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NUSAPl and O-GlcNAcylation were reported to be hyper-activated in many kinds of cancers and involved in the advanced progression of cancers. In bladder cancer, O-GlcNAc transferase (OGT) expresses in patients' urine samples, with no expression in healthy individuals, indicating O-GlcNAcylation might involve in the occurrence and development of bladder cancer. Therefore, the present study aims to investigate the effects of O-GlcNAcylation in bladder cancer and if it can regulate NUSAP1 protein. Western blot, immunohistochemistry, and PCR w