Hyperglycemia upregulates thioredoxin interacting protein (TXNIP) expression, which in turn induces ROS production, inflammatory and fibrotic responses in the diabetic kidney. Dysregulation of autophagy contributes to the development of diabetic nephropathy. However, the interaction of TXNIP with autophagy/mitophagy in diabetic nephropathy is unknown. In this study, streptozotocin-induced diabetic rats were given TXNIP DNAzyme or scrambled DNAzyme for 12 weeks respectively. Fibrotic markers, mitochondrial function and mitochondrial reactive oxygen species (mtROS) were assessed in kidneys. Tubular autophagy and mitophagy were determined in kidneys from both human and rats with diabetic nephropathy. TXNIP and autophagic signaling molecules were examined. TXNIP DNAzyme dramatically attenuated extracellular matrix deposition in the diabetic kidneys compared to the control DNAzyme. Accumulation of autophagosomes and reduced autophagic clearance were shown in tubular cells of human diabetic compared to non-diabetic kidneys, which was reversed by TXNIP DNAzyme. High glucose induced mitochondrial dysfunction and mtROS production and inhibited mitophagy in proximal tubular cells, which was reversed by TXNIP siRNA. TXNIP inhibition suppressed diabetes-induced BNIP3 expression and activation of the mTOR signaling pathway. Collectively, hyperglycemia-induced TXNIP contributes to the dysregulation of tubular autophagy and mitophagy in diabetic nephropathy through activation of the mTOR signaling pathway.

Acute kidney injury (AKI) and chronic kidney disease (CKD) are rising in global prevalence and cause significant morbidity for patients. Current treatments are limited to slowing instead of stabilising or reversing disease progression. In this review, we describe mesenchymal stem cells (MSCs) and their constituents, extracellular vesicles (EVs) as being a novel therapeutic for CKD. MSC-derived EVs (MSC-EVs) are membrane-enclosed particles, including exosomes, which carry genetic information that mimics the phenotype of their cell of origin. MSC-EVs deliver their cargo of mRNA, miRNA, cytokines, and growth factors to target cells as a form of paracrine communication. This genetically reprograms pathophysiological pathways, which are upregulated in renal failure. Since the method of exosome preparation significantly affects the quality and function of MSC-exosomes, this review compares the methodologies for isolating exosomes from MSCs and their role in tissue regeneration. More specifically, it summarises the therapeutic efficacy of MSC-EVs in 60 preclinical animal models of AKI and CKD and the cargo of biomolecules they deliver. MSC-EVs promote tubular proliferation and angiogenesis, and inhibit apoptosis, oxidative stress, inflammation, the epithelial-to-mesenchymal transition, and fibrosis, to alleviate AKI and CKD. By reprogramming these pathophysiological pathways, MSC-EVs can slow or even reverse the progression of AKI to CKD, and therefore offer potential to transform clinical practice.

The Ca²⁺-activated K⁺ channel KCa3.1 mediates cellular signaling processes associated with dysfunction of vasculature. However, the role of KCa3.1 in diabetic nephropathy is unknown. We sought to assess whether KCa3.1 mediates the development of renal fibrosis in two animal models of diabetic nephropathy. Wild-type and KCa3.1^−/− mice, and secondly eNOS^−/− mice, had diabetes induced with streptozotocin and then were treated with/without a selective inhibitor of KCa3.1 (TRAM34). Our results show that the albumin-to-creatinine ratio significantly decreased in diabetic KCa3.1^−/− mice compared with diabetic wild-type mice and in diabetic eNOS^−/− mice treated with TRAM34 compared with diabetic mice. The expression of monocyte chemoattractant protein-1 (MCP-1), intercellular adhesion molecule 1 (ICAM1), F4/80, plasminogen activator inhibitor type 1 (PAI-1), and type III and IV collagen significantly decreased (P < 0.01) in kidneys of diabetic KCa3.1^−/− mice compared with diabetic wild-type mice. Similarly, TRAM34 reduced the expression of the inflammatory and fibrotic markers described above in diabetic eNOS^−/− mice. Furthermore, blocking the KCa3.1 channel in both animal models led to a reduction of transforming growth factor-β1 (TGF-β1) and TGF-β1 type II receptor (TβRII) and phosphorylation of Smad2/3. Our results provide evidence that KCa3.1 mediates renal fibrosis in diabetic nephropathy through the TGF-β1/Smad signaling pathway. Blockade of KCa3.1 may be a novel target for therapeutic intervention in patients with diabetic nephropathy.

MicroRNAs (miRNAs) comprise of a novel class of endogenous small noncoding RNAs that frequently downregulate the expression of target genes. Recent reports suggest that miRNA-200b prevents epithelial-to-mesenchymal transition (EMT) in cancer cells by targeting the E-box binding transcription factors Zinc finger E-box-binding homeobox 1 (ZEB1) and Zinc finger E-box-binding homeobox 2 (ZEB2). About 35% of active fibroblasts are derived from EMT which is central to the development of progressive renal fibrosis. Hence, this study was designed to assess the effect of miRNA-200b on transforming growth factor-β (TGF-β1)-induced fibrotic responses in renal tubular cells. Morphologically, human kidney-2 cells transfected with miRNA-200b retained their epithelial cell characteristics when exposed to TGF-β1. miRNA-200b significantly increased E-cadherin (P < 0.001) and reduced fibronectin mRNA and protein expression (both P < 0.01) independent of phospho-Smad2/3 and phospho-p38 and p42/44 signaling. Increased E-cadherin expression was associated with decreased expression of ZEB1 and ZEB2 and repression of fibronectin was mediated through direct targeting of the fibronectin mRNA, demonstrated using pMIR luciferase reporter assay and site-directed mutagenesis. These results suggest that miRNA-200b suppresses TGF-β1-induced EMT via inhibition of ZEB1 and ZEB2 and the extracellular matrix protein fibronectin by directing targeting of its 3′UTR mRNA, independent of pathways directly involved in TGF-β1 signaling.

Autophagy is emerging as an important pathway in many diseases including diabetic nephropathy. It is acknowledged that oxidative stress plays a critical role in autophagy dysfunction and diabetic nephropathy and KCa3.1 blockade ameliorates diabetic renal fibrosis through inhibiting TGF-β1 signaling pathway. To identify the role of KCa3.1 in dysfunctional tubular autophagy in diabetic nephropathy, human proximal tubular cells (HK2) transfected with scrambled or KCa3.1 siRNAs were exposed to TGF-β1 for 48 h, then autophagosome formation, the autophagy marker LC3, signaling molecules PI3K, Akt and mTOR and oxidative stress marker nitrotyrosine were examined respectively. In vivo, LC3, nitrotyrosine and phosphorylated mTOR were examined in kidneys of diabetic KCa3.1+/+ and KCa3.1−/− mice. The results demonstrated that TGF-β1 increased the formation of autophagic vacuoles, LC3 expression and phosphorylation of PI3K, Akt and mTOR in scrambled siRNA transfected HK2 cells compared to control cells, which was reversed in KCa3.1 siRNA transfected HK2 cells. In vivo, expression of LC3 and nitrotyrosine and phosphorylation of mTOR were significantly increased in kidneys of diabetic KCa3.1+/+ mice compared to non-diabetic mice, which were attenuated in kidneys of diabetic KCa3.1−/− mice. These results suggest that KCa3.1 activation contributes to dysfunctional tubular autophagy in diabetic nephropathy through PI3K/Akt/mTOR signaling pathways.