Notch1 Expression In Bladder Cancer Cells Mediate The Polarization Of Macrophages In The Tumor Micro-environment

Objective:To investigate the expression and clinical significance of NOTCH1 in bladder cancer tissue;to detect the expression of NOTCH1 and macrophage markers CD68 and CD163 in bladder cancer tissue by immunofluorescence,and to classify the relationship among the three;to detect bladder cancer cells Expression of NOTCH1 in lines.By constructing in vitro co-culture system,T24 cells were co-cultured with THP-1 cells,and proliferation ability of T24 cells and expression of CD68 and CD163 were detected after co-culture.NOTCH1 inhibitor DAPT was added into T24 cells in co-culture system,and the proliferation ability and expression of CD68 and CD163 of T24 cells after administration were detected.To investigate whether NOTCH1 affects the progression of bladder cancer by regulating macrophage polarization.Methods:The paraffin specimens of 62 cases of bladder cancer tissues were selected,and multiplex immunofluorescence staining was used to detect the expressions of NOTCH1 and macrophage markers CD68 and CD163 in bladder cancer tissues.The expression of NOTCH1 was detected by WB and RT-QPCR in 20 fresh bladder cancer specimens and their adjacent adjacent specimens.Human ureteral epithelial immortalized cells SV-HUC1 were selected as control,and T24 and 5637 cells were used as experimental groups.The expression level of NOTCH1 was detected by WB and RT-QPCR.THP-1 cells were induced into macrophages by PMA,and an in vitro co-culture system was constructed.T24 cells were co-cultured with macrophage cells after induction.The NOTCH1 inhibitor DAPT was added to T24 cells,and T24 cells were detected by MTT before and after administration.The proliferation ability was detected by Real-time PCR to detect the expressions of CD68 and CD163.Results:1.Immunofluorescence staining showed that the expression of NOTCH1 was proportional to the expression of M1 macrophages and inversely proportional to M2 macrophages.And the expression of NOTCH1 is related to the prognosis of patients,the higher the expression,the better the prognosis;2.Real-time PCR and Western Blot in fresh bladder cancer specimens and adjacent specimens showed that the expression of NOTCH1 in bladder cancer was down-regulated;3.In T24,5637 cells,the expression of NOTCH1 is lower than that in SV-HUC1cells;4.Co-culture of T24 cells and macrophages promotes the polarization of macrophages to M2 type;5.The supernatant of co-culture of T24 cells and macrophages promotes the proliferation of T24 cells;6.The expression of NOTCH1 was down-regulated after co-culture of T24 cells and macrophages;7.After inhibiting the NOTCH1 expression of T24 cells in the co-culture system,the proliferation ability of T24 cells became stronger,and the polarization of macrophages to M2 type was more obvious.Conclusion:NOTCH1 is down-regulated in bladder cancer tissues and cells,and can inhibit the proliferation of bladder cancer.Co-culture of T24 cells with Mφ cells downregulated the expression of NOTCH1,which inhibited the development of bladder cancer by promoting m1-type polarization of macrophages.