Abstract: Objective To investigate the role and mechanism of histone H3 in regulating the proliferation and apoptosis of bladder cancer cells via nuclear factor-κB (NF-κB). Methods Bladder cancer tissues and adjacent normal tissues were collected after surgical resection. Normal urinary tract epithelial cell line SV-HUC-1 and bladder cancer cell lines BIU-87, J82, RT4, UM-UC-3, and T24 were cultured to detect the levels of nuclear protein NF-κB and total protein Survivin. T24 cells were divided into groups and treated with different concentrations of histone H3 (50, 100, or 200 mg/L), or a combination of 200 mg/L histone H3 with 200 μmol/L NF-κB inhibitor PDTC. The cell proliferation (D₄₅₀ value), apoptotic rate, and the levels of NF-κB in the nuclear protein, Survivin in the total protein, and cleaved caspase-9 and cleaved caspase-3 were detected. Results The levels of NF-κB and Survivin in bladder cancer tissues were higher than those in adjacent normal tissues. In BIU-87, J82, RT4, UM-UC-3, and T24 cells, their NF-κB and Survivin levels were higher than those in SV-HUC-1 cells (P<0.05). T24 cells, which showed the most significantly upregulated expression of NF-κB and Survivin, were selected for subsequent experiments. Histone H3 increased the D₄₅₀ value and the levels of NF-κB and Survivin in T24 cells in a concentration-dependent manner, and inhibited the apoptotic rate and the levels of cleaved caspase-9 and cleaved caspase-3 (P<0.05). After treatment with 200 mg/L histone H3 combined with 200 μmol/L NF-κB inhibitor PDTC, their D₄₅₀ value and the levels of NF-κB and Survivin in T24 cells decreased, compared with those in the 200 mg/L histone H3 group, while their apoptotic rate and the levels of cleaved caspase-9 and cleaved caspase-3 increased, compared with those in the 200 mg/L histone H3 group (P<0.05). Conclusions Extracellular histone H3 promotes cell proliferation and inhibits apoptosis in bladder cancer cells by activating the NF-κB/Survivin pathway.