ROS/NLRP3/Caspase-1 Pathway Contributes to TCP Particles-Induced Pyroptosis of Calvaria Osteocytes in the Mouse Osteolysis Model

Wear particles-induced inflammatory osteolysis, a major contributing factor of aseptic loosening, affects long-term survival of orthopedic prostheses. Increasing observations have demonstrated that osteocytes death is involved in wear particles-induced periprosthetic osteolysis, but the underling molecular mechanisms remain unclear. We chose micro-sized tricalcium phosphate (TCP) particles to simulate TCP wear debris from loosened TCP implants, which have been widely used as bone substitute biomaterials in orthopedic and dental surgery for almost 40 years, and investigated the direct biological effects of the particles on calvaria osteocytes in vivo . Results showed that TCP wear particles triggered nod-like receptor protein 3 (NLRP3) inflammasome activation and pyroptosis in calvaria osteocytes, as indicated by increased osteocytes death, up-regulation of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), cleaved Caspase-1 and interleukin (IL)-1β, elevation of IL-1β, IL-18 and lactic dehydrogenase (LDH), which were all inhibited by the Caspase-1 inhibitor VX765. Moreover, the NLRP3 inhibitor MCC950 reduced TCP particles-induced Caspase-1 cleavage, production of IL-18, IL-1β and LDH, and pyroptotic death in calvaria osteocytes. Further experiments revealed that the NLRP3-mediated pyroptosis pathway was mediated by reactive oxygen species (ROS) in TCP particles-induced cell death of calvaria osteocytes, since a ROS scavenger (N-acetyl-cysteine, NAC) efficiently prevented NLRP3 inflammasome activation and pyroptosis. Taken together, TCP wear particles induce pyroptosis of calvaria osteocytes through activation of ROS/NLRP3/Caspase-1 pathway, which is involved in osteoclastogenesis and periprosthetic osteolysis. Our findings strongly suggest that pyroptosis may play a significant role in wear particles-induced cell death of osteocytes and periprosthetic osteolysis, and will provide a novel evidence for elucidating its pathophysiology.