Masayuki Iyoda

TLR4 Links Podocytes with the Innate Immune System to Mediate Glomerular Injury

Toll-like receptors (TLR) classically recognize pathogen-associated danger signals but are also activated via endogenous ligands. For evaluation of their role in inflammatory kidney disease, the function of TLR was analyzed in two mouse models of cryoglobulinemic membranoproliferative glomerulonephritis (MPGN; mice transgenic for thymic stromal lymphopoietin [TSLP], with or without deletion of the Fcgamma receptor IIb). Expression of TLR1 through 9 and TLR11 mRNA was detectable in whole kidneys and in isolated glomeruli of wild-type mice, with TLR3 and TLR4 having the highest absolute levels of expression. TLR1, 2, and 4 were increased in TSLP transgenic mice and even higher in TSLP transgenic FcgammaRIIb-deficient mice. TLR5 through 9 and 11 were upregulated to similar degrees in TSLP transgenic and TSLP transgenic FcgammaRIIb-deficient mice. Immunohistochemical studies of nephritic glomeruli localized TLR4 protein to podocytes. Cultured podocytes also expressed TLR4, and stimulation with TLR4-specific ligands resulted in a marked induction of chemokines; this was reduced by specific knockdown of TLR4 with siRNA. Fibrinogen, a potential endogenous TLR4 ligand, was shown to induce a similar profile of chemokines. In conclusion, it was demonstrated that TLR4 is constitutively expressed by podocytes and is upregulated in MPGN, where it may mediate glomerular injury by modulating expression of chemokines; therefore, TLR4 may link podocytes with the innate immune system to mediate MPGN triggered by the deposition of immune complexes.

IL-17A and IL-17F stimulate chemokines via MAPK pathways (ERK1/2 and p38 but not JNK) in mouse cultured mesangial cells: synergy with TNF-α and IL-1β

We investigated the role of IL-17 family members IL-17A and IL-17F in the induction of chemokines in mouse cultured mesangial cells (SV40 MES 13 cells). We evaluated the expression of the chemokines monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-2 (MIP-2) by ELISA and real-time RT-PCR (Q-PCR). Activation of MAPK was assessed by immunoblotting. IL-17RA and IL-17RC were inhibited by small interfering RNA (siRNA). We found that IL-17A or IL-17F stimulation of mesangial cells led to both a dose- and time-dependent increase in MCP-1 and MIP-2 release. This effect was dependent on mRNA transcription and protein translation. Both also enhanced TNF-alpha- and IL-1beta-mediated MCP-1 and MIP-2 release in the cells. Additionally, we observed that IL-17A and IL-17F induced MAPK (p38 MAPK, ERK1/2, and JNK) activation and that pharmacological inhibitors of p38 MAPK (SB203580) and ERK1/2 (U0126), but not JNK (SP600125), blocked the IL-17A/IL-17F-mediated MCP-1 and MIP-2 release. Mesangial cells expressed IL-17RA and IL-17RC, and the IL-17A-mediated MCP-1 and MIP-2 release was significantly blocked by soluble IL-17RA. Furthermore, inhibition of either IL-17RA or IL-17RC expression via siRNA led to significant reduction of IL-17A/IL-17F-stimulated chemokine production. We conclude that IL-17A and IL-17F induce the production of chemokines MCP-1 and MIP-2 via MAPK pathways (p38 MAPK and ERK1/2), as well as mRNA transcription and protein translation and have synergistic effects with TNF-alpha and IL-1beta in cultured mesangial cells.

Sildenafil, a phosphodiesterase type 5 inhibitor, attenuates diabetic nephropathy in non-insulin-dependent Otsuka Long-Evans Tokushima Fatty rats

BACKGROUND It is well established that the pathogenesis of diabetic nephropathy is associated with abnormalities of renal nitric oxide (NO) generation. Many of the biological actions of NO are mediated by cGMP, which is rapidly degraded by phosphodiesterases. In this study, we evaluated the renoprotective effects of sildenafil (SIL), an inhibitor of phosphodiesterase‐5, in type 2 diabetic rats.

Nilotinib Attenuates Renal Injury and Prolongs Survival in Chronic Kidney Disease

The tyrosine kinase inhibitor imatinib is beneficial in experimental renal diseases, but the effect of the new tyrosine kinase inhibitor nilotinib on the progression of renal failure is unknown. We administered either nilotinib or vehicle to Sprague-Dawley rats beginning 2 weeks after 5/6 nephrectomy (Nx) or laparotomy and continuing for 8 weeks. Serum creatinine levels were significantly lower in the nilotinib group after 6 and 8 weeks of treatment. Furthermore, nilotinib-treated rats had less proteinuria, attenuated glomerulosclerosis and tubulointerstitial damage, and reduced macrophage infiltration into the tubulointerstitium. Treatment with nilotinib also significantly decreased renal cortical expression of profibrogenic genes, such as IL-1β and monocyte chemotactic protein-1, which correlated closely with the tubulointerstitial damage score and ED1-positive macrophages score. In addition, nilotinib treatment significantly prolonged survival. Taken together, these results suggest that nilotinib may limit the progression of chronic kidney disease.

Preventive and therapeutic effects of imatinib in Wistar-Kyoto rats with anti-glomerular basement membrane glomerulonephritis

Imatinib is a selective tyrosine kinase inhibitor that can block activity of the platelet-derived growth factor receptor (PDGFR) and that has immunomodulatory effects on various cell types. Here we measured the protective effects of imatinib in Wistar-Kyoto rats with nephrotoxic serum nephritis, a kidney disease model where CD8+ T cells and macrophages play pathogenetic roles. Groups of animals were given imatinib from one day before up to 13 days following induction of nephritis and from day 7 to 20 following disease induction. Compared to control rats, at each time point imatinib treatment caused significantly less proteinuria, lowered serum blood urea nitrogen and creatinine, and decreased the number of glomeruli with necrosis, crescents, and fibrin deposits. Imatinib-treated rats had a significant reduction in glomerular macrophage accumulation and reduced renal cortical PDGFR-beta and M-CSF receptor mRNA expression. Using colocalization we found that glomerular macrophages had reduced IL-1beta and MCP-1 protein expression. Late imatinib treatment significantly reduced proteinuria, serum blood urea nitrogen, and creatinine, and reversed renal histopathological changes. We show that imatinib has renoprotective and therapeutic properties and provide pre-clinical work that will need to be confirmed in patients with crescentic glomerulonephritis.

Imatinib Suppresses Cryoglobulinemia and Secondary Membranoproliferative Glomerulonephritis

Imatinib is a receptor tyrosine kinase inhibitor that blocks the activity of c-Abl, c-Kit, and PDGF receptors. We tested the protective effects of imatinib in thymic stromal lymphopoietin transgenic mice, a model of cryoglobulinemia and associated membranoproliferative glomerulonephritis (MPGN), in which some glomerular manifestations likely result from PDGF receptor activation. Surprising, administration of imatinib beginning at weaning suppressed production of cryoglobulin, attenuating both the renal injury and systemic features of cryoglobulinemia. Flow cytometry suggested that inhibition of B cell development in the bone marrow likely caused the reduction in cryoglobulin production. In addition, administration of imatinib to thymic stromal lymphopoietin transgenic mice with established MPGN also diminished cryoglobulin production and reversed the renal and systemic lesions. These data suggest that treatment with imatinib may be a novel therapeutic approach for cryoglobulinemia and MPGN in humans.

Renin-Angiotensin System Blockade Is Renoprotective in Immune Complex–Mediated Glomerulonephritis

Blockade of the renin-angiotensin system is renoprotective in a variety of chronic nephropathies, but the direct effect of such treatment in active, immune complex-mediated glomerulonephritis is unknown. This study investigated the short- and long-term effects of an angiotensin-converting enzyme inhibitor (enalapril) and an angiotensin II type 1 receptor blocker (losartan) in thymic stromal lymphopoietin transgenic (TSLPtg) mice, which develop mixed cryoglobulinemia and severe cryoglobulinemia-associated membranoproliferative glomerulonephritis. Enalapril and losartan each reduced hypertension, proteinuria, glomerular extracellular matrix deposition, and mesangial cell activation in TSLPtg mice. These renoprotective effects were not observed with hydralazine treatment, despite a similar antihypertensive effect. Treatment with enalapril or losartan also decreased renal plasminogen activator inhibitor-1 in TSLPtg mice, assessed by immunohistochemistry and quantitative real-time reverse transcriptase-PCR. None of the treatments affected immune complex deposition or macrophage infiltration. Overall, enalapril- and losartan-treated TSLPtg mice survived significantly longer than untreated TSLPtg mice. These studies demonstrate that angiotensin blockade may provide renoprotective benefits, independent of its BP-lowering effect, in the treatment of active immune complex-mediated glomerulonephritis.

Localization of TGF-β Signaling Intermediates Smad2, 3, 4, and 7 in Developing and Mature Human and Mouse Kidney

Smad proteins are signaling intermediates of the TGF-β superfamily and are involved in a range of biological activities including development and immune responses. We studied the expression of TGF-β-receptor activated Smads (Smad2 and Smad3), the common partner Smad (Smad4), an inhibitory Smad (Smad7), and the activated (phosphorylated) Smad2 (pSmad2) in developing and adult kidneys of humans and mice. These studies demonstrate associated expression of these Smads in multiple renal cell types in all developmental stages and in mature non-diseased kidneys. Smad expression is in general most widespread at the earliest stages of nephron development and diminishes as components of the nephrons become more differentiated. Paucity of Smad expression in mesangial cells in contrast to widespread expression of these Smads in glomerular visceral epithelial cells in both developing and mature kidneys was remarkable. Divergent and less extensive expression of Smad4, compared with other Smad proteins, was also demonstrated in tubules of human kidneys. Based on the observed expression patterns, these findings demonstrate, for the first time, expression of the TGF-β-receptor-activated Smad2 and Smad3, the common mediator Smad4, and the inhibitory Smad7 in the developing human fetal kidney, extending observations previously made in rodent systems to humans.

Long- and short-term treatment with imatinib attenuates the development of chronic kidney disease in experimental anti-glomerular basement membrane nephritis

BACKGROUND Imatinib is a selective tyrosine kinase inhibitor that can block platelet-derived growth factor (PDGF) receptor activity. Imatinib is also known as an anti-inflammatory agent. We examined the therapeutic effects of long- or short-term imatinib treatment in Wistar-Kyoto (WKY) rats with established anti-glomerular basement membrane (GBM) nephritis. METHODS Nephrotoxic serum (NTS) nephritis was induced in WKY rats on day 0. Groups of animals were given either imatinib or vehicle daily by intraperitoneal injection, from day 7 to day 49 in the long-term treatment study, and from day 7 to 13 in the short-term treatment study; all rats were sacrificed at day 50. RESULTS In long-term treatment, imatinib showed marked renoprotective effects; imatinib suppressed proteinuria, improved renal function, attenuated the development of glomerulosclerosis and tubulointerstitial injury and reduced the expression levels of collagen type I and transforming growth factor-beta (TGF-β) in renal cortex. The key finding of the present study was that short-term treatment with imatinib also significantly attenuated the development of renal injury until day 50, although the degree of renoprotection was slightly inferior to that of long-term treatment. CONCLUSIONS These results suggest that administration of imatinib is a promising strategy for limiting the progression of glomerulonephritis (GN) to end-stage renal failure. In particular, a short period of treatment at an early stage of GN is more beneficial in terms of cost-effectiveness and reduction of adverse effects in comparison to a continuous and long period of treatment.

Microscopic polyangiitis after silicone breast implantation.

We describe the case of a patient who developed microscopic polyangiitis (MPA) after silicone breast implantation. A 60-year-old woman who had undergone silicone breast implantation was admitted to our hospital with complaints of general malaise and hematoproteinuria. She was diagnosed as having MPA with evidence of acute progressive renal failure, pulmonary hemorrhage, and positivity for myeloperoxidase-antineutrophil cytoplasmic antibody (ANCA). A renal biopsy showed severe necrotizing and crescentic glomerulonephritis with arteriolitis. The patient received high-dose steroids and plasma exchange treatment, but died of progressive pulmonary hemorrhage and multiple cerebral hemorrhage. Silicone implantation is associated with scleroderma, systemic lupus erythematosus, and rheumatoid arthritis. This case report indicates the possibility of the development of MPA after silicone breast implantation.