Study Of Serum Bradykinin Levels As A Diagnostic Marker In Cervical Cancer And Its Potential Mechanisms To Promote VEGF Expression Via BDKRB2

Purpose: To explore the role of bradykinin(BK)as a serum marker in the diagnosis of cervical cancer,and the role of BK in the development of cervical cancer and its underlying mechanism.Methods: 1.Serum samples from 130 patients with cervical cancer(including 65 pairs before and after surgery,65 preoperative patients),35 patients with cervical intraepithelial neoplasia(CIN)(preoperative and postoperative pairing),and 35 patients with uterine leiomyoma.Collect their clinical case information at the same time,as well as SCCA results from laboratory department.ELISA was used to detect BK levels in the above serum samples.The ROC curve was plotted to evaluate the diagnostic efficacy of BK and SCCA,and the combined diagnostic efficacy of both.2.We searched for data on the bdkrb2 gene on the Oncomine database to compare the expression level of bdkrb2 gene between cervical cancer tissue and normal cervical tissue.Paraffin-embedded cervical cancer tissue specimens(44 cases),CIN tissue samples(12 cases),and uterine fibroid cervical epithelial tissue samples(8 cases)were collected.Immunohistochemistry was used to detect the protein expression of BDKRB2 in each tissue.3.Tube formation assays were performed to test the effects of BK or BDKRB2 antagonist HOE140 on tube formation of human umbilical vein endothelial cells(HUVEC).Western blot,q RT-PCR and ELISA assays were used to detect the effects of BK or HOE140 on the expression of vascular endothelial growth factor(VEGF)in human cervical cancer cell lines(Si Ha and He La).4.Construction of cervical cancer cell lines(Si Ha and He La)with overexpression / knockout of BDKRB2.Western blot,q RT-PCR,ELISA assays were used to detect the effects of BDKRB2 on VEGF expression in cervical cancer cell lines.5.The cervical cancer(Si Ha)subcutaneous tumor models were constructed to detect the effects of BDKRB2 expression on cervical cancer growth and tumor angiogenesis.Results: 1.Compared with CIN and uterine leiomyoma,the level of serum BK in cervical cancer patients is significantly increased,while the level of BK is decreased after surgical treatment.Moreover,in the diagnosis of cervical cancer,the combined detection of BK and squamous cell-associated antigen(SCCA)has a higher diagnostic efficiency than the detection of SCCA and BK,respectively.2.BDKRB2 expression was significantly upregulated in the cervical cancer vs.the normal samples in one dataset of Oncomine online database.Immunohistochemical detection show the protein expression of BDKRB2 in cervical cancer and CIN tissues were significantly higher than normal cervical epithelial tissues.3.Tube formation assays,Western blot,q RT-PCR,ELISA detection show that with BK treatment,HUVEC angiogenesis and VEGF expression increased in Si Ha and He La cells.While HUVEC angiogenesis and VEGF expression decreased with administration of BDKRB2 antagonist HOE140.4.In BDKRB2 overexpression / knockout cervical cancer cells,Western blot,q RT-PCR,and ELISA test results show the levels of VEGF in BDKRB2-overexpressed cells were markedly increased,while BDKRB2?knockdown cervical cancer cells exhibit the opposite effect.5.In vivo experiments show tumor angiogenesis increased and tumor growth accelerated in tumor-bearing nude mice with overexpressed BDKRB2;while tumor angiogenesis decreased and tumor growth slowed in tumor-bearing nude mice with under-expressed BDKRB2.Conclusion: BK may be an auxiliary diagnostic marker for cervical cancer,and combined detection of BK and SCCA may improve the diagnostic efficiency of cervical cancer;BK up-regulates VEGF expression through BDKRB2,thus promoting tumor angiogenesis may be a potential mechanism for the development of cervical cancer.