Exploration of the role of macrophage mitochondrial damage-mediated excessive pyroptosis in ITP

Abstract Introduction Targeting macrophages is expected to become an important entry point for the study of the pathogenesis of ITP and the search for therapeutic targets. Pyroptosis, a form of programmed cell death, serves as a crucial innate immune response mechanism. Mitochondria are crucial structures for cellular fate and inflammatory responses, and their damage is closely related to pyroptosis. Whether mitochondrial damage occurs in ITP, and its relationship with macrophage pyroptosis, remain to be investigated. This study will explore the role of macrophages in ITP pathogenesis from the novel perspective of mitochondrial damage and pyroptosis, providing a theoretical basis for developing innovative therapeutic strategies targeting the macrophage “mitochondria-pyroptosis-immune regulation” axis. Methods Single-cell sequencing was performed on PBMCs from 5 patients with ITP (2 newly diagnosed and 3 refractory cases). The pyroptosis score was calculated using the pyroptosis characteristic gene set. The NLRP3/Caspase-1 pathway molecules and pyroptotic products (IL-1β/IL-18/HMGB1) in the peripheral blood and bone marrow samples of the patients were detected. The THP-1 macrophages were induced to undergo pyroptosis using Nigericin + LPS or the plasma of ITP patients, thereby establishing an in vitro model. The mitochondrial division inhibitor Mdivi-1 (targeting Drp1) was used for intervention, and changes in mitochondrial membrane potential, MtROS levels, and pyroptosis pathway molecules were detected. A passive ITP mouse model was established, and both pyroptosis inhibitor disulfiram and Mdivi-1 were used for intervention to evaluate changes in mitochondrial function, pyroptotic products, and platelet levels. Results Single-cell sequencing shows that the pyroptosis score of monocytes in refractory ITP patients is 4 times higher than that in the newly diagnosed group, and the NLRP3 inflammasome and NOD-like receptor pathways are significantly activated. In ITP patients, there are mitochondrial damage, excessive pyroptosis and excessive activation of the NLRP3/Caspase-1-dependent pathway in peripheral blood mononuclear cells and bone marrow macrophages; the levels of IL-1β, IL-18 and HMGB1 are more obvious in recurrent ITP patients. Mdivi-1 can inhibit the mitochondrial damage of macrophages induced by Nigericin + LPS and the plasma of ITP patients. It can increase the mitochondrial membrane potential, reduce the level of MtROS, and decrease the pyroptosis of macrophages and the excessive activation of the NLRP3/Caspase-1 pathway. Both disulfiram and Mdivi-1 can reduce mitochondrial damage in bone marrow and spleen macrophages of ITP mice, decrease the levels of pyroptosis-related products, and increase the platelet levels of ITP mice. Conclusions Mitochondrial damage induces excessive pyroptosis in macrophages through the classical pyroptosis pathway mediated by NLRP3/Caspase-1, thereby contributing to the pathogenesis of ITP. This pathological process holds promise as a potential target for ITP intervention.