FSP1-specific SMAD2 knockout in renal tubular, endothelial, and interstitial cells reduces fibrosis and epithelial-to-mesenchymal transition in murine STZ-induced diabetic nephropathy

Abstract

Extracellular matrix deposition during tubulointerstitial fibrosis (TIF), a central pathological process in patients with diabetic nephropathy (DN), is driven by locally activated, disease-relevant myofibroblasts. Myofibroblasts can arise from various cellular sources, e.g., tubular epithelial cells via a process named epithelial-to-mesenchymal transition (EMT). Transforming growth factor beta 1 (TGF-β1) and its downstream Smad signaling play a critical role in both TIF and EMT. Whereas Smad3 is one central mediator, the role of the other prominently expressed variant, Smad2, is not completely understood. In this study, we sought to analyze the role of renal Smad2 in the development of TIF and EMT during streptozotocin-induced DN by using a fibroblast-specific protein 1 (FSP1)-promotor-driven SMAD2 knockout mouse model with decreased tubular, endothelial, and interstitial Smad2 expression. In contrast to wild-type diabetic mice, diabetic SMAD2 knockout mice showed the following features: (1) significantly reduced DN and TIF (shown by KIM1 expression; periodic acid Schiff staining; collagen I and III, fibronectin, and connective tissue growth factor deposition); (2) significantly reduced tubular EMT-like changes (e.g., altered Snail1, E-cadherin, matrix metalloproteinase 2, and vimentin deposition); and (3) significantly decreased expression of myofibroblast markers (α-smooth muscle actin, FSP1). As one mechanism for the protection against diabetes-induced TIF and EMT, decreased Smad3 protein levels and, as a possible consequence, reduced TGF-β1 levels were observed in diabetic SMAD2 knockout mice. Our findings thus support the important role of Smad2 for pro-fibrotic TGF-β/Smad3 signaling in experimental DN.

New insights into TGF-β/Smad signaling in tissue fibrosis

“…The mice with constitutive deletion of Smad2 were embryonic lethal [44]. Conditional deletion of Smad2 could significantly attenuate renal fibrosis, tubular EMT-like changes and the levels of myofibroblast markers in diabetic nephropathy mice, as well as decreased Smad3 and TGF-β1 [45]. Besides, Smad2 deletion promoted fibrosis through enhanced TGF-β/Smad3 signaling, and increased autoinduction of TGF-β1.…”

Transforming Growth Factor-Beta1 in Diabetic Kidney Disease

“…Conversely, unlike Smad3, the function of Smad2 in DKD is unclear. Overexpression of Smad2 attenuated TGF-β1induced phosphorylated Smad3 and collagen expression, whereas deletion of Smad2 promoted renal fibrosis via substantially enhanced Smad3 signaling (Meng et al, 2010;Loeffler et al, 2018). Although Smad2 interacts with Smad3 physically, Smad2 and -3 may compete for phosphorylation in response to TGF-β1 stimulation.…”

Diverse Role of TGF-β in Kidney Disease

“…In canonical TGF-β signaling, Smad2, and Smad3 are two key downstream mediators that are highly activated in the fibrotic kidney (Wang et al, 2006;Chung et al, 2010b;Zhou et al, 2010;Loeffler et al, 2018). Although Smad2 and Smad3 bind together, their functional roles are distinct.…”

“…However, a recent finding that conditional deletion of Smad2 from TECs accelerates renal fibrosis reveals a protective role of Smad2 in renal fibrosis (Meng et al, 2010). In addition, FSP1-specific Smad2 knockout in renal tubular, endothelial, and interstitial cells is also reported to reduce renal fibrosis and epithelial-to-mesenchymal transition in murine streptozotocin (STZ)-induced diabetic nephropathy (Loeffler et al, 2018).…”