ZnT8 Exerts Anti-apoptosis of Kidney Tubular Epithelial Cell in Diabetic Kidney Disease Through TNFAIP3-NF-κB Signal Pathways.

Apoptosis of kidney tubular epithelial cells contributes to diabetic kidney disease (DKD) pathophysiology, but the mechanisms are not fully understood. Zinc transporter protein member 8 (ZnT8, SLC30A8) is a susceptive gene in diabetes. Here, we aim to investigate whether ZnT8 has effects on pathophysiology of DKD. The animal groups include control, ZnT8KO mice, STZ-induced, and ZnT8-KO-STZ. STZ-induced DKD was performed in male C57BL/6 J mice and in ZnT8-KO mice. High glucose (HG)-induced apoptosis in a normal rat kidney tubular epithelial cell line (NRK-52E cells) was performed in vitro. Transfection of hZnT8-EGFP or TNFAIP3 siRNA was done in NRK-52E cells. Flow cytometry with Annexin V-FITC/PI double staining and TUNEL analysis was performed for the detection of apoptosis. Gene expression at mRNA and protein levels was examined with real-time RT-PCR and Western blot. Urine albumin to creatinine ratio, proinflammatory cytokines, and apoptosis were enhanced in kidneys of STZ and ZnT8-KO-STZ mice compared to control or ZnT8-KO mice. ZnT8 overexpression after hZnT8-EGFP transfection decreased HG-stimulated inflammatory activity and apoptosis in NRK-52E cells. Furthermore, treatment with ZnSO4 blunted HG-induced apoptosis and NF-κB activation. ZnSO4 increased the abundance of zinc-finger protein TNF-α-induced protein 3 (TNFAIP3). Also, ZnT8 over-expression after hZnT8-EGFP transfection significantly ameliorates HG-induced NF-κB-dependent transcriptional activity and apoptotic protein expressions in NRK-52E cells, but the inhibitory effect of ZnT8 was significantly abolished with TNFAIP3 siRNA. Our study provides evidence that ZnT8 has protective effects against apoptosis of renal tubular epithelial cells through induction of TNFAIP3 and subsequent suppression of the NF-κB pathway.

Effects of Curcumin on High Glucose-Induced Epithelial-to-Mesenchymal Transition in Renal Tubular Epithelial Cells Through the TLR4-NF-κB Signaling Pathway.

OBJECTIVE: Diabetic kidney disease (DKD) is a microvascular complication in diabetes mellitus, while tubuloepithelial to mesenchymal transition (EMT) of mature tubular epithelial cells is a key point in the early development and progression of renal interstitial fibrosis. The present study aimed to investigate the protective effects of Curcumin on EMT and fibrosis in cultured normal rat kidney tubular epithelial cell line (NRK-52E). METHODS: By using immunofluorescence staining and Western blot protocols, in vitro experiments were designed to analyze EMT markers, including collagen I and E-cadherin in high glucose (HG) exposed NRK-52E cells and to detect the expression levels of phosphorylated-NF-κB, TLR4 and reactive oxygen species (ROS) after Curcumin pre-treatment. With co-treatment with TAK242, these molecules in the TLR4-NF-κB signaling pathway were further evaluated. RESULTS: Curcumin decreased the HG-induced EMT levels and ROS production in NRK-52E cells. Furthermore, Curcumin was found to inhibit the TLR4-NF-κB signaling activation in HG-induced EMT of NRK-52E cells. CONCLUSION: The present study provides evidence suggesting a novel mechanism that Curcumin exerts the anti-fibrosis effects via inhibiting activation of the TLR4-NF-κB signal pathway and consequently protecting the HG-induced EMT in renal tubular epithelial cells. Thereby, TLR4-NF-κB may be a useful target for therapeutic intervention in DKD.