Abstract
OBJECTIVE: To investigate whether cisplatin induces tumor necrosis factor-α (TNF-α) secretion in human head and neck squamous cell carcinoma (HNSCC) cells to trigger RIP1/RIP3/MLKL-dependent necroptosis of the cells. METHODS: HNSCC cell lines HN4 and SCC4 treated with cisplatin (CDDP) or the combined treatment with CDDP and z-VAD-fmk (a caspase inhibitor) or Nec-1 (a necroptosis inhibitor) for 24 h were examined for changes in cell viability using CCK8 assay and expressions of caspase-8 and necroptosis pathway proteins (RIP1/RIP3/MLKL) using Western blotting. The changes in migration of the cells were assessed with cell scratch assay, and the expressions of epithelial-mesenchymal transition (EMT) marker proteins N-cadherin, vimentin, and E-cadherin as well as the expressions of NF-κB (p65) and TNF-α were detected with Western blotting. RESULTS: The IC(50) of cisplatin was 10 μg/mL in HN4 cells and 15 μg/mL in SCC4 cells. Cisplatin treatment significantly decreased the expressions of caspase-8, N-cadherin and vimentin and increased the expressions of Ecadherin, the necroptosis pathway proteins (RIP1/RIP3/MLKL), TNF-α, and NF-κB (p65), and these changes were obviously inhibited by treatment with Nec-1. Cisplatin stimulation also significantly lowered migration of the cells, and this inhibitory effect was strongly attenuated by Nec-1 treatment. CONCLUSION: Cisplatin activates nuclear factor-κB signaling in HNSCCs to promote TNF-α autocrine and induce RIP1/RIP3/MLKL-dependent necroptosis, thus leading to inhibition of cell proliferation.