Keratinocyte necroptosis promotes the progression of radiation-induced oral mucositis.

IMPORTANCE: Radiation-induced oral mucositis (RIOM) is a prevalent complication arising from radiation therapy for tumors or combined radiotherapy, but the therapeutic options available remain limited. Understanding its underlying mechanisms is crucial for developing effective interventions. OBJECTIVES: To investigate whether keratinocyte necroptosis contributes to RIOM pathogenesis and evaluate the effects of RIPK3/MLKL inhibition. METHODS: A mouse model of RIOM was established with varying irradiation doses. Tongue tissues were analyzed via histological staining, immunohistochemistry, and Western blot. In vitro, keratinocytes were irradiated and treated with RIPK3 or MLKL inhibitors. Subsequently, cell viability, necroptosis, and inflammatory cytokine expression were assessed using CCK-8, LDH release, Western blot, flow cytometry and RT-qPCR. RESULTS: In irradiated mouse tongues, p-RIPK3/RIPK3 and p-MLKL/MLKL ratios were significantly elevated (P < 0.01), accompanied by heightened expression levels of IL-1beta and IL-6. Similar findings were observed in keratinocytes, which, after 12 Gy irradiation for 2.5 days, reduced cell viability (P < 0.001), enhanced necroptotic marker expression (P < 0.001), and increased inflammatory cytokine levels (P < 0.001). Furthermore, treatment with RIPK3 inhibitor GSK'872 or MLKL inhibitor GW806742X significantly reduced irradiation-induced keratinocyte cell death (P < 0.001), LDH release (P < 0.001) and the expression of inflammatory cytokines (P < 0.01). CONCLUSIONS: This study provides evidence that RIPK3/MLKL-mediated necroptosis in keratinocytes contributes to the pathogenesis of RIOM. Inhibiting this pathway reduces cell death and inflammation, suggesting a promising therapeutic target for the treatment of RIOM.