ELP2-NLRP3-GSDMD/GSDME-mediated pyroptosis is induced by TNF-α in MC3T3-E1 cells during osteogenic differentiation

Tumour necrosis factor-alpha (TNF-alpha) is a cytokine involved in systemic inflammation. TNF-alpha slows down osteogenic differentiation, which may contribute to poor bone development in the inflammatory microenvironment. TNF-alpha inhibits osteogenic differentiation by activating the JAK-STAT3 pathway, of which Signal transducer and activator of transcription 3 (STAT3)-interacting protein 1 (StIP1, also known as elongator complex protein 2, ELP2) is a key protein in the JAK-STAT3 pathway. We investigated whether and how ELP2 activation mediates the TNF-alpha-induced pyroptosis during osteoblastic differentiation. Using in vitro cell cultures of preosteoblastic MC3T3-E1 cells, we found that TNF-alpha exposure causes cell pyroptosis in an inflammatory microenvironment during osteoblastic differentiation. Bioinformatics, protein docking model and co-immunoprecipitation analysis revealed an association between ELP2, STAT3 and NLRP3. Forced ELP2 expression promoted MC3T3-E1 cells pyroptosis, with an increase in the expression of STAT3, NLRP3 inflammasome, GSDMD/GSDME, osteoblast marker genes, and the activity of alkaline phosphatase. In contrast, ELP2 silencing ameliorated MC3T3-E1 cells pyroptosis, and osteogenic differentiation, especially after TNF-alpha stimulation. The TNF-alpha-induced cells pyroptosis during osteoblastic differentiation was therefore mediated by ELP2. These results suggest that ELP2 is upregulated at the pyroptosis of MC3T3-E1 cells and inhibits osteogenic differentiation in response to TNF-alpha through NLRP3-GSDMD/GSDME activation.