Glomerular Macrophage Accumulation Research Articles

Elevated apolipoprotein C3 augments diabetic kidney disease and associated atherosclerosis in type 2 diabetes.

Diabetes increases the risk of both cardiovascular disease and kidney disease. Notably, most of the excess cardiovascular risk in people with diabetes is in those with kidney disease. Apolipoprotein C3 (APOC3) is a key regulator of plasma triglycerides, and it has recently been suggested to play a role in both type 1 diabetes-accelerated atherosclerosis and kidney disease progression. To investigate if APOC3 plays a role in kidney disease in people with type 2 diabetes, we analyzed plasma levels of APOC3 from the Veterans Affairs Diabetes Trial. Elevated baseline APOC3 levels predicted a greater loss of renal function. To mechanistically test if APOC3 plays a role in diabetic kidney disease and associated atherosclerosis, we treated black and tan, brachyury, WT and leptin-deficient (OB; diabetic) mice, a model of type 2 diabetes, with an antisense oligonucleotide (ASO) to APOC3 or a control ASO, all in the setting of human-like dyslipidemia. Silencing APOC3 prevented diabetes-augmented albuminuria, renal glomerular hypertrophy, monocyte recruitment, and macrophage accumulation, partly driven by reduced ICAM1 expression. Furthermore, reduced levels of APOC3 suppressed atherosclerosis associated with diabetes. This suggests that targeting APOC3 might benefit both diabetes-accelerated atherosclerosis and kidney disease.

A Novel Type 2 Diabetes Mouse Model of Combined Diabetic Kidney Disease and Atherosclerosis

Diabetic kidney disease and atherosclerotic disease are major causes of morbidity and mortality associated with type 2 diabetes (T2D), and diabetic kidney disease is a major cardiovascular risk factor. The black and tan, brachyury (BTBR) mouse strain with leptin deficiency (Lepob) has emerged as one of the best models of human diabetic kidney disease. However, no T2D mouse model of combined diabetic kidney disease and atherosclerosis exists. Our goal was to generate such a model. To this end, the low-density lipoprotein (LDL) receptor was targeted for degradation via inducible degrader of the LDL receptor (IDOL) overexpression, using liver-targeted adenoassociated virus serotype DJ/8 (AAV-DJ/8) in BTBR wild-type and BTBR Lepob mice. Liver-targeted IDOL-AAV-DJ/8 increased plasma LDL cholesterol compared with the control enhanced green fluorescent protein AAV-DJ/8. IDOL-induced dyslipidemia caused formation of atherosclerotic lesions of an intermediate stage, which contained both macrophages and smooth muscle cells. BTBR Lepob mice exhibited diabetic kidney disease. IDOL-induced dyslipidemia worsened albuminuria and glomerular macrophage accumulation but had no effect on mesangial expansion or podocyte numbers. Thus, by inducing hepatic degradation of the LDL receptor, we generated a T2D model of combined kidney disease and atherosclerosis. This model provides a new tool to study mechanisms, interactions, and treatment strategies of kidney disease and atherosclerosis in T2D.

Crucial Role of Mesangial Cell-derived Connective Tissue Growth Factor in a Mouse Model of Anti-Glomerular Basement Membrane Glomerulonephritis

Connective tissue growth factor (CTGF) coordinates the signaling of growth factors and promotes fibrosis. Neonatal death of systemic CTGF knockout (KO) mice has hampered analysis of CTGF in adult renal diseases. We established 3 types of CTGF conditional KO (cKO) mice to investigate a role and source of CTGF in anti-glomerular basement membrane (GBM) glomerulonephritis. Tamoxifen-inducible systemic CTGF (Rosa-CTGF) cKO mice exhibited reduced proteinuria with ameliorated crescent formation and mesangial expansion in anti-GBM nephritis after induction. Although CTGF is expressed by podocytes at basal levels, podocyte-specific CTGF (pod-CTGF) cKO mice showed no improvement in renal injury. In contrast, PDGFRα promoter-driven CTGF (Pdgfra-CTGF) cKO mice, which predominantly lack CTGF expression by mesangial cells, exhibited reduced proteinuria with ameliorated histological changes. Glomerular macrophage accumulation, expression of Adgre1 and Ccl2, and ratio of M1/M2 macrophages were all reduced both in Rosa-CTGF cKO and Pdgfra-CTGF cKO mice, but not in pod-CTGF cKO mice. TGF-β1-stimulated Ccl2 upregulation in mesangial cells and macrophage adhesion to activated mesangial cells were decreased by reduction of CTGF. These results reveal a novel mechanism of macrophage migration into glomeruli with nephritis mediated by CTGF derived from mesangial cells, implicating the therapeutic potential of CTGF inhibition in glomerulonephritis.

The pathogenic role of macrophage in lupus nephritis

Lupus nephritis is major manifestation of systemic lupus erythematosus and could cause nephrotic syndrome or chronic kidney disease might lead to end-stage renal failure. The pathogenesis of macrophage as well as lymphocyte impairment had been described in lupus nephritis. The interstitial macrophage accumulation and interstitial change or fibrosis is more important than glomerular immunoglobulin deposition or glomerular macrophage accumulation in terms of renal outcome and survival proportion. The expressions of macrophage associated proteins such as CCL2/MCP-1, MIP-1 family and their receptors, CCR2, CCR1 and CCR5 are major target of therapeutic strategy for improving renal illness. The blockade of these chemokines or chemokine receptors ameliorates renal impairment without reducing glomerular immunoglobulin deposition. Deletion of CSF-1 signaling pathway represented more excellent effect in experimental lupus nephritis. The effect of specific antagonist for macrophage associated proteins, specific thyrosine kinase inhibitor for macrophage signaling pathway on glomerulonephritis in lupus prone mice had been reported with evaluation of renal leukocyte infiltration, anti-DNA antibody reduction, the amount of proteinuria, and their survival. The depletion of macrophage could be useful therapeutic tool including M2 macrophage and have synergistic effect with other immunomodulating agents.

CD36 and Na/K-ATPase-α1 Form a Proinflammatory Signaling Loop in Kidney

Proatherogenic, hyperlipidemic states demonstrate increases in circulating ligands for scavenger receptor CD36 (eg, oxidized low-density lipoprotein [oxLDL]) and the Na/K-ATPase (eg, cardiotonic steroids). These factors increase inflammation, oxidative stress, and progression of chronic kidney disease. We hypothesized that diet-induced obesity and hyperlipidemia potentiate a CD36/Na/K-ATPase-dependent inflammatory paracrine loop between proximal tubule cells (PTCs) and their associated macrophages and thereby facilitate development of chronic inflammation and tubulointerstitial fibrosis. ApoE(-/-) and apoE(-/-)/cd36(-/-) mice were fed a high-fat diet for ≤32 weeks and examined for physiologic and histologic changes in renal function. Compared with apoE(-/-), apoE(-/-)/cd36(-/-) mice had improved creatinine clearance and blood pressure which corresponded histologically with less glomerular and tubulointerstitial macrophage accumulation, foam cell formation, oxidant stress, and interstitial fibrosis. Coimmunopreciptation and a cell surface fluorescence-based crosslinking assay showed that CD36 and Na/K-ATPase α-1 colocalized in PTCs and macrophages, and this association was increased by oxLDL or the cardiotonic steroid ouabain. OxLDL and ouabain also increased activation of Src and Lyn in PTCs. Cell-free conditioned medium from PTCs treated with oxLDL or ouabain increased macrophage migration. OxLDL, ouabain, or plasma isolated from high-fat diet-fed mice stimulated reactive oxygen species production in PTCs, which was inhibited by N-acetyl-cysteine, apocynin, or Na/K-ATPase α-1 knockdown. These data suggest that ligands generated in hyperlipidemic states activate CD36 and the Na/K-ATPase and potentiate an inflammatory signaling loop involving PTCs and their associated macrophages, which facilitates the development of chronic inflammation, oxidant stress, and fibrosis underlying the renal dysfunction common to proatherogenic, hyperlipidemic states.

Contrasting Dose–Effects of Multi-glycoside of Tripterygium wilfordii HOOK. f. on Glomerular Inflammation and Hepatic Damage in Two Types of Anti-Thy1.1 Glomerulonephritis

Multi-glycoside, one of the extracted compounds from Tripterygium wilfordii HOOK. f. (GTW), has been shown to be clinically effective in suppressing glomerular inflammation in chronic kidney disease. However, its clinical application is often limited by its cytotoxic actions on the liver. This study was performed to contrast the dose–effects of GTW on glomerular inflammation and hepatic damage in two types of anti-Thy1.1 glomerulonephritis (GN). Rats with acute or chronic anti-Thy1.1 GN were either left untreated (the Vehicle group) or treated with a high or low dose of GTW and sacrificed on day 7 or day 45. GTW was administrated 3 days before or at the same time as the antibody injection and lasted until sacrifice. GTW at high dose ameliorated glomerular macrophage accumulation, mesangial proliferation, proteinuria, and interleukin-2 expression in the acute anti-Thy1.1 GN model, but caused structural and functional lesions in the liver. In contrast, GTW at low dose improved activated macrophage and T lymphocyte infiltration, mesangial injury, proteinuria, and interleukin-2 and interferon-γ expressions without hepatic toxicity in the chronic model of GN induced by anti-Thy1.1 antibody. In conclusion, GTW at low dose not only effectively inhibits glomerular inflammation but also avoids severe injuries to the liver.

Effects of p38 mitogen-activated protein kinase inhibition on anti-neutrophil cytoplasmic autoantibody pathogenicity in vitro and in vivo

ObjectiveTo determine whether inhibition of p38 mitogen-activated protein kinase (p38MAPK) reduces the pathogenicity of anti-neutrophil cytoplasmic autoantibodies (ANCAs) in vitro and in vivo.MethodsThe effects of the p38MAPK-specific inhibitor AR-447 were...

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.

Blockade of the c-Jun amino terminal kinase prevents crescent formation and halts established anti-GBM glomerulonephritis in the rat

Macrophages induce acute renal injury in anti-glomerular basement membrane (GBM) glomerulonephritis. This operates, in part, via activation of the c-Jun amino terminal kinase (JNK) signaling pathway. However, it is unknown whether inhibition of JNK signaling is effective once the proinflammatory response is established in the injured kidney. This study examined whether blockade of JNK signaling could halt disease progression, including crescent formation, in a model of severe crescentic anti-GBM glomerulonephritis. WKY rats were immunized with sheep IgG and then injected with sheep anti-GBM serum (day 0). Animals were treated with the JNK inhibitor, CC-401, vehicle alone, or no treatment from day 7 until being killed on day 24 of disease. Untreated animals at day 7 showed significant proteinuria, focal glomerular lesions, marked glomerular macrophage and T-cell accumulation, and upregulation of proinflammatory mediators (TNF-α, iNOS, MMP-12). Untreated and vehicle-treated groups displayed severe glomerulonephritis at day 24 with renal impairment and worsening proteinuria. These animals had severe glomerular lesions, with 60% of glomeruli exhibiting fibrocellular crescents, in association with increased macrophage and T-cell accumulation (including macrophage giant cells) and a further increase in mRNA levels of TNF-α, iNOS, MMP-12, and TGF-β1. In contrast, CC-401 treatment prevented renal impairment, suppressed proteinuria, and prevented severe glomerular and tubulointerstitial lesions, including crescent formation and granulomatous-like lesions. These protective effects were independent of glomerular macrophage and T-cell accumulation, and of the humoral immune response. CC-401 treatment inhibited expression of both pro- and antiinflammatory molecules (interleukin-10 and heme oxygenase-1). In addition, IL-1 induced MMP-12 and IL-10 production by cultured macrophages was found to be JNK dependent. In conclusion, blockade of JNK signaling provides substantial protection against the progression of crescentic anti-GBM glomerulonephritis, which may be, in part, due to inhibition of the macrophage proinflammatory response.

CD100 Enhances Dendritic Cell and CD4+ Cell Activation Leading to Pathogenetic Humoral Responses and Immune Complex Glomerulonephritis

CD100, a member of the semaphorin family, is a costimulatory molecule in adaptive immune responses by switching off CD72's negative signals. However, CD100's potential pathogenetic effects in damaging immune responses remain largely unexplored. We tested the hypothesis that CD100 plays a pathogenetic role in experimental immune complex glomerulonephritis. Daily injection of horse apoferritin for 14 days induced immune complex formation, mesangial proliferative glomerulonephritis and proteinuria in CD100-intact (CD100+/+) BALB/c mice. CD100-deficient (CD100-/-) mice were protected from histological and functional glomerular injury. They exhibited reduced deposition of Igs and C3 in glomeruli, reduced MCP-1 and MIP-2 intrarenal mRNA expression, and diminished glomerular macrophage accumulation. Attenuated glomerular injury was associated with decreased Ag-specific Ig production, reduced CD4+ cell activation and cytokine production. Following Ag injection, CD4+ cell CD100 expression was enhanced and dendritic cell CD86 expression was up-regulated. However, in CD100-/- mice, dendritic cell CD86 (but not CD80) up-regulation was significantly attenuated. Following i.p. immunization, CD86, but not CD80, promotes early Ag-specific TCR-transgenic DO11.10 CD4+ cell proliferation and IFN-gamma production, suggesting that CD100 expression enables full expression of CD86 and consequent CD4+ cell activation. Transfer of CD100+/+ DO11.10 cells into CD100-/- mice resulted in decreased proliferation demonstrating that CD100 from other sources in addition to CD100 from Ag-specific CD4+ cells plays a role in initial T cell proliferation. Although T cell-B cell interactions also may be relevant, these studies demonstrate that CD100 enhances pathogenetic humoral immune responses and promotes the activation of APCs by up-regulating CD86 expression.

Monocyte chemoattractant protein-1 promotes the development of diabetic renal injury in streptozotocin-treated mice

Diabetic nephropathy involves a renal inflammatory response induced by the diabetic milieu. Macrophages accumulate in diabetic kidneys in association with the local upregulation of monocyte chemoattractant protein-1 (MCP-1); however, the contribution of macrophages to renal injury and the importance of MCP-1 to their accrual are unclear. Therefore, we examined the progression of streptozotocin (STZ)-induced diabetic nephropathy in mice deficient in MCP-1 in order to explore the role of MCP-1-mediated macrophage accumulation in the development of diabetic kidney damage. Renal pathology was examined at 2, 8, 12 and 18 weeks after STZ treatment in MCP-1 intact (+/+) and deficient (-/-) mice with equivalent blood glucose and hemoglobin A1c levels. In MCP-1(+/+) mice, the development of diabetic nephropathy was associated with increased kidney MCP-1 production, which occurred mostly in tubules, consistent with our in vitro finding that elements of the diabetic milieu (high glucose and advanced glycation end products) directly stimulate tubular MCP-1 secretion. Diabetes of 18 weeks resulted in albuminuria and elevated plasma creatinine in MCP-1(+/+) mice, but these aspects of renal injury were largely suppressed in MCP-1(-/-) mice. Protection from nephropathy in diabetic MCP-1(-/-) mice was associated with marked reductions in glomerular and interstitial macrophage accumulation, histological damage and renal fibrosis. Diabetic MCP-1(-/-) mice also had a smaller proportion of kidney macrophages expressing markers of activation (inducible nitric oxide synthase or sialoadhesin) compared to diabetic MCP-1(+/+) mice. In conclusion, our study demonstrates that MCP-1-mediated macrophage accumulation and activation plays a critical role in the development of STZ-induced mouse diabetic nephropathy.

Granulocyte Macrophage Colony-Stimulating Factor Expression by Both Renal Parenchymal and Immune Cells Mediates Murine Crescentic Glomerulonephritis

GM-CSF has previously been demonstrated to be important in crescentic glomerulonephritis (GN). As both renal parenchymal cells and infiltrating inflammatory cells produce GM-CSF, their separate contributions to inflammatory renal injury were investigated by creation of two different types of GM-CSF chimeric mice: (1) GM-CSF-deficient (GM-CSF-/-)-->wild-type (WT) chimeras with leukocytes that are unable to produce GM-CSF and (2) WT-->GM-CSF-/- chimeras with deficient renal cell GM-CSF expression. Crescentic anti-glomerular basement membrane GN was induced in WT, GM-CSF(-/-)-->WT chimeras, WT-->GM-CSF-/- chimeras, and GM-CSF-/- mice by planting an antigen (sheep globulin) in their glomeruli. WT mice developed severe crescentic GN, whereas GM-CSF-/- were protected from development of disease. Glomerular T cell recruitment, CD40+ glomerular cells, and renal IFN-gamma and TNF expression were similar in both chimeras and WT mice but significantly reduced in GM-CSF-/- mice, indicating that either leukocyte or renal sources of GM-CSF are sufficient to drive these aspects of the inflammatory response. Restricted expression of GM-CSF revealed a major role for renal cell-derived GM-CSF but a minor role for leukocyte-derived GM-CSF in the formation of cellular crescents; glomerular MHC II expression; serum creatinine; and monocyte chemoattractant protein-1, vascular cellular adhesion molecule, and IL-1beta expression. Glomerular macrophage accumulation, proteinuria, and interstitial infiltrate were equivalent in both chimeric groups but intermediate between WT and GM-CSF-/-, indicating that both sources are required for the full development of glomerular injury in crescentic GN.

Aggravation of Anti-Myeloperoxidase Antibody-Induced Glomerulonephritis by Bacterial Lipopolysaccharide: Role of Tumor Necrosis Factor-α

Aggravation of Anti-Myeloperoxidase Antibody-Induced Glomerulonephritis by Bacterial Lipopolysaccharide: Role of Tumor Necrosis Factor-α

Expression of CX3CL1/fractalkine by mesangial cells in vitro and in acute anti-Thy1 glomerulonephritis in rats.

Mesangial cells (MCs) can promote glomerular macrophage accumulation in glomerulonephritis through production of a variety of chemokines. This study investigated the potential of MCs to synthesize CX3CL1/fractalkine, a CX3C chemokine, both in vitro and in acute anti-Thy1 glomerulonephritis in rats. Anti-Thy1 glomerulonephritis was induced in Wistar rats by a single injection of mouse anti-rat Thy1.1 antibody intravenously. Glomerular mRNAs for CX3CL1/fractalkine, CCL2/monocyte chemoattractant protein (MCP)-1, and their cognate receptors, CX3CR1 and CCR2, were determined by northern blot analysis or reverse-transcription polymerase chain reaction. CX3CL1/fractalkine mRNA and protein expression in vivo was localized by in situ hybridization and immunohistochemistry. Monocytes/macrophages and activated MCs were detected by immunohistochemistry. Regulation of CX3CL1/fractalkine expression in cultured MCs was determined by northern and western blot analysis. After induction of anti-Thy1 disease, glomerular CX3CL1/fractalkine mRNA was significantly up-regulated, peaking at 2 h and sustaining into day 5 of the nephritis. A corresponding increase in urinary CX3CL1/fractalkine protein was evident after day 1 of the nephritis, but became more prominent during the MC proliferative phase (days 3-5). Meanwhile, induction of glomerular CCL2/MCP-1 mRNA and urinary CCL2/MCP-1 protein occurred within 24 h, and was barely detectable after day 3 of the nephritis. Urinary CCL2/MCP-1, but not CX3CL1/fractalkine, correlated with glomerular macrophage accumulation (r = 0.936, P<0.01) and glomerular CCR2 mRNA expression (r = 0.965, P<0.01). In contrast, only urinary CX3CL1/fractalkine coincided temporally to glomerular mRNA for CX3CR1 (r = 0.809, P < 0.01). Combined in situ hybridization and immunohistochemistry revealed that activated MCs were a major source for CX3CL1/fractalkine mRNA and protein during days 3-5 of the nephritis. Incubation of cultured MCs with tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, platelet-derived growth factor (PDGF)-AB or basic fibroblast growth factor (bFGF) significantly up-regulated CX3CL1/fractalkine mRNA and protein expression. This cytokine- and growth factor-stimulated CX3CL1/fractalkine expression could be abolished by the nuclear factor-kappaB inhibitors, curcumin and MG132. Our data demonstrate that activated MCs are a source for the augmented glomerular CX3CL1/fractalkine expression during the proliferative phase of acute anti-Thy1 glomerulonephritis. Up-regulation of MC CX3CL1/fractalkine by TNF-alpha, IL-1beta, PDGF-AB and bFGF is mediated, at least in part, via the nuclear factor-kappaB signalling pathway. The differential expression of CCL2/MCP-1 and CX3CL1/fractalkine may sequentially recruit distinct subsets of monocytes to the glomerulus during acute anti-Thy1 glomerulonephritis.

Up-regulation of macrophage colony-stimulating factor (M-CSF) and migration inhibitory factor (MIF) expression and monocyte recruitment during lipid-induced glomerular injury in the exogenous hypercholesterolaemic (ExHC) rat.

Although macrophages play an important role in lipid-induced glomerular injury, we know little of the mechanisms by which hyperlipidaemia induces monocyte recruitment. This study investigated the role of M-CSF and macrophage MIF in monocyte recruitment during the development of lipid-induced glomerular injury in the susceptible ExHC rat strain. Groups of five ExHC rats were fed a high cholesterol diet (HCD) containing 3% cholesterol, 0.6% sodium cholate and 15% olive oil, and killed after 3 days, 1, 2 or 6 weeks. Control animals were killed on day 0 or after 6 weeks on a normal diet. Animals were hypercholesterolaemic 3 days after the induction of the HCD, but showed no change in plasma triglycerides over the 6-week period. Glomerular macrophage accumulation was first evident at 1-2 weeks and increased up to week 6, when macrophage-derived foam cells were seen in almost all glomeruli, and segmental lesions and mild proteinuria were also evident. Combined in situ hybridization and immunohistochemistry staining demonstrated that, coincident with the induction of hypercholesterolaemia on day 3, there was marked up-regulation of M-CSF and MIF mRNA expression by intrinsic glomerular cells (mostly mesangial cells and podocytes) which preceded monocyte recruitment. There was a highly significant correlation between the number of M-CSF and MIF-positive cells and glomerular macrophage accumulation over the 6-week period. Although some glomerular macrophages and foam cells exhibited M-CSF and MIF expression, the major source of these molecules was intrinsic glomerular cells. No local macrophage proliferation was observed during the development of glomerular lesions. In conclusion, hypercholesterolaemia caused marked up-regulation of M-CSF and MIF expression by intrinsic glomerular cells, which correlated with monocyte recruitment and the development of lipid-induced glomerular injury. This is the first study to implicate local synthesis of MIF in the pathogenesis of lipid-induced lesions.

Heterogeneity of antigen expression explains controversy over glomerular macrophage accumulation in mouse glomerulonephritis.

Many antibody labelling studies have suggested that there are few or no glomerular macrophages in mouse models of glomerulonephritis, despite the presence of a prominent interstitial macrophage infiltrate. These findings conflict with studies of human and rat glomerulonephritis. Therefore, we examined whether heterogeneity of macrophage antigen expression could explain this apparent discrepancy. Kidneys were collected from normal mice and mice killed at 2 and 10 days after induction of accelerated anti-glomerular basement membrane (GBM) glomerulonephritis. Following fixation, macrophages were detected by immunoperoxidase staining in serial kidney sections using antibodies recognising CD11b, F4/80 and CD68. Induction of anti-GBM nephritis caused a progressive increase in glomerular and interstitial leukocytes. At days 2 and 10, there were more CD68+ macrophages in glomeruli than macrophages expressing CD11b or F4/80. At day 10, CD68+ macrophages accounted for almost all glomerular CD45+ total leukocytes. In contrast, CD11b+ and F4/80+ macrophages at day 10 accounted for only 65 and 13% of glomerular leukocytes, respectively. However, in the interstitium the number of macrophages expressing CD68, CD11b and F4/80 were not different. Antibody detection of mouse CD68 identifies all glomerular macrophages in mouse anti-GBM nephritis, indicating a similar infiltrate to that seen in human and rat anti-GBM nephritis. Our finding of substantial heterogeneity in glomerular macrophage antigen expression in this model suggests that previous studies of mouse glomerulonephritis may have underestimated glomerular macrophages and their role in glomerular injury.

Adoptive transfer studies demonstrate that macrophages can induce proteinuria and mesangial cell proliferation

Glomerular macrophage accumulation is a feature of proliferative human and experimental glomerulonephritis. However, our understanding of the role of macrophages in the induction of renal injury is based upon indirect evidence from depletion studies, most of which lack specificity for this cell type. Therefore, an adoptive transfer approach was used to directly assess the potential of macrophages to induce renal injury. Accelerated anti-glomerular basement membrane (anti-GBM) disease was induced in rats by immunization with sheep IgG (day -5), followed by administration of sheep anti-rat GBM serum (day 0), with animals killed on day 2. To facilitate the adoptive transfer studies, immunized animals were made leukopenic by cyclophosphamide (CyPh) given on day -2. Bone marrow-derived (BM) or NR8383 macrophages were transferred by tail vein injection 24 hours after injection of anti-GBM serum, with animals killed 3 or 24 hours after transfer. Pretreatment with CyPh prevented glomerular leukocyte accumulation and completely inhibited proteinuria, glomerular cell proliferation and hypercellularity in accelerated anti-GBM disease. Adoptive transfer led to significant glomerular accumulation of BM or NR8383 macrophages within 3 hours of injection, and this was still evident 24 hours later. Adoptive transfer of BM or NR8383 macrophages induced proteinuria (63 +/- 16 BM vs. 5 +/- 2 mg/24 h CyPh control; P < 0.001), glomerular cell proliferation (5.1 +/- 1.2 BM vs. 0.5 +/- 0.1 PCNA+ cells/gcs CyPh; P < 0.001) and glomerular hypercellularity (51.2 +/- 2.0 BM vs. 41.9 +/- 0.9 nuclei/gcs CyPh; P < 0.001). The degree of renal injury correlated with the number of transferred glomerular macrophages. Two-color immunostaining demonstrated that most glomerular proliferative cell nuclear antigen+ (PCNA+) proliferating cells were OX-7+ mesangial cells. CyPh treatment did not prevent up-regulation of glomerular intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule (VCAM-1) expression or an increase in urinary monocyte chemoattractant protein-1 (MCP-1) excretion. This study provides the first direct evidence that macrophages can induce renal injury in terms of proteinuria and mesangial cell proliferation.

Crescentic glomerulonephritis is diminished in fibrinogen-deficient mice

Crescentic forms of glomerulonephritis are characterized by the accumulation of fibrin and cells in Bowman's space and are associated with a rapid loss of renal function. Accumulation of fibrin in the glomerular tufts is thought to promote macrophage infiltration and glomerular injury. To directly explore the role of fibrin(ogen) in the development of crescentic glomerulonephritis, antiglomerular basement membrane nephritis was induced in fibrinogen-deficient and control mice. Glomeruli from control mice developed severe disease including fibrin deposits, inflammatory cell accumulation, and crescent formation (46.3 +/- 7.3% of glomeruli). Fibrinogen-deficient mice developed significantly milder disease with fewer glomerular crescents (24.0 +/- 4.7% of glomeruli; P < 0.03). Glomerular macrophage accumulation was diminished in fibrinogen-deficient mice (0.9 +/- 0.4 macrophages/glomerular cross section) relative to control mice (3.9 +/- 1.4 macrophages/glomerular cross section; P < 0.03). Finally, renal function as assessed by serum creatinine was better maintained in fibrinogen-deficient mice. These results indicate that although fibrin(ogen) is not essential for the development of glomerular crescents, it contributes significantly to the pathogenesis of crescentic glomerulonephritis by promoting glomerular macrophage accumulation and impairing filtration.

Crescentic glomerulonephritis is diminished in fibrinogen-deficient mice

First published August 21, 2001; 00.1152/ajprenal.0002.2001.— Crescentic forms of glomerulonephritis are characterized by the accumulation of fibrin and cells in Bowman's space and are associated with a rapid loss of renal function. Accumulation of fibrin in the glomerular tufts is thought to promote macrophage infiltration and glomerular injury. To directly explore the role of fibrin(ogen) in the development of crescentic glomerulonephritis, antiglomerular basement membrane nephritis was induced in fibrinogen-deficient and control mice. Glomeruli from control mice developed severe disease including fibrin deposits, inflammatory cell accumulation, and crescent formation (46.3 ± 7.3% of glomeruli). Fibrinogen-deficient mice developed significantly milder disease with fewer glomerular crescents (24.0 ± 4.7% of glomeruli; P &lt; 0.03). Glomerular macrophage accumulation was diminished in fibrinogen-deficient mice (0.9 ± 0.4 macrophages/glomerular cross section) relative to control mice (3.9 ± 1.4 macrophages/glomerular cross section; P &lt; 0.03). Finally, renal function as assessed by serum creatinine was better maintained in fibrinogen-deficient mice. These results indicate that although fibrin(ogen) is not essential for the development of glomerular crescents, it contributes significantly to the pathogenesis of crescentic glomerulonephritis by promoting glomerular macrophage accumulation and impairing filtration.

Interleukin 12 induces crescentic glomerular lesions in a high IgA strain of ddY mice, independently of changes in IgA deposition.

Our recently established high immunoglobulin (Ig)A inbred strain (HIGA) of ddY mice showed constantly high serum IgA levels, progressive mesangial sclerosis accompanied by IgA deposits, and elevated renal expression of transforming growth factor (TGF)-beta, mimicking IgA nephropathy. In the present study, we assessed the role of the immune system, especially of T cells, in this strain. The in vitro production of interferon (IFN)-gamma, interleukin (IL)-4 and TGF-beta1 by splenic CD4+ T cells was assessed in HIGA mice at 14 and 28 weeks of age by comparison with age-matched C57BL/6 and BALB/c mice, T-helper (Th) 1, and Th2 prone controls respectively. Moreover, recombinant murine IL-12 was administered intraperitoneally to HIGA mice and serum IgA and renal lesions were analysed. The production of IFN-gamma by splenic CD4+ T cells was markedly upregulated in HIGA mice at both ages as compared with age-matched C57BL/6 and BALB/c mice. Although splenic CD4+ T cells from HIGA mice produced less IL-4 than those from BALB/c mice at both ages, the former produced significantly more IL-4 with age, which contrasted with the age-associated decrease in the latter. Moreover, TGF-beta1 production of these cells in HIGA mice was equal to or greater than that in the two groups of control mice at both ages. Daily intraperitoneal administration of IL-12 for 1 week significantly enhanced crescent formation with glomerular macrophage accumulation and interstitial cell infiltration, whereas it reduced the serum IgA level. In HIGA mice, Th1 is markedly upregulated from a young age and there is an age-associated Th2 increase with TGF-beta1 upregulation in helper T cells. The former may be related to the exacerbation of inflammatory renal lesions on IL-12 administration, while the latter may contribute to increased IgA production, leading to glomerular IgA deposition and progressive glomerulosclerosis in HIGA mice. The pathogenic role of T cell function and fluctuation of these subsets, especially the Th1/Th2 balance, is crucial to the immunopathological phenotype of the renal lesions in HIGA mice.