A novel redox signalling axis mediates nicotine-induced podocyte injury.

INTRODUCTION: Nicotine has been shown to induce Nlrp3 (nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3) inflammasome activation, thereby contributing to podocyte injury. However, the molecular mechanisms underlying nicotine-induced Nlrp3 inflammasome activation and subsequent podocyte dysfunction remain largely unknown. The present study investigated whether membrane raft (MR)-associated redox signalling plays a critical role in nicotine-induced Nlrp3 inflammasome activation and podocyte injury. METHODS AND RESULTS: Nicotine treatment induced membrane raft clustering in podocytes in a dose-dependent manner. Upon nicotine stimulation, the NADPH oxidase subunits gp91phox and p47phox were recruited and aggregated within membrane raft clusters, leading to the formation of a membrane raft redox signalling platform. The formation of this signalling platform was significantly inhibited by pretreatment with the membrane raft disruptor methyl-β-cyclodextrin (MCD), the NADPH oxidase inhibitor diphenyleneiodonium (DPI), or the acid sphingomyelinase (Asm) inhibitor amitriptyline. Nicotine also markedly enhanced the colocalization of Nlrp3 with ASC and caspase-1, indicating inflammasome assembly, and significantly increased caspase-1 activity, IL-1β production, desmin expression, podocyte permeability, and apoptosis compared with control cells. These effects were significantly attenuated by pretreatment with MCD, DPI, the caspase-1 inhibitor WEHD, or amitriptyline. Furthermore, immunofluorescence analysis demonstrated that nicotine significantly reduced podocin expression, whereas pretreatment with MCD, DPI, WEHD, or amitriptyline largely preserved podocin expression. CONCLUSION: Our findings demonstrate that membrane raft-associated redox signalling serves as a critical upstream mechanism driving nicotine-induced NLRP3 inflammasome activation and podocyte injury.