Blocking Intracellular TXNIP Shuttling Attenuates Reactive Oxygen Species-Associated NLRP3 Inflammasome Activation Induced by Tubulointerstitial Fibrosis in Diabetic Kidney Disease

Background: Accumulating evidence underscores the pivotal role of tubulointerstitial fibrosis (TIF) in the progression of diabetic kidney disease (DKD). TXNIP is an important regulatory protein that elicits the generation of reactive oxygen species (ROS) that involved in the fibrotic pathogenesis of DKD. However, the potential mechanism of TXNIP in DKD is not yet well understood. Methods: RNA sequencing and RT-qPCR were used to detect the most significant expression of protein in high glucose-stimulated renal tubular epithelial cells (RTECs). Co-immunoprecipitation, ARE luciferase assay and chromatin immunoprecipitation assay were used to identify the transcriptional relationships between STAT6 and TXNIP. Immunofluorescence and western blot were used to detect the TXNIP shuttling. TXNIP-mediated NLRP3 inflammasome activation were examined via TXNIP siRNA and overexpression in vitro and knockdown in RTECs using AAV-shRNA in vivo. Correspondingly, the efficacy of Astragaloside IV (AS-IV), an active ingredient from traditional formula of Huangqi decoction, was examined in vitro and in vivo. Results: We found that TXNIP was highly expressed in high glucose-stimulated RTECs and the renal tubules of patients with DKD. Experimental data showed that the TXNIP expression was positively associated with the development of TIF in DKD. p-STAT6 could bind to the promoter of TXNIP and promote its transcription in the nucleus and the interaction of STAT6 and TXNIP could enhance the NLRP3 inflammasome activation. Subsequent the TXNIP shuttling from nucleus to mitochondria increased the NLRP3-mediated TIF both in high glucose-stimulated renal tubular epithelial cells and in db/db mouse. Knockdown of TXNIP both in vitro and in vivo delayed the ROS associated inflammasome activation. Meanwhile, TXNIP was the target for Astragaloside IV to exert inhibitory effect on the STAT6/TXNIP/NLRP3-mediated inflammatory response. Conclusion: These findings collectively substantiate a critical role of TXNIP shuttling to control inflammasome activation, and presenting AS-IV as a promising therapeutic ingredient for inhibiting TIF in DKD. Funding: Private Foundation Support