title: Nod-like receptor protein 3 (NLRP3) inflammasome activation and podocyte injury via thioredoxin-interacting protein (TXNIP) during hyperhomocysteinemia.
abstract: NADPH oxidase-derived reactive oxygen species (ROS) have been reported to activate NLRP3 inflammasomes resulting in podocyte and glomerular injury during hyperhomocysteinemia (hHcys). However, the mechanism by which the inflammasome senses ROS is still unknown in podocytes upon hHcys stimulation. The current study explored whether thioredoxin-interacting protein (TXNIP), an endogenous inhibitor of the antioxidant thioredoxin and ROS sensor, mediates hHcys-induced NLRP3 inflammasome activation and consequent glomerular injury. In cultured podocytes, size exclusion chromatography and confocal microscopy showed that inhibition of TXNIP by siRNA or verapamil prevented Hcys-induced TXNIP protein recruitment to form NLRP3 inflammasomes and abolished Hcys-induced increases in caspase-1 activity and IL-1β production. TXNIP inhibition protected podocytes from injury as shown by normal expression levels of podocyte markers, podocin and desmin. In vivo, adult C57BL/6J male mice were fed a folate-free diet for 4 weeks to induce hHcys, and TXNIP was inhibited by verapamil (1 mg/ml in drinking water) or by local microbubble-ultrasound TXNIP shRNA transfection. Evidenced by immunofluorescence and co-immunoprecipitation studies, glomerular inflammasome formation and TXNIP binding to NLRP3 were markedly increased in mice with hHcys but not in TXNIP shRNA-transfected mice or those receiving verapamil. Furthermore, TXNIP inhibition significantly reduced caspase-1 activity and IL-1β production in glomeruli of mice with hHcys. Correspondingly, TXNIP shRNA transfection and verapamil attenuated hHcys-induced proteinuria, albuminuria, glomerular damage, and podocyte injury. In conclusion, our results demonstrate that TXNIP binding to NLRP3 is a key signaling mechanism necessary for hHcys-induced NLRP3 inflammasome formation and activation and subsequent glomerular injury.
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298. Electronic address: pli@vcu.edu.
authors.forename: Justine M
authors.initials: JM
authors.lastname: Abais
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298.
authors.forename: Min
authors.initials: M
authors.lastname: Xia
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298.
authors.forename: Guangbi
authors.initials: G
authors.lastname: Li
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298.
authors.forename: Yang
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298.
authors.forename: Sabena M
authors.initials: SM
authors.lastname: Conley
authors.affiliation: Internal Medicine Virginia Commonwealth University, School of Medicine, Richmond, Virginia 23298.
authors.forename: Todd W B
authors.initials: TWB
authors.lastname: Gehr
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298.
authors.forename: Krishna M
authors.initials: KM
authors.lastname: Boini
authors.affiliation: Departments of Pharmacology and Toxicology and School of Medicine, Richmond, Virginia 23298.
authors.forename: Pin-Lan
authors.initials: PL
authors.lastname: Li
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020170:Caspase 1 / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D020138:Hyperhomocysteinemia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D058847:Inflammasomes / Q000235:genetics / Q000378:metabolism*; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D050199:Podocytes / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D013879:Thioredoxins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014665:Vasodilator Agents / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
chemical_list: D002352:Carrier Proteins; C587672:IL1B protein, mouse; D058847:Inflammasomes; D053583:Interleukin-1beta; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C492716:Nlrp3 protein, mouse; C445672:Txnip protein, mouse; D014665:Vasodilator Agents; D013879:Thioredoxins; D014700:Verapamil; D020170:Caspase 1
keywords: Glomerulosclerosis; Homocysteine; Inflammation; Kidney; Podocyte; Redox Signaling; Thioredoxin