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Rolf A.k. Stahl - One of the best experts on this subject based on the ideXlab platform.
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the il 23 th17 axis contributes to renal injury in Experimental Glomerulonephritis
Journal of The American Society of Nephrology, 2009Co-Authors: Hansjoachim Paust, Janeric Turner, Oliver M Steinmetz, Anett Peters, Felix Heymann, Christoph Holscher, Gunter Wolf, Christian Kurts, Hanswilli Mittrucker, Rolf A.k. StahlAbstract:T cells infiltrate the kidney in both human and Experimental Glomerulonephritis, and several lines of evidence indicate that T cell-mediated tissue damage plays an important role in the immunopathogenesis of renal inflammatory diseases. However, the functions of the different T cell subsets, particularly the recently identified interleukin-17 (IL-17)-producing T cells (Th17 cells), are incompletely understood in Glomerulonephritis. Here, we identified renal IL-17-producing T cells in the T cell-mediated model of nephrotoxic nephritis in mice. In vitro, IL-17 enhanced the production of the proinflammatory chemokines CCL2/MCP-1, CCL3/MIP-1α, and CCL20/LARC, which are implicated in the recruitment of T cells and monocytes, in mouse mesangial cells. To determine the function of Th17 cells in renal inflammation, we induced nephrotoxic nephritis in IL-23 p19−/− mice, which have reduced numbers of Th17 cells, and in IL-17−/− mice, which are deficient in the effector cytokine IL-17 itself. In comparison with nephritic wild-type mice, IL-23 p19−/− mice demonstrated less infiltration of Th17 cells, and both IL-23 p19−/− and IL-17−/− mice developed less severe nephritis as measured by renal function, albuminuria, and frequency of glomerular crescent formation. These results demonstrate that the IL-23/IL-17 pathway significantly contributes to renal tissue injury in Experimental Glomerulonephritis. Targeting the IL-23/Th17 axis may be a promising therapeutic strategy for the treatment of proliferative and crescentic Glomerulonephritis.
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The IL-23/Th17 Axis Contributes to Renal Injury in Experimental Glomerulonephritis
Journal of the American Society of Nephrology : JASN, 2009Co-Authors: Hansjoachim Paust, Janeric Turner, Oliver M Steinmetz, Anett Peters, Felix Heymann, Christoph Holscher, Gunter Wolf, Christian Kurts, Hanswilli Mittrucker, Rolf A.k. StahlAbstract:T cells infiltrate the kidney in both human and Experimental Glomerulonephritis, and several lines of evidence indicate that T cell-mediated tissue damage plays an important role in the immunopathogenesis of renal inflammatory diseases. However, the functions of the different T cell subsets, particularly the recently identified interleukin-17 (IL-17)-producing T cells (Th17 cells), are incompletely understood in Glomerulonephritis. Here, we identified renal IL-17-producing T cells in the T cell-mediated model of nephrotoxic nephritis in mice. In vitro, IL-17 enhanced the production of the proinflammatory chemokines CCL2/MCP-1, CCL3/MIP-1alpha, and CCL20/LARC, which are implicated in the recruitment of T cells and monocytes, in mouse mesangial cells. To determine the function of Th17 cells in renal inflammation, we induced nephrotoxic nephritis in IL-23 p19(-/-) mice, which have reduced numbers of Th17 cells, and in IL-17(-/-) mice, which are deficient in the effector cytokine IL-17 itself. In comparison with nephritic wild-type mice, IL-23 p19(-/-) mice demonstrated less infiltration of Th17 cells, and both IL-23 p19(-/-) and IL-17(-/-) mice developed less severe nephritis as measured by renal function, albuminuria, and frequency of glomerular crescent formation. These results demonstrate that the IL-23/IL-17 pathway significantly contributes to renal tissue injury in Experimental Glomerulonephritis. Targeting the IL-23/Th17 axis may be a promising therapeutic strategy for the treatment of proliferative and crescentic Glomerulonephritis.
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Effects of different PPARγ-agonists on MCP-1 expression and monocyte recruitment in Experimental Glomerulonephritis
Kidney international, 2002Co-Authors: Ulf Panzer, André Schneider, Gunter Wolf, Gunther Zahner, Friedrich Thaiss, Youfei Guan, Rüdiger Reinking, Sigrid Harendza, Rolf A.k. StahlAbstract:Effects of different PPARγ-agonists on MCP-1 expression and monocyte recruitment in Experimental Glomerulonephritis. Background Activators of peroxisome proliferator activated receptor γ (PPARγ) have been shown to modulate chemokine expression in isolated monocytes/macrophages (M/M) and to exert anti-inflammatory effects in some models of Experimental inflammatory diseases. We evaluated the effects of different forms of PPARγ activators in a model of Experimental Glomerulonephritis (GN) in rats. Methods GN was induced in rats by application of an anti-thymocyte antibody (ATS). Nephritic rats were treated with two synthetic PPARγ ligands of the thiazolidinedione (TZD) group, troglitazone (200 mg/kg/day) and ciglitazone (100 mg/kg/day), and with a natural ligand 15d-PGJ 2 (1.5 mg/day). Twenty-four hours after induction of the GN, the glomerular mRNA expression of the chemokine monocyte chemoattractant protein-1 (MCP-1) and the cognate chemokine receptor CCR-2 were examined by Northern blotting and RT-PCR. The glomerular M/M infiltration was determined by immunohistology. The activation of the transcription factors PPARγ, nuclear factor-βB (NF-βB) and activator protein-1 (AP-1) in glomeruli was analyzed by electrophoretic mobility shift assay. Results Induction of GN up-regulated glomerular nuclear protein binding of NF-βB and AP-1. Treatment of nephritic rats with troglitazone and ciglitazone augmented nuclear PPARγ and AP-1 DNA binding but did not affect NF-βB binding. TZD enhanced glomerular MCP-1 expression and increased glomerular M/M recruitment. In contrast, 15d-PGJ 2 attenuated NF-βB activation and did not affect AP-1 activity or MCP-1 expression. Conclusion Our data show that PPARγ activators of the TZD group, but not 15d-PGJ 2 , enhance MCP-1 expression and M/M infiltration in the induction phase of Experimental GN. The results demonstrate that TZD and 15d-PGJ 2 may exert different effects in the immune response in Experimental GN. Our study underscores the need to critically evaluate whether PPARγ ligands will have beneficial or possibly deleterious effects in GN.
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Monocyte chemoattractant protein-1 and osteopontin differentially regulate monocytes recruitment in Experimental Glomerulonephritis
Kidney international, 2001Co-Authors: Ulf Panzer, Gunter Wolf, Gunther Zahner, Friedrich Thaiss, Udo Helmchen, Rüdiger Reinking, Petra Barth, Mariola Reszka, Rolf A.k. StahlAbstract:Monocyte chemoattractant protein-1 and osteopontin differentially regulate monocytes recruitment in Experimental Glomerulonephritis. Background This study evaluated the mechanisms of monocyte/macrophage (M/M) infiltration in a rat model of anti-glomerular basement membrane Glomerulonephritis (GN). We focused on chemokines and osteopontin, which are known regulators of M/M recruitment. Methods Using immunohistology, in situ hybridization, and Northern blotting, the expression levels of chemokines and osteopontin were evaluated in isolated glomeruli and tubules 4, 10, and 20 days after the induction of GN. In vivo blocking experiments were performed by application of neutralizing antibodies against osteopontin and monocyte chemoattractant protein-1 (MCP-1). Results In nephritic animals, high glomerular MCP-1 and RANTES (regulated upon activation normal T cell expressed and secreted) expression levels were observed on days 4 and 10. The tubular expression of MCP-1, however, was only slightly enhanced. In contrast, tubular osteopontin production was maximally stimulated (day 10) and paralleled with peaks of albuminuria and tubulointerstitial M/M infiltration. Application of an anti-osteopontin antibody ameliorated tubulointerstitial and glomerular M/M recruitment, whereas treatment with an anti-MCP-1 antibody selectively reduced glomerular M/M recruitment. However, tubulointerstitial M/M infiltration remained unchanged. Conclusion These studies show that chemokines and osteopontin are differentially expressed in glomeruli and tubules in this model of GN. Chemokines play a primary role in the glomeruli, whereas osteopontin has a predominant role in tubulointerstitial M/M recruitment. The roles of chemokines and osteopontin may thus be dependent on the renal compartment and on the disease model.
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The chemokine receptor antagonist AOP-RANTES reduces monocyte infiltration in Experimental Glomerulonephritis
Kidney international, 1999Co-Authors: Ulf Panzer, André Schneider, Jill Wilken, Darren A. Thompson, Stephen B. H. Kent, Rolf A.k. StahlAbstract:The chemokine receptor antagonist AOP-RANTES reduces monocyte infiltration in Experimental Glomerulonephritis. Background This study was designed to evaluate the role of the novel chemokine receptor antagonist amino-oxypentane RANTES (AOP-RANTES), which blocks the binding of macrophage inflammatory protein-1α (MIP-1α), MIP-1β, and RANTES to the chemokine receptor-5 (CCR-5) on the infiltration of monocytes in Experimental Glomerulonephritis. Methods Rats were treated twice daily with 12.5 μg AOP-RANTES following an induction of anti–rat-thymocyte antibody-mediated Glomerulonephritis. The white blood cell count, glomerular monocyte infiltration, chemokine expression, and collagen type IV deposition were assessed. Results The induction of Glomerulonephritis increased glomerular monocyte/macrophage (M/M) infiltration at 24 hours and at 5 days was still higher than in controls. AOP-RANTES prevented glomerular M/M infiltration at 24 hours and at 5 days. This was paralleled by reduced glomerular collagen type IV deposition as a fibrotic marker in nephritic animals. Conclusion These data show that the CCR-5 chemokine receptor antagonist AOP-RANTES ameliorates M/M infiltration and improves glomerular pathology in Experimental Glomerulonephritis. The use of chemokine receptor antagonists may offer a new therapeutic option in inflammatory renal injuries.
Masatsugu Hori - One of the best experts on this subject based on the ideXlab platform.
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DNAzyme for TGF-β suppressed extracellular matrix accumulation in Experimental Glomerulonephritis
Kidney international, 2004Co-Authors: Yoshitaka Isaka, Enyu Imai, Hiroyuki Nakamura, Masayuki Mizui, Yoshitsugu Takabatake, Masaru Horio, Hiroshi Kawachi, Fujio Shimizu, Masatsugu HoriAbstract:DNAzyme for TGF-β suppressed extracellular matrix accumulation in Experimental Glomerulonephritis. Background We developed an electroporation-mediated gene transfer method targeting glomerular mesangial cells. Injecting DNA solution via renal artery followed by electric pulses using tweezers-type electrodes could result in efficient transfection in mesangial cells. Therefore, this gene transfer system opened a feasible strategy to manipulate the function of several cytokines and growth factors in mesangial cells. Recently, a new generation of catalytic nucleic acid composed of DNA, named DNA enzyme (DNAzyme), has been developed. Method We generated a DNAzyme (TGFDE) targeting transforming growth factor-β1 (TGF-β1), and examined the therapeutic effect of TGFDE in vitro and in vivo. Results In cultured rat mesangial cells, treatment with TGFDE blocked TGF-β1 mRNA expression, and thereby suppressed type I collagen mRNA expression. Next, we introduced TGFDE or scrambled DNAzyme (TGFSCR) into anti-Thy-1 model of nephritic rats by electroporation 3 days after disease induction. Northern blot analysis and immunohistochemical staining demonstrated that glomerular message and protein expression of TGF-β1, α-smooth muscle actin (α-SMA), and type I collagen were suppressed in TGFDE-transfected nephritic rats compared with untreated nephritic rats and TGFSCR-transfected rats on day 7. Consequently, we observed significant reduction in glomerular matrix score in TGFDE-transfected nephritic rats. Conclusion Inhibition of TGF-β1 expression by electroporation-mediated DNAzyme transfer might be useful for the therapy of Glomerulonephritis.
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Electroporation-mediated PDGF receptor-IgG chimera gene transfer ameliorates Experimental Glomerulonephritis.
Kidney international, 2001Co-Authors: Hiroyuki Nakamura, Yoshitaka Akagi, Yoshitaka Isaka, Enyu Imai, Michiko Tsujie, Tetsuo Sudo, Noriko Ohno, Masatsugu HoriAbstract:Electroporation-mediated PDGF receptor-IgG chimera gene transfer ameliorates Experimental Glomerulonephritis. Background Mesangial cell proliferation and phenotypic alteration occur in an early phase of glomerular injury and precede increased extracellular matrix accumulation. A critical growth factor responsible for mesangial proliferation is platelet-derived growth factor (PDGF), which has proved to be a potent mitogen. Methods We generated a chimeric cDNA encoding an extracellular domain of the β-PDGF receptor fused with IgG-Fc, termed PDGFR/Fc, and examined the feasibility of gene therapy targeting PDGF using PDGFR/Fc. Results Chimeric PDGFR/Fc molecule completely inhibited the tyrosine phosphorylation of β-PDGF receptors and cellular proliferation induced by PDGF in vitro. We then introduced the PDGFR/Fc expression vector into the muscle of anti–Thy-1 model of glomerulonephritic rats by electroporation. The plasma concentration of chimeric PDGFR/Fc levels was 244.4 ± 89.8 ng/mL four days after transfection. On day 5, PDGFR / Fc gene transfer significantly reduced the number of PCNA-positive cells and glomerular cell numbers by 59.6 and 23.2%, respectively. Northern blot analysis demonstrated that glomerular mRNA levels of α-smooth muscle action, transforming growth factor-β1, and type I collagen were also suppressed on days 5 and 7 by the PDGFR / Fc transfection. There was a significant reduction in the matrix score of the transfected nephritic rats (2.91 ± 0.75 and 2.06 ± 0.95; disease control group vs. treated group, P Conclusion These results suggest that gene therapy by the manipulation of PDGF action using electroporation-mediated PDGFR / Fc gene transfer to the skeletal muscle might be a useful treatment for mesangioproliferative Glomerulonephritis.
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Gene therapy by transforming growth factor-β receptor-IgG Fc chimera suppressed extracellular matrix accumulation in Experimental Glomerulonephritis
Kidney international, 1999Co-Authors: Yoshitaka Isaka, Yoshitaka Akagi, Yasufumi Kaneda, Wayne A. Border, Michiko Tsujie, Tetsuo Sudo, Noriko Ohno, Nancy A. Noble, Yutaka Ando, Masatsugu HoriAbstract:Gene therapy by transforming growth factor-β receptor-IgG Fc chimera suppressed extracellular matrix accumulation in Experimental Glomerulonephritis. Background The evidence that transforming growth factor-β (TGF-β) is a key mediator in the pathogenesis of fibrotic diseases is now supported by several lines of investigation. This evidence provides a certain base for targeting TGF-β as an antifibrotic agent. Methods We generated a chimeric cDNA, termed TGFβRII/Fc, encoding an extracellular domain of the TGF-β type II receptor fused to the IgG-Fc domain, and tested whether TGFβRII/Fc could be a novel strategy for treating glomerular diseases. Results In cultured BNul-7 cells, recombinant TGFβRII/Fc reversed the antiproliferative response induced by TGF-β1. In addition, TGFβRII/Fc diminished the TGF-β1–induced production of EIIIA-positive fibronectin in cultured normal rat kidney cells. We then introduced the chimeric cDNA into the muscle of the nephritic rats by the hemagglutinating virus of Japan liposome–mediated gene transfer method in order to block the TGF-β activity in nephritic glomeruli through systemic delivery of chimeric molecules. Treatment with TGFβRII/Fc gene transfection could suppress the glomerular TGF-β mRNA in nephritic rats with a comparable effect in the reduction of extracellular matrix accumulation. Conclusion GFβRII/Fc successfully inhibited the action of TGF-β in vitro and in vivo , and gene therapy by chimeric TGFβRII/Fc might be feasible for the therapy of glomerulosclerosis.
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Gene transfer and kidney disease.
Journal of Nephrology, 1998Co-Authors: Enyu Imai, Yoshitaka Akagi, Yoshitaka Isaka, Yasufumi Kaneda, Masatsugu HoriAbstract:There is little doubt that molecular biological intervention therapy has come of age and its potential is arousing tremendous excitement. A gene transfer technique, the HVJ-liposome method, is now applicable as a tool for the dissection of molecular aspects in the pathophysiology of renal diseases, and for gene therapy in Experimental Glomerulonephritis. Overexpressed transforming growth factor (TGF)-beta in the normal rat glomeruli, by gene transfer of TGF-beta cDNA, leads to glomerulosclerosis. However, inhibition of the TGF-beta action by antisense oligonucleotides can suppress the development of the Experimental Glomerulonephritis. We investigated whether in vivo gene transfer of chimeric proteins, composed of the extracellular domain of TGF-beta type II receptor fused with IgC-Fc, suppresses Experimental Glomerulonephritis. The expression of TGF-beta in glomeruli was suppressed and so was the extracellular matrix expansion. Taken together with clinical observation of up-regulation of TGF-beta in various glomerulopathies, the dys-regulation of the TGF-beta is important in the development of glomerulosclerosis, and manipulation of this overexpression may prove a novel therapeutic approach for slowing the progression of the disease.
William G. Couser - One of the best experts on this subject based on the ideXlab platform.
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the cyclin kinase inhibitor p21cip1 waf1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis
Kidney International, 1999Co-Authors: Yoon-goo Kim, Charles E. Alpers, James B Brugarolas, Richard J. Johnson, William G. CouserAbstract:The cyclin kinase inhibitor p21 CIP1/WAF1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis. Background During glomerulogenesis, visceral glomerular epithelial cells (VECs) exit the cell cycle and become terminally differentiated and quiescent. In contrast to other resident glomerular cells, VECs undergo little if any proliferation in response to injury. However, the mechanisms for this remain unclear. Cell proliferation is controlled by cell-cycle regulatory proteins where the cyclin-dependent kinase inhibitor p21 Cip1,WAF1 (p21) inhibits cell proliferation and is required for differentiation of many nonrenal cell types. Methods To test the hypothesis that p21 is required to maintain a differentiated and quiescent VEC phenotype, Experimental Glomerulonephritis was induced in p21 knockout (-/-) and p21 wild-type (+/+) mice with antiglomerular antibody. DNA synthesis (proliferating cell nuclear antigen, bromodeoxyuridine staining), VEC proliferation (multilayers of cells in Bowman's space), matrix accumulation (periodic acid-Schiff, silver staining), apoptosis (TUNEL), and renal function (serum urea nitrogen) were studied on days 5 and 14 ( N = 6 per time point). VECs were identified by location, morphology, ezrin staining, and electron microscopy. VEC differentiation was measured by staining for Wilms' tumor-1 gene. Results Kidneys from unmanipulated p21-/- mice were histologically normal and did not have increased DNA synthesis, suggesting that p21 was not required for the induction of VEC terminal differentiation. Proliferating cell nuclear antigen and bromodeoxyuridine staining was increased 4.3- and 3.3-fold, respectively, in p21-/- mice with Glomerulonephritis ( P P P Conclusion These results show that the cyclin kinase inhibitor p21 is not required by VECs to attain a terminally differentiated VEC phenotype. However, the loss of p21, in disease states, is associated with VEC re-entry into the cell cycle and the development of a dedifferentiated proliferative phenotype.
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The cyclin kinase inhibitor p21CIP1/WAF1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis.
Kidney international, 1999Co-Authors: Yoon-goo Kim, Charles E. Alpers, James B Brugarolas, Richard J. Johnson, William G. Couser, Stuart J. ShanklandAbstract:The cyclin kinase inhibitor p21 CIP1/WAF1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis. Background During glomerulogenesis, visceral glomerular epithelial cells (VECs) exit the cell cycle and become terminally differentiated and quiescent. In contrast to other resident glomerular cells, VECs undergo little if any proliferation in response to injury. However, the mechanisms for this remain unclear. Cell proliferation is controlled by cell-cycle regulatory proteins where the cyclin-dependent kinase inhibitor p21 Cip1,WAF1 (p21) inhibits cell proliferation and is required for differentiation of many nonrenal cell types. Methods To test the hypothesis that p21 is required to maintain a differentiated and quiescent VEC phenotype, Experimental Glomerulonephritis was induced in p21 knockout (-/-) and p21 wild-type (+/+) mice with antiglomerular antibody. DNA synthesis (proliferating cell nuclear antigen, bromodeoxyuridine staining), VEC proliferation (multilayers of cells in Bowman's space), matrix accumulation (periodic acid-Schiff, silver staining), apoptosis (TUNEL), and renal function (serum urea nitrogen) were studied on days 5 and 14 ( N = 6 per time point). VECs were identified by location, morphology, ezrin staining, and electron microscopy. VEC differentiation was measured by staining for Wilms' tumor-1 gene. Results Kidneys from unmanipulated p21-/- mice were histologically normal and did not have increased DNA synthesis, suggesting that p21 was not required for the induction of VEC terminal differentiation. Proliferating cell nuclear antigen and bromodeoxyuridine staining was increased 4.3- and 3.3-fold, respectively, in p21-/- mice with Glomerulonephritis ( P P P Conclusion These results show that the cyclin kinase inhibitor p21 is not required by VECs to attain a terminally differentiated VEC phenotype. However, the loss of p21, in disease states, is associated with VEC re-entry into the cell cycle and the development of a dedifferentiated proliferative phenotype.
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Molecular mechanisms of Experimental Glomerulonephritis: an overview
Nephrology, 1997Co-Authors: William G. Couser, Stuart J. Shankland, Masaomi Nangaku, Richard J. JohnsonAbstract:Summary: Most glomerular diseases are autoimmune in nature with injury occurring as a consequence of antibody reacting with antigens on glomerular cell membranes or in glomerular matrix, or immune complex formation occurring on or near glomerular cell membrnes. Many of the structural and functional consequences of these events result from the response of resident glomerular cells to the injury, or may be induced by inflammatory effector cells derived from the circulation. In terms of glomerular cells, injury to the glomerular epithelial cell (GEC) may be induced by: (i) non-complement fixing antibodies which mimic the glomerular lesions of minimal change nephrotic syndrome/focal glomerular sclerosis; or (ii) by antibodies which fix complement leading to GEC attack by C5b-9 resulting in a lesion analogous to membranous nephropathy. C5b-9 also mediates antibody induced injury to the mesangial cell resulting in a mesangioproliferative Glomerulonephritis (IgA nephropathy, systemic lupus erythematosus; SLE) as well as to the glomerular endothelial cell (thrombotic microangiopathy, haemolytic uraemic syndrome). the effects of C5b-9 may be lytic (mesangial cell, glomerular endothelial cell; GEN) or sublytic (GEC) resulting in stimulation of local oxident and protease production. Both lytic and sublytic effects are substantially modulated by cell-bound complement regulatory proteins such as CD59 and Crry. When fixed or planted antigens are present in larger quantities and accessible to the circulation, complement activation generates chemotactic factors leading to neutrophil infiltration and producing injury through the myeloperoxidase (MPO)-H2-O2-halide system, a mechanism substantially augmented by platelet-neutrophil interaction (post-infectious Glomerulonephritis [GN], SLE). Macrophages may also be localized in glomeruli by either immune adherence mechanisms or as a consequence of cell mediated immune reactions in the glomerulus. Macrophages differ from neutrophils in producing large amounts of transforming growth factor-β (TGF-β) and procoagulants which contribute to crescent formation rapidly progressive Glomerulonephritis (RPGN).
Richard J. Johnson - One of the best experts on this subject based on the ideXlab platform.
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the cyclin kinase inhibitor p21cip1 waf1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis
Kidney International, 1999Co-Authors: Yoon-goo Kim, Charles E. Alpers, James B Brugarolas, Richard J. Johnson, William G. CouserAbstract:The cyclin kinase inhibitor p21 CIP1/WAF1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis. Background During glomerulogenesis, visceral glomerular epithelial cells (VECs) exit the cell cycle and become terminally differentiated and quiescent. In contrast to other resident glomerular cells, VECs undergo little if any proliferation in response to injury. However, the mechanisms for this remain unclear. Cell proliferation is controlled by cell-cycle regulatory proteins where the cyclin-dependent kinase inhibitor p21 Cip1,WAF1 (p21) inhibits cell proliferation and is required for differentiation of many nonrenal cell types. Methods To test the hypothesis that p21 is required to maintain a differentiated and quiescent VEC phenotype, Experimental Glomerulonephritis was induced in p21 knockout (-/-) and p21 wild-type (+/+) mice with antiglomerular antibody. DNA synthesis (proliferating cell nuclear antigen, bromodeoxyuridine staining), VEC proliferation (multilayers of cells in Bowman's space), matrix accumulation (periodic acid-Schiff, silver staining), apoptosis (TUNEL), and renal function (serum urea nitrogen) were studied on days 5 and 14 ( N = 6 per time point). VECs were identified by location, morphology, ezrin staining, and electron microscopy. VEC differentiation was measured by staining for Wilms' tumor-1 gene. Results Kidneys from unmanipulated p21-/- mice were histologically normal and did not have increased DNA synthesis, suggesting that p21 was not required for the induction of VEC terminal differentiation. Proliferating cell nuclear antigen and bromodeoxyuridine staining was increased 4.3- and 3.3-fold, respectively, in p21-/- mice with Glomerulonephritis ( P P P Conclusion These results show that the cyclin kinase inhibitor p21 is not required by VECs to attain a terminally differentiated VEC phenotype. However, the loss of p21, in disease states, is associated with VEC re-entry into the cell cycle and the development of a dedifferentiated proliferative phenotype.
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The cyclin kinase inhibitor p21CIP1/WAF1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis.
Kidney international, 1999Co-Authors: Yoon-goo Kim, Charles E. Alpers, James B Brugarolas, Richard J. Johnson, William G. Couser, Stuart J. ShanklandAbstract:The cyclin kinase inhibitor p21 CIP1/WAF1 limits glomerular epithelial cell proliferation in Experimental Glomerulonephritis. Background During glomerulogenesis, visceral glomerular epithelial cells (VECs) exit the cell cycle and become terminally differentiated and quiescent. In contrast to other resident glomerular cells, VECs undergo little if any proliferation in response to injury. However, the mechanisms for this remain unclear. Cell proliferation is controlled by cell-cycle regulatory proteins where the cyclin-dependent kinase inhibitor p21 Cip1,WAF1 (p21) inhibits cell proliferation and is required for differentiation of many nonrenal cell types. Methods To test the hypothesis that p21 is required to maintain a differentiated and quiescent VEC phenotype, Experimental Glomerulonephritis was induced in p21 knockout (-/-) and p21 wild-type (+/+) mice with antiglomerular antibody. DNA synthesis (proliferating cell nuclear antigen, bromodeoxyuridine staining), VEC proliferation (multilayers of cells in Bowman's space), matrix accumulation (periodic acid-Schiff, silver staining), apoptosis (TUNEL), and renal function (serum urea nitrogen) were studied on days 5 and 14 ( N = 6 per time point). VECs were identified by location, morphology, ezrin staining, and electron microscopy. VEC differentiation was measured by staining for Wilms' tumor-1 gene. Results Kidneys from unmanipulated p21-/- mice were histologically normal and did not have increased DNA synthesis, suggesting that p21 was not required for the induction of VEC terminal differentiation. Proliferating cell nuclear antigen and bromodeoxyuridine staining was increased 4.3- and 3.3-fold, respectively, in p21-/- mice with Glomerulonephritis ( P P P Conclusion These results show that the cyclin kinase inhibitor p21 is not required by VECs to attain a terminally differentiated VEC phenotype. However, the loss of p21, in disease states, is associated with VEC re-entry into the cell cycle and the development of a dedifferentiated proliferative phenotype.
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Dissociation of mesangial cell migration and proliferation in Experimental Glomerulonephritis
Kidney international, 1999Co-Authors: Leah Haseley, Christian Hugo, Michael A. Reidy, Richard J. JohnsonAbstract:Dissociation of mesangial cell migration and proliferation in Experimental Glomerulonephritis. Background Recently, we documented that following in vivo mesangial cell (MC) ablation in the Thy1 model, reconstitution of the mesangium occurs by a coordinated proliferation and migration of Thy1 (OX-7)-positive cells originating from the hilus and extraglomerular mesangium. We investigated the role of basic fibroblast growth factor (bFGF) in the mediation of these events. Methods Rats were injected with antithymocyte serum and 48 hours later were pulsed with 3 H-thymidine to label proliferating cells. Ninety minutes later, a baseline renal biopsy was obtained, and rats were injected with neutralizing anti-bFGF antibodies or control IgG. Sacrificial biopsies were obtained at 96 hours of disease. Using computer image analysis, biopsies from both time points were quantitated for the number of radiolabeled MC (proliferation) and their mean distance from the hilus (migration). The effect of bFGF on the migration of MCs in culture was examined using a chemotactic assay. Results At sacrifice, autoradiographs of rats receiving anti-bFGF had significantly fewer radiolabeled MCs as compared with rats receiving control IgG (8.7 ± 1.9 vs. 14.7 ± 3.5, P = 0.0001), yielding an overall 40% reduction in proliferation. There was no difference, however, in the final distance of radiolabeled MCs from the glomerular hilus in the two groups, indicating that the administration of anti-bFGF did not effect MC migration in this model. In an in vitro chemotactic assay, MCs migrated in response to platelet-derived growth factor (PDGF) BB (20 ng/ml), but did not migrate in response to bFGF at a wide range of concentrations (0.5 to 50 ng/ml). Conclusions These studies demonstrate that bFGF is an important mediator of MC proliferation but that it does not significantly influence MC migration. This is the first demonstration showing that the mediators effecting proliferation can be dissociated from those mediating migration in renal injury.
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Molecular mechanisms of Experimental Glomerulonephritis: an overview
Nephrology, 1997Co-Authors: William G. Couser, Stuart J. Shankland, Masaomi Nangaku, Richard J. JohnsonAbstract:Summary: Most glomerular diseases are autoimmune in nature with injury occurring as a consequence of antibody reacting with antigens on glomerular cell membranes or in glomerular matrix, or immune complex formation occurring on or near glomerular cell membrnes. Many of the structural and functional consequences of these events result from the response of resident glomerular cells to the injury, or may be induced by inflammatory effector cells derived from the circulation. In terms of glomerular cells, injury to the glomerular epithelial cell (GEC) may be induced by: (i) non-complement fixing antibodies which mimic the glomerular lesions of minimal change nephrotic syndrome/focal glomerular sclerosis; or (ii) by antibodies which fix complement leading to GEC attack by C5b-9 resulting in a lesion analogous to membranous nephropathy. C5b-9 also mediates antibody induced injury to the mesangial cell resulting in a mesangioproliferative Glomerulonephritis (IgA nephropathy, systemic lupus erythematosus; SLE) as well as to the glomerular endothelial cell (thrombotic microangiopathy, haemolytic uraemic syndrome). the effects of C5b-9 may be lytic (mesangial cell, glomerular endothelial cell; GEN) or sublytic (GEC) resulting in stimulation of local oxident and protease production. Both lytic and sublytic effects are substantially modulated by cell-bound complement regulatory proteins such as CD59 and Crry. When fixed or planted antigens are present in larger quantities and accessible to the circulation, complement activation generates chemotactic factors leading to neutrophil infiltration and producing injury through the myeloperoxidase (MPO)-H2-O2-halide system, a mechanism substantially augmented by platelet-neutrophil interaction (post-infectious Glomerulonephritis [GN], SLE). Macrophages may also be localized in glomeruli by either immune adherence mechanisms or as a consequence of cell mediated immune reactions in the glomerulus. Macrophages differ from neutrophils in producing large amounts of transforming growth factor-β (TGF-β) and procoagulants which contribute to crescent formation rapidly progressive Glomerulonephritis (RPGN).
Nancy A. Noble - One of the best experts on this subject based on the ideXlab platform.
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A mutant, noninhibitory plasminogen activator inhibitor type 1 decreases matrix accumulation in Experimental Glomerulonephritis
The Journal of clinical investigation, 2003Co-Authors: Yufeng Huang, Wayne A. Border, Masashi Haraguchi, Daniel A. Lawrence, Nancy A. NobleAbstract:In fibrotic renal disease, elevated TGF-β and angiotensin II lead to increased plasminogen activator inhibitor type 1 (PAI-1). PAI-1 appears to reduce glomerular mesangial matrix turnover by inhibiting plasminogen activators, thereby decreasing plasmin generation and plasmin-mediated matrix degradation. We hypothesized that therapy with a mutant human PAI-1 (PAI-1R) that binds to matrix vitronectin but does not inhibit plasminogen activators, would enhance plasmin generation, increase matrix turnover, and decrease matrix accumulation in Experimental Glomerulonephritis. Three Experimental groups included normal, untreated disease control, and PAI-1R–treated nephritic rats. Plasmin generation by isolated day 3 glomeruli was dramatically decreased by 69%, a decrease that was reversed 43% (P < 0.02) by in vivo PAI-1R treatment. At day 6, animals treated with PAI-1R showed significant reductions in proteinuria (48%, P < 0.02), glomerular staining for periodic acid–Schiff positive material (33%, P < 0.02), collagen I (28%, P < 0.01), collagen III (34%, P < 0.01), fibronectin (48%, P < 0.01), and laminin (41%, P < 0.01), and in collagen I (P < 0.01) and fibronectin mRNA levels (P < 0.02). Treatment did not alter overexpression of TGF-β1 and PAI-1 mRNAs, although TGF-β1 protein was significantly reduced. These observations strongly support our hypothesis that PAI-1R reduces glomerulosclerosis by competing with endogenous PAI-1, restoring plasmin generation, inhibiting inflammatory cell infiltration, decreasing local TGF-β1 concentration, and reducing matrix accumulation.
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t-PA promotes glomerular plasmin generation and matrix degradation in Experimental Glomerulonephritis.
Kidney international, 2001Co-Authors: Masashi Haraguchi, Wayne A. Border, Yufeng Huang, Nancy A. NobleAbstract:t-PA promotes glomerular plasmin generation and matrix degradation in Experimental Glomerulonephritis. Background In addition to its well-known role in degrading fibrin, recent evidence suggests that plasmin degrades matrix proteins and activates prometalloproteinases. Plasmin is generated from plasminogen by tissue plasminogen activator (t-PA). We hypothesized that t-PA treatment increases plasmin generation in nephritic glomeruli and degrades pathological matrix leading to a therapeutic reduction in matrix accumulation. Methods Anti–Thy-1 nephritis was induced by injection of OX-7 antibody. Rats were given twice daily intravenous injections of saline (disease control group) or human recombinant t-PA (rt-PA; 1 mg/kg body weight) on days 3 through 5. Proteinuria, glomerular matrix protein staining, and glomerular mRNA levels for transforming growth factor-β1 (TGF-β1), fibronectin, and plasminogen activator inhibitor type 1 (PAI-1) were evaluated at day 6. Localization of rt-PA, plasmin generation by glomeruli in vitro, and glomerular production and content of active TGF-β1 were also investigated. Results Compared with disease control animals, proteinuria and staining score for periodic acid-Schiff (2.75 ± 0.17 vs. 1.41 ± 0.09), fibronectin-EDA+ (19 ± 2 vs. 14 ± 1), laminin (35 ± 2 vs. 25 ± 2), type I collagen (33 ± 1 vs. 21 ± 3), and type IV collagen (27 ± 2 vs. 23 ± 1) were significantly reduced in treated rats ( P Conclusions These results show that injected rt-PA binds to fibrin in nephritic glomeruli, thus increasing plasmin generation and promoting pathological matrix degradation without activating latent TGF-β. Agents that increase plasmin generation, such as t-PA, may have potential as antifibrotic therapies.
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Angiotensin II blockade and low-protein diet produce additive therapeutic effects in Experimental Glomerulonephritis.
Kidney international, 2000Co-Authors: Harm Peters, Wayne A. Border, Nancy A. NobleAbstract:Angiotensin II blockade and low-protein diet produce additive therapeutic effects in Experimental Glomerulonephritis. Background Transforming growth factor-β (TGF-β) overexpression plays a key role in the accumulation of extracellular matrix in acute and chronic renal diseases. Recent studies have suggested that the degree of reduction in pathological TGF-β overexpression can be used as a therapeutic index to evaluate the antifibrotic potential of pharmacological angiotensin II (Ang II) blockade in renal disease. Using this target, we found that treatment with the angiotensin I-converting enzyme inhibitor enalapril or the Ang II type 1 receptor antagonist losartan reduced TGF-β overexpression more effectively at doses clearly higher than those required to control blood pressure. However, both forms of Ang II blockade were only partially effective in normalizing TGF-β expression. This study investigated whether a greater antifibrotic, TGF-β–reducing benefit can be achieved when Ang II blockade is combined with dietary protein restriction. Methods Mesangioproliferative Glomerulonephritis was induced in male Sprague-Dawley rats on a normal-protein diet. Treatment with a low-protein diet and/or maximally effective doses of enalapril or losartan was started one day after disease induction. On the fifth day, 24-hour urine protein excretion was measured. On the sixth day, cortical kidney tissue was taken for periodic acid-Schiff staining. Isolated glomeruli were used for mRNA extraction or were placed in culture for determination of production of TGF-β1, the matrix protein fibronectin, and the protease inhibitor plasmin activator inhibitor type 1 (PAI-1) by enzyme-linked immunosorbent assay. Results Compared with untreated nephritic animals on a normal-protein diet, a single treatment with enalapril, losartan, or low-protein diet significantly reduced glomerular TGF-β production, albeit to a similar degree of approximately 45%. A moderate, but significant further reduction in pathological TGF-β expression of a total of 65% for enalapril and 60% for losartan was achieved when these drugs were combined with low-protein feeding. This reduction in TGF-β overexpression paralleled decreased proteinuria, glomerular matrix accumulation, and overproduction of fibronectin and PAI-1. Conclusions Ang II blockade and low-protein diet have additive effects on disease reduction, suggesting that disease progression in humans with chronic renal failure may be slowed more effectively when Ang II blockade and low-protein diet are combined. Since maximal pharmacological Ang II inhibition was used, it is likely that dietary protein restriction further reduces pathological TGF-β overexpression by mechanisms different from those of enalapril or losartan.
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Tandem antifibrotic actions of L-arginine supplementation and low protein diet during the repair phase of Experimental Glomerulonephritis
Kidney international, 2000Co-Authors: Harm Peters, Wayne A. Border, Nancy A. NobleAbstract:Tandem antifibrotic actions of L-arginine supplementation and low protein diet during the repair phase of Experimental Glomerulonephritis. Background Based upon the central role transforming growth factor-beta (TGF-β) overexpression appears to play in renal fibrotic diseases, we have recently advocated reduction of TGF-β as a therapeutic target. As part of efforts to determine the strength of this approach, we have undertaken studies to quantitate the effects of currently used and promising therapies in terms of their potential to reduce markers of disease in anti-thymocyte-serum (ATS)-Glomerulonephritis in the rat. Here we assess the therapeutic effect of L-arginine supplementation, which has been shown to reduce fibrosis in a number of hypertensive models, given alone or in combination with low protein diet and started 24 hours after disease induction. Methods Glomerulonephritis was induced by intravenous injection of OX-7 monoclonal antibody into 200g Sprague-Dawley rats. Twenty-four hours later animals were placed in groups that were either untreated, treated with 1% L-arginine in drinking water or 6% protein diets or both. On the fifth day of disease 24-hour urine specimens were collected and systemic blood pressure was measured. On the sixth day rats were anesthetized. Kidneys were perfused, tissue was taken for PAS staining and glomeruli were isolated. Aliquots of glomeruli were used for RNA preparation and for culture to determine 72-hour production of TGF-β, fibronectin and plasminogen activator-type 1 (PAI-1), which were assayed by ELISA on culture supernatants. Measures of nitrate and nitrite (NOx) production included plasma NOx, urinary NOx and glomerular production of NOx in culture. Results All disease measures except proteinuria and including matrix accumulation, TGF-β, fibronectin and PAI-1 production and mRNA expression for TGF-β, fibronectin and PAI-1 were significantly and similarly reduced by about 50% in groups treated with L-arginine or with low protein diet. Proteinuria was reduced in low protein treated but not in L-arginine supplemented rats. Neither systemic blood pressure nor measures of NO synthesis showed differences between groups that could be attributed to L-arginine supplementation. In contrast, disease-related increases in glomerular production of NOx were markedly reduced by low protein. Combined therapy resulted in small, but statistically significant decreases in most measures of disease. Conclusions L-arginine supplementation reduces fibrotic disease in ATS-induced Glomerulonephritis if started after disease induction. The absence of evidence for increased NO production related to L-arginine supplementation suggests that L-arginine is acting here through different pathways from those demonstrated in hypertensive models of disease. The data support the ideas that TGF-β reduction is a valid therapeutic target and that quantitation of TGF-β reduction is a useful approach for comparing antifibrotic drug candidates.
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L-Arginine supplementation increases mesangial cell injury and subsequent tissue fibrosis in Experimental Glomerulonephritis.
Kidney international, 1999Co-Authors: Harm Peters, Wayne A. Border, Nancy A. NobleAbstract:l-Arginine supplementation increases mesangial cell injury and subsequent tissue fibrosis in Experimental Glomerulonephritis. Background Mesangial cell lysis in the antithymocyte serum (ATS)-induced model of Glomerulonephritis is dependent on the generation of cytotoxic nitric oxide (NO) through transient induction of NO synthase (iNOS). We hypothesized that increased availability of l-arginine (L-Arg) during mesangial cell lysis might provide iNOS with increased substrate leading to increased lysis, and that this increased lysis would be reflected in more severe fibrotic disease at day 6. Methods mRNA and placed in culture to study the dependence of NO production on L-Arg concentration. In rats sacrificed at six days, L-Arg supplementation was stopped 16hours after ATS injection. Fibrotic disease was evaluated by urinary protein excretion, histological assessment of glomerular cell number, matrix accumulation, and production of transforming growth factor-β1 and matrix components fibronectin and plasminogen activator inhibitor type-1 (PAI-1) by isolated glomeruli in culture. Results At six hours, the glomerular cell number was significantly reduced by ATS injection ( P P P P P P P P P Conclusions The results indicate that if given during disease induction, L-Arg supplementation can enhance iNOS-dependent tissue injury by providing increased substrate. Although the increase in injury with L-Arg supplementation was small, it led to increased fibrosis at day 6. These data predict that in diseases with repeated iNOS-dependent tissue injury, L-Arg supplementation may produce cumulative increases in tissue fibrosis.
