Tubular Overexpression of Gremlin Induces Renal Damage Susceptibility in Mice (original) (raw)
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Tubular Overexpression of Gremlin in Transgenic Mice Aggravates Renal Damage in Diabetic Nephropathy
American Journal of Physiology - Renal Physiology, 2015
Diabetic nephropathy (DN) is currently a leading cause of end-stage renal failure worldwide. Gremlin was identified as a gene differentially expressed in mesangial cells exposed to high glucose and in experimental diabetic kidneys. We have described that Gremlin is highly expressed in biopsies from patients with diabetic nephropathy, predominantly in areas of tubulointerstitial fibrosis. In streptozotocin (STZ)-induced experimental diabetes, Gremlin deletion using Grem1 heterozygous knockout mice or by gene silencing, ameliorates renal damage. To study the in vivo role of Gremlin in renal damage, we developed a diabetic model induced by STZ in transgenic (TG) mice expressing human Gremlin in proximal tubular epithelial cells. The albuminuria/creatinuria ratio, determined at week 20 after treatment, was significantly increased in diabetic mice but with no significant differences between transgenic (TG/STZ) and wild-type mice (WT/STZ). To assess the level of renal damage, kidney tissu...
Nephron Experimental Nephrology, 2012
brotic genes, such as TGF-β 1 , and augmented the production of ECM proteins, including type I collagen. The blockade of endogenous Gremlin with small interfering RNA inhibited TGF-β 1 -induced ECM upregulation. In tubular epithelial cells Gremlin also increased profibrotic genes and caused EMT changes: phenotypic modulation to myofibroblast-like morphology, loss of epithelial markers and in duction of mesenchymal markers. Moreover, Gremlin gene silencing inhibited TGF-β 1 -induced EMT changes. Conclusions: Gremlin directly activates profibrotic events in cul tured renal fibroblasts and tubular epithelial cells. Moreover, endogenous Gremlin blockade inhibited TGF-β-mediated matrix production and EMT, suggesting that Gremlin could be a novel therapeutic target for renal fibrosis.
Transplantation Proceedings, 2008
Background. Chronic allograft nephropathy (CAN) is the most frequent cause of chronic dysfunction and late loss of renal allografts. Epithelial mesenchymal transition (EMT) has been identified as responsible for the presence of activated interstitial fibroblasts (myofibroblasts) and transforming growth factor beta (TGF-)/Smad is the key signaling mediator. It has been proposed that the bone morphogenetic protein 7 (BMP-7) antagonist, Gremlin, could participate in EMT, as a downstream mediator of TGF-. Methods. We evaluated 33 renal allograft biopsies, 16 of which showed CAN, versus 17 controls. By in situ hybridization we studied the expression of TGF- and Gremlin mRNA. Gremlin, BMP-7, E-cadherin, and ␣-smooth muscle actin (␣-SMA) proteins were evaluated by immunohistochemistry and Smad3 activation by Southwestern. In cultured human tubuloepithelial cells (HK2 cell line), Gremlin induction by TGF- was studied by confocal microscopy. Results. Among renal biopsies of transplanted patients with CAN, we detected upregulation of TGF- in colocalization with Gremlin (RNA and protein), mainly in areas of tubulointerstitial fibrosis. In the same tubules, we observed decreased expression of E-cadherin and induction of vimentin and ␣-SMA. BMP-7 was significantly decreased in the CAN biopsies. In addition, HK2 stimulated with TGF- (1 ng/mL) induced Gremlin production at 72 hours. Conclusion. We postulated that Gremlin is a downstream mediator of TGF-, suggesting a role for Gremlin in EMT observed in CAN.
Frontiers in pharmacology, 2018
Chronic kidney disease (CKD) is emerging as an important health problem due to the increase number of CKD patients and the absence of an effective curative treatment. Gremlin has been proposed as a novel therapeutic target for renal inflammatory diseases, acting via Vascular Endothelial Growth Factor Receptor-2 (VEGFR2). Although many evidences suggest that Gremlin could regulate renal fibrosis, the receptor involved has not been yet clarified. Gremlin, as other TGF-β superfamily members, regulates tubular epithelial to mesenchymal transition (EMT) and, therefore, could contribute to renal fibrosis. In cultured tubular epithelial cells Gremlin binding to VEGFR2 is linked to proinflammatory responses. Now, we have found out that in these cells VEGFR2 is also involved in the profibrotic actions of Gremlin. VEGFR2 blockade by a pharmacological kinase inhibitor or gene silencing diminished Gremlin-mediated gene upregulation of profibrotic factors and restored changes in EMT-related gene...
Gremlin regulates renal inflammation via vascular endothelial growth factor receptor 2 pathway
The Journal of Pathology, 2015
Inflammation is a main feature of progressive kidney disease. Gremlin binds to bone morphogenetic proteins (BMPs), acting as an antagonist and regulating nephrogenesis and fibrosis among other processes. Gremlin also binds to vascular endothelial growth factor receptor-2 (VEGFR2) in endothelial cells to induce angiogenesis. In renal cells, Gremlin regulates proliferation and fibrosis, but there are no data about inflammatory-related events. We have investigated the direct effects of Gremlin in the kidney, evaluating whether VEGFR2 is a functional Gremlin receptor. Administration of recombinant Gremlin to murine kidneys induced rapid and sustained activation of VEGFR2 signalling, located in proximal tubular epithelial cells. Gremlin bound to VEGFR2 in these cells in vitro, activating this signalling pathway independently of its action as an antagonist of BMPs. In vivo, Gremlin caused early renal damage, characterized by activation of the nuclear factor-κB pathway linked to up-regulation of proinflammatory factors and infiltration of immune inflammatory cells. VEGFR2 blockade diminished Gremlin-induced renal inflammatory responses. The link between Gremlin/VEGFR2 and NF-κB/inflammation was confirmed in vitro. Gremlin overexpression was associated to VEGFR2 activation in human renal disease and in the unilateral ureteral obstruction experimental model, where VEGFR2 kinase inhibition diminished renal inflammation. Our data show that a Gremlin/VEGFR2 axis participates in renal inflammation and could be a novel target for kidney disease.
Nephrology Dialysis Transplantation, 2007
Background. Recent evidence in vitro and in vivo suggests that gremlin, a bone morphogenetic protein antagonist, is participating in tubular epithelial mesenchymal transition (EMT) in diabetic nephropathy as a downstream mediator of TGF-b. Since EMT also occurs in parietal epithelial glomerular cells (PECs) leading to crescent formation, we hypothesized that gremlin could participate in this process. With this aim we studied its expression in 30 renal biopsies of patients with pauci-immune crescentic nephritis. Methods. Gremlin was detected by in situ hybridization (ISH) and immunohistochemistry (IMH) and TGF-b by ISH and Smads by southwestern histochemistry (SWH). Phosphorylated Smad2, CTGF, BMP-7, PCNA, a-SMA, synaptopodin, CD-68, and phenotypic markers of PECs (cytokeratin, E-cadherin), were detected by IMH. In cultured human monocytes, gremlin and CTGF induction by TGF-b was studied by western blot. Results. We observed strong expression of gremlin mRNA and protein in cellular and fibrocellular crescents corresponding to proliferating PECs and monocytes, in co-localization with TGF-b. A marked over-expression of gremlin was also observed in tubular and infiltrating interstitial cells, correlating with tubulointerstitial fibrosis (r ¼ 0.59; P < 0.01). A nuclear Smad activation in the same tubular cells, that are expressing TGF-b and gremlin, was detected. In human cultured monocytes, TGF-b induced gremlin production while CTGF expression was not detected. Conclusion. We postulate that gremlin may play a role in the fibrous process in crescentic nephritis, both in glomerular crescentic and tubular epithelial cells. The co-localization of gremlin and TGF-b expression found in glomeruli and tubular cells suggest that gremlin may be important in mediating some of the pathological effects of TGF-b.
Gremlin, A Potential Urinary Biomarker of Anca-Associated Crescentic Glomerulonephritis
Scientific Reports, 2019
Gremlin renal overexpression has been reported in diabetic nephropathy, pauci-immune crescentic glomerulonephritis and chronic allograft nephropathy and has been implicated in the pathophysiology of the progression of renal damage. However, it is unknown whether urinary Gremlin can be associated with renal functional status, renal biopsy findings and outcome. To examine these associations we studied 20 patients with ANCA+ renal vasculitis and very high urinary Gremlin (354 ± 76 ug/gCr), 86 patients with other glomerular diseases and moderately elevated urinary Gremlin (83 ± 14 ug/gCr) and 11 healthy controls (urinary Gremlin 11.3 ± 2.4 ug/gCr). Urinary Gremlin was significantly correlated with renal expression of Gremlin (r = 0.64, p = 0.013) observed in cellular glomerular crescents, tubular epithelial cells and interstitial inflammatory cells. Moreover, urinary Gremlin levels were correlated with the number of glomerular crescents (r = 0.53; p < 0.001), renal CD68 positive cells (r = 0.71; p < 0.005), tubulointerstitial fibrosis (r = 0.50; p < 0.05), and serum creatinine levels (r = 0.60; p < 0.001). Interestingly, Gremlin expression was colocalized with CD68, CD163 (monocyte/macrophage markers) and CCL18 positive cells. ROC curve analysis showed that the cutoff value of urinary Gremlin in glomerular diseases as 43 ug/gCr with 72% of sensitivity and 100% of specificity [AUC: 0.96 (CI 95% 0.92-0.99] (p < 0.001). For ANCA+ renal vasculitis the value of urinary Gremlin of 241 ug/gCr had 55% of sensitivity and 100% of specificity [AUC: 0.81 (CI 95% 0.68-0.94) (p < 0.001]. Based on these results we propose that urinary Gremlin represents a non-invasive biomarker in ANCA+ renal vasculitis, and suggest a role of Gremlin in the formation of crescents. Gremlin was identified as one of developmental genes, re-expressed during renal damage that could play a role in the progression of diabetic nephropathy and other chronic renal diseases 1. Gremlin belongs to a family of bone morphogenetic proteins (BMPs) antagonists 2-4 that have a well-recognized role in organogenesis and fibrotic related disorders 5-7 Gremlin is induced by TGF-β in renal cells 2 , and it is a downstream mediator of TGF-β, activating the Smad pathway and inducing epithelial-mesenchymal transition 8. Moreover, in a BMP-independent manner, Gremlin could contribute to renal inflammation via vascular endothelial growth factor receptor 2 pathway 9. Gremlin knockout mice die of renal aplasia, lung defects and limb malformations 7,10-14 , and we have recently shown that Gremlin tubular overexpression aggravates folic acid-induced and streptozotocin-induced renal damage 15,16. In human diabetic nephropathy, Gremlin is most prominently expressed in podocytes and areas of tubulointerstitial fibrosis, colocalized with TGF-β expression and correlated with tubulointerstitial score damage 17,18. Gremlin has also been observed in various nephropathies, notably in pauci-immune crescentic vasculitis where strong expression of Gremlin mRNA and protein were observed at glomerular crescents, and in tubular and infiltrating interstitial cells 19. However, it is not known if Gremlin is present in the urine during kidney injury and whether it could be a novel biomarker with diagnostic or prognostic value in glomerular diseases. The present study was done to answer these questions.
Nephrology Dialysis Transplantation, 2008
Background. Recent evidence in vitro and in vivo suggests that gremlin, a bone morphogenetic protein antagonist, is participating in tubular epithelial mesenchymal transition (EMT) in diabetic nephropathy as a downstream mediator of TGF-b. Since EMT also occurs in parietal epithelial glomerular cells (PECs) leading to crescent formation, we hypothesized that gremlin could participate in this process. With this aim we studied its expression in 30 renal biopsies of patients with pauci-immune crescentic nephritis. Methods. Gremlin was detected by in situ hybridization (ISH) and immunohistochemistry (IMH) and TGF-b by ISH and Smads by southwestern histochemistry (SWH). Phosphorylated Smad2, CTGF, BMP-7, PCNA, a-SMA, synaptopodin, CD-68, and phenotypic markers of PECs (cytokeratin, E-cadherin), were detected by IMH. In cultured human monocytes, gremlin and CTGF induction by TGF-b was studied by western blot. Results. We observed strong expression of gremlin mRNA and protein in cellular and fibrocellular crescents corresponding to proliferating PECs and monocytes, in co-localization with TGF-b. A marked over-expression of gremlin was also observed in tubular and infiltrating interstitial cells, correlating with tubulointerstitial fibrosis (r ¼ 0.59; P < 0.01). A nuclear Smad activation in the same tubular cells, that are expressing TGF-b and gremlin, was detected. In human cultured monocytes, TGF-b induced gremlin production while CTGF expression was not detected. Conclusion. We postulate that gremlin may play a role in the fibrous process in crescentic nephritis, both in glomerular crescentic and tubular epithelial cells. The co-localization of gremlin and TGF-b expression found in glomeruli and tubular cells suggest that gremlin may be important in mediating some of the pathological effects of TGF-b.
Expression of Gremlin, a Bone Morphogenetic Protein Antagonist, in Human Diabetic Nephropathy
American Journal of Kidney Diseases, 2005
Background: We report the induction of gremlin, a bone morphogenetic protein antagonist, in cultured human mesangial cells exposed to high glucose and transforming growth factor  (TGF-) levels in vitro and kidneys from diabetic rats in vivo. Methods: Gremlin expression was assessed in human diabetic nephropathy by means of in situ hybridization, immunohistochemistry, and real-time polymerase chain reaction and correlated with clinical and pathological indices of disease. Results: Gremlin was not expressed in normal human adult kidneys. Conversely, abundant gremlin expression was observed in human diabetic nephropathy. Although some gremlin expression was observed in occasional glomeruli, gremlin expression was most prominent in areas of tubulointerstitial fibrosis, where it colocalized with TGF- expression. Gremlin messenger RNA levels correlated directly with renal dysfunction, determined by means of serum creatinine level, but not with proteinuria level. There was a strong correlation between gremlin expression and tubulointerstitial fibrosis score. Conclusion: In aggregate, these results indicate that the developmental gene gremlin reemerges in the context of tubulointerstitial fibrosis in diabetic nephropathy and suggests a role for TFG- as an inducer of gremlin expression in this context. Am J Kidney Dis 45:1034-1039.