BMAL1 downregulation drives PANoptosis and the osteogenic differentiation impairment of PDLSCs by ERK/AP‐1 signaling pathway

BACKGROUND: One strategy to delay bone loss in periodontitis involves maintaining the osteogenic differentiation function of periodontal ligament stem cells (PDLSCs). The core circadian gene BMAL1 influences the fate of mesenchymal stem cells and is essential for regulating pyroptosis, apoptosis, and necroptosis. PANoptosis, a novel form of programmed cell death, simultaneously activates all 3 pathways. This study focuses on the role of BMAL1, the process of PANoptosis, and the osteogenic impairment of PDLSCs. METHODS: A mouse periodontitis model was established to evaluate the expression of BMAL1 and osteogenic factors. We stimulated PDLSCs with lipopolysaccharide (LPS) and used a Western blot to detect PANoptosis-related factors. Osteogenic factors in PDLSCs were assessed using real-time quantitative polymerase chain reaction (RT-qPCR), Western blot, alkaline phosphatase, and alizarin red staining. The expression of ERK pathway proteins was examined by immunofluorescence and Western blot to investigate how BMAL1 regulates PANoptosis under inflammatory conditions. RESULTS: Treatment with LPS leads to the downregulation of BMAL1 expression, which subsequently induces RIPK1-PANoptosome-mediated PANoptosis in PDLSCs, impairing their osteogenic differentiation function. Inhibition of the RIPK1-PANoptosome with Nec-1S improved the expression of osteogenic differentiation-related genes and proteins. Overexpression of BMAL1 using the synthetic ligand SR1078 alleviated these detrimental effects. Inhibition of the ERK pathway with U0126 reduced the expression of its downstream target AP-1, effectively reversing the impact of BMAL1 on PANoptosis. CONCLUSIONS: The downregulation of BMAL1 triggers PANoptosis in PDLSCs, leading to impaired osteogenic function under inflammatory conditions. This study provides new insights into the pathogenesis of periodontitis and suggests novel targets for its prevention and treatment. PLAIN LANGUAGE SUMMARY: Periodontitis is a chronic inflammatory condition of the oral cavity marked by the destruction of periodontal attachment and resorption of alveolar bone. One strategy to delay alveolar bone loss in periodontitis involves maintaining the osteogenic differentiation function of periodontal ligament stem cells (PDLSCs). The circadian rhythm influences the fate of mesenchymal stem cells, with the core circadian gene BMAL1 playing a crucial role in regulating pyroptosis, apoptosis, and necroptosis. PANoptosis is a novel form of programmed cell death, encompassing pyroptosis, apoptosis, and necroptosis, which may play a role in regulating the osteogenic activity of PDLSCs. Our study aims to detect the role of PANoptosis of PDLSCs in periodontitis and elucidate the underlying relationship between BMAL1 and PANoptosis. We found that treatment with lipopolysaccharide leads to the downregulation of BMAL1 expression, which subsequently induces RIPK1-PANoptosome-mediated PANoptosis in PDLSCs, impairing their osteogenic differentiation function. Notably, inhibition of the RIPK1-PANoptosome improved the expression of osteogenic differentiation-related genes and proteins. Mechanistic exploration revealed that BMAL1 downregulation induces PANoptosis in PDLSCs through the ERK/AP-1 signaling pathway. This study highlights the potential therapeutic targets for mitigating bone loss in periodontitis.