of cervical cancer,S100A16 mRNA and protein expression were increased in cervical cancer tissues compared to adjacent tissues,and the difference was statistically significant(p<0.05).(3)By analyzing the clinical pathological characteristics of cervical cancer and S100A16,it was found that high expression of S100A16 was related to pathological grade and muscle infiltration of cervical cancer,and the difference was statistically significant(p<0.05).(4)Successfully constructed S100A16 overexpression/low expression cervical cancer cell lines Hela and CaSki cells.Divided into si-S100A16 group and si-NC group.Compared with si-NC group,the expression level of S100A16 was significantly down-regulated in si-S100A16 group(p<0.05).(5)Compared with si-S100A16 group,the number of cell proliferation and migration in si-NC group were significantly increased,and the apoptosis rate in si-S100A 16 group was significantly increased,and the difference was statistically significant(p<0.05).(6)Compared with the si-NC group,cells in the si-S100A16 group were significantly reduced in G0/G1 phase,but significantly increased in G2/M phase and S phase(p<0.01).(7)Compared with the si-NC group,the expression levels of LC3II/I and Beclinl were significantly increased(p<0.01),while the expression level of p62 was significantly decreased(p<0.05)in Hela and CaSki cells of si-S100A16 group.(8)In the subcutaneous transplantation tumor experiment,compared with shRNA-NC,the tumors of shRNA-S 100A16 group were obviously alleviated(P<0.05);compared with OV-NC group,the tumor weight of OV-S100A16 group was significantly increased,and the difference was statistically significant(p<0.05).Immunohistochemical method was used to detect the expression of Ki-67 in subcutaneous transplantation tumor tissues.The expression of Ki-67 in shRNA-S 100A16 group tumor tissues was significantly lower than that in shRNA-NC group,while the expression of Ki-67 in OV-S100A16 group was significantly higher than that in OV-NC group(p<0.05).(9)In the subcutaneous transplantation tumor experiment,compared with NC-S100A16 group,the expression of MMP-2 and MMP-9 in OV-S100A16 group was significantly higher,and the expression of E-cadherin was down-regulated.(10)Through WB experiment,the effect of knocking down S100A16 on the PI3K/AKT/mTOR signaling pathway in cervical cancer cells was detected,the expression of p-PI3K,p-Akt and p-mTOR in Hela and CaSki cells of si-S100A16 group was significantly decreased(p<0.05),After blocking PI3K,the proliferation and migration of cervical cancer cells were inhibited(p<0.05).(11)In nude mouse transplantation tumor,compared with shRNA-NC group,the expression of p-PI3K,p-Akt,p-mTOR and mTOR in shRNA-S 100A16 group was significantly decreased(p<0.05),and the expression of PI3K was significantly increased(p<0.05).Compared with ov-NC group,the expression of p-PI3K,p-Akt and p-mTOR in ov-S100A16 group was significantly increased(p<0.05),and the expression of mTOR was significantly decreased(p<0.05).Conclusion:(1)In cervical cancer tissue,S100A16 is an independent risk factor that affects the poor prognosis of cervical cancer patients,indicating that S100A16 plays an important role in the pathogenesis of cervical cancer and may be a clinical target for diagnosis and treatment.(2)S100A16 can affect the growth cycle of cervical cancer cells,inhibit the apoptosis rate of cervical cancer cells,promote the proliferation and migration of cervical cancer cells,suppress the autophagy ability of cervical cancer cells,and promote the epithelial-mesenchymal transition of cervical cancer cells.(3)S100A16 may promote the proliferation,migration,and EMT of cervical cancer cells by activating the PI3K/AKT/mTOR signaling pathway and is associated with poor prognosis of cervical cancer,suggesting that it may be a potential molecular marker for targeting cervical cancer treatment and prognosis assessment. |
PDF Full Text Request