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Tubulointerstitial fibrosis is observed in diabetic nephropathy. It is still debated whether tubular cells, undergoing epithelial-mesenchymal transition (EMT) in high glucose (HG) conditions, may contribute to interstitial fibrosis development. In this study, we investigated the phenotypic and molecular EMT-like changes and the alteration of inflammatory and fibrogenic secretome induced by HG in human primary tubular cell cultures. Taking advantage of this in vitro cell model composed of proximal and distal tubular cells, we showed that HG-treated tubular cells acquired a fibroblast-like morphology with increased cytoplasmic stress fibers, maintaining the expression of the epithelial markers specific of proximal and distal tubular cells. HG increased Snail1, miRNA210 and Vimentin mesenchymal markers, decreased N-cadherin expression and migration ability of primary tubular cells, while E-cadherin expression and focal adhesion distribution were not affected. Furthermore, HG treatment of tubular cells altered the inflammatory cytokine secretion creating a secretome able to enhance the proliferation and migration of fibroblasts. Our findings show that HG promotes an activated state of partial EMT in human tubular primary cells and induces a pro-inflammatory and pro-fibrogenic microenvironment, supporting the active role of tubular cells in diabetic nephropathy onset., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Torsello et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Aims: Diabetic nephropathy is a major consequence of inflammation developing in type 1 diabetes, with interleukin‐8 (IL‐8)‐CXCR1/2 axis playing a key role in kidney disease progression. In this study, we investigated the therapeutic potential of a CXCR1/2 non‐competitive allosteric antagonist (Ladarixin) in preventing high glucose‐mediated injury in human podocytes and epithelial cells differentiated from renal stem/progenitor cells (RSC) cultured as nephrospheres. Materials and Methods: We used human RSCs cultured as nephrospheres through a sphere‐forming functional assay to investigate hyperglycemia‐mediated effects on IL‐8 signalling in human podocytes and tubular epithelial cells. Results: High glucose impairs RSC self‐renewal, induces an increase in IL‐8 transcript expression and protein secretion and induces DNA damage in RSC‐differentiated podocytes, while exerting no effect on RSC‐differentiated epithelial cells. Accordingly, the supernatant from epithelial cells or podocytes cultured in high glucose was able to differentially activate leucocyte‐mediated secretion of pro‐inflammatory cytokines, suggesting that the crosstalk between immune and non‐immune cells may be involved in disease progression in vivo. Conclusions: Treatment with Ladarixin during RSC differentiation prevented high glucose‐mediated effects on podocytes and modulated either podocyte or epithelial cell‐dependent leucocyte secretion of pro‐inflammatory cytokines, suggesting CXCR1/2 antagonists as possible pharmacological approaches for the treatment of diabetic nephropathy. [ABSTRACT FROM AUTHOR]

Renal tubular cells are involved in the tubular interstitial fibrosis observed in diabetic nephropathy. It is debated whether epithelial–mesenchymal transition (EMT) affects tubular cells, which under highglucose conditions overproduce transforming growth factor-β (TGF-β), a fibrogenic cytokine involved in interstitial fibrosis development. Our study investigated the involvement of non-receptor tyrosine kinase Arg (also called Abl2) in TGF-β production. Human primary tubular cell cultures exposed to high-glucose conditions were used. These cells showed an elongated morphology, stress fibers and vimentin increment but maintained most of the epithelial marker expression and distribution. In these cells exposed to high glucose, which overexpressed and secreted active TGF-β1, Arg protein and activity was downregulated. A further TGF-β1 increase was induced by Arg silencing with siRNA, as with the Arg tyrosine kinase inhibitor Imatinib. In the cells exposed to high glucose, reactive oxygen species (ROS)-dependent Arg kinase downregulation induced both RhoA activation, through p190RhoGAPA (also known as ARHGAP35) modulation, and proteasome activity inhibition. These data evidence a new specific involvement of Arg kinase into the regulation of TGF-β1 expression in tubular cells under high-glucose conditions and provide cues for new translational approaches in diabetic nephropathy. [ABSTRACT FROM AUTHOR]