Prognosis Of Squamous Cell Carcinoma Of Bladder And Molecular Mechanism Of HSDL2 Regulating KLF5 On Proliferation Of Bladder Cancer

Objective:The relationship between malignant bladder cancer and clinicopathological features and prognosis was observed by SEER database.In addition,To investigate the influence of HSDL2 gene on the proliferation and invasion of Bladder cancer maligant meningioma cell line,and to discuss whether the KLF5 gene make the the proliferation and invasiveness effects.Methods:1.The clinicopathological features of patients with bladder cancer were downloaded from SEER database.Significant independent prognostic factors were analyzed by univariate and multivariate Cox analysis,and nomogram prediction model was established.2.Immunohistochemistry was used to detect HSDL2 expression in 50 cases of invasive bladder cancer and adjacent tissues,and the relationship between the expression level and clinicopathological features and survival time of patients with invasive bladder cancer was analyzed.3.Patients with high HSDL2 expression tend to have more tumors,deeper local tumor infiltration,and lymph node metastasis.4.The expression of HSDL2 m RNA in normal urothelial cell lines nhucs and bladder cancer cell lines 5637 and T24 were detected by Q-PCR.5.The expression of HSDL2 in bladder cancer cell line T24 was silenced.The expression of HSDL2 m RNA and protein was detected by Q-PCR and Western blot.6.The expression of HSDL2 in bladder cancer cell line T24 was silenced.MTT assay and plate clone test were used to detect the cell proliferation.7.The expression of HSDL2 in bladder cancer cell line T24 was silenced,and the apoptosis was detected by flow cytometry.8.Immunoprecipitation assay was used to detect the correlation between HSDL2 and KLF5.9.The expression of HSDL2 in bladder cancer cell line T24 was silenced.The proliferation ability of bladder cancer cell line T24 was detected by nude mice tumorigenesis test in vivo.10.The in vivo fluorescence imaging of nude mice in each group was analyzed37 days after inoculation.Results:1.A total of 595 cases of bladder squamous cell carcinoma were enrolled by SEER data and randomly divided into training set(n=416)and validation set(n=179).Univariate and multivariate significant variables were included in nomogram prediction model,including AJCC T,AJCC M,age,tumor size and AJCC N.2.Nomograms for predicting overall survival by training set and validation set indicated that c-index was 0.768(95% CI: 0.741-0.795)and 0.717(95% CI:0.671-0.763),respectively.The c-index of TNM staging was significantly lower than nomogram model in training set of 0.580(95% CI: 0.543-0.617)and validation set of0.540(95% CI: 0.484-0.596).The calibration curve and DCA results showed that the prediction model established by nomogram could predict the 1-year and 3-year survival rate well.In addition,compared with AJCC’s TNM staging,the nomogram prediction model has a wide range of threshold probability and net income.3.The expression of HSDL2 m RNA in bladder cancer cell lines 5637 and T24 was higher than that in normal urothelial cell lines nhucs.4.Hst22 and hst24 were successfully transfected into bladder cancer cells.5.The expression of HSDL2 in bladder cancer cell line T24 was silenced.MTT assay and plate cloning test showed that the cell proliferation was decreased.6.Cell apoptosis was detected by flow cytometry.The results showed that silencing the expression of HSDL2 in T24 cells significantly increased the apoptosis rate.7.Immunoprecipitation assay showed that HSDL2 was silenced and KLF5 was down regulated in T24 cells.8.The results of tumor formation in nude mice in vivo showed that silencing the expression of HSDL2 in T24 cell line of bladder cancer reduced the tumorigenicity and weight of tumor.9.In vivo fluorescence imaging analysis of nude mice showed that the fluorescence intensity of HSDL2 silencing group was significantly lower than that of control group.Conclusions:1.Univariate and multivariate COX results suggested that AJCC T,AJCC M,age,tumor size,and AJCC N were independent prognostic factors influencing bladder malignant squamous cell carcinoma;2.The nomogram prediction model established by clinicopathological features can well predict the 1-year and 3-year survival rates of bladder squamous cell carcinoma;3.HSDL2 can affect the proliferation of bladder cancer;4.HSDL2 may play a role by regulating the expression level of KLF5...