Small molecule inhibitor of Thioredoxin Interacting Protein (TXNIP) attenuates Nicotine-Induced Podocyte Injury and Glomerular Sclerosis

Background: Previous studies strongly identify nicotine as a predisposing factor for chronic kidney disease (CKD) onset and progression to end-stage renal disease (ESRD), clear mechanistic understanding of nicotine induced renal dysfunction remains less understood. The Thioredoxin (TRX) system, comprising NADPH, TRX reductase and TRX, is central to cellular redox balance and protection against cellular oxidative stress. Upregulation in Thioredoxin interacting protein (TXNIP) expression sequesters reduced TRX and affects the TRX cellular disulfide oxidoreductase system. Our recent study postulated that NADPH oxidase dependent O2•− production (reactive oxygen species (ROS)) mediates nicotine-induced NLRP3 inflammasome activation and leads to podocyte injury. However, it remains unknown how NADPH oxidase-derived ROS is sensed by NLRP3 inflammasomes in podocytes upon nicotine stimulation. Hence, the current study assessed whether TXNIP mediates nicotine-induced NLRP3 inflammasome activation and consequent podocyte injury. Methods: TXNIP/NLRP3 molecular interaction was assessed via co-immunoprecipitation; Inflammasome formation and immunofluorescence expressions were analyzed via confocal microscopy; IL-1β production was measured using ELISA; Caspase-1 enzymatic activity was measured by colorimetric assay; Apoptotic induction was assessed by fluorescence-based TUNEL assay. Results: Co-immunoprecipitation based molecular interaction studies showed that TXNIP strongly interacts with NLRP3 upon nicotine stimulation as compared to control podocytes. However, pre-treatment with TXNIP-specific inhibitor SRI-37330 attenuates TXNIP/NLRP3 interaction. Confocal microscopic analysis suggests that nicotine significantly augments the colocalization of NLRP3 with apoptosis associated speck-like protein containing a caspase recruitment domain (ASC), NLRP3 with caspase-1 and NLRP3 with TXNIP in podocytes compared to control cells. However, pre-treatment with TXNIP inhibiting SRI-37330 abolished the nicotine-induced NLRP3/ASC, NLRP3/Caspase-1 and NLRP3/TXNIP co-localization. Nicotine treatment significantly increases active caspase-1 expression (6-fold), caspase-1 enzymatic activity and IL-1β production compared to control cells. However, prior treatment with TXNIP inhibiting SRI-37330 attenuated the nicotine-induced active caspase-1 expression, caspase-1 activity and IL-1β production. Further immunofluorescence studies confirm that nicotine significantly decreases podocin and nephrin (2.5-fold) expression (markers of podocyte damage) compared to control cells. However, pretreatment with TXNIP inhibiting SRI-37330 attenuated the nicotine-induced podocin and nephrin downregulation. Moreover, confocal and biochemical analysis showed that nicotine treatment significantly increased desmin expression, podocyte permeability and apoptosis relative to control cells. However, pre-treatment with TXNIP inhibiting SRI-37330 significantly attenuates nicotine-induced desmin expression, cell permeability and apoptotic induction. Conclusion: Cumulatively, our studies suggest that NLRP3/TXNIP interaction is an important signaling mediator of nicotine-induced NLRP3 inflammasome activation and subsequent podocyte injury. This project was supported by NIH grant, R01HL148711. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.