Intrarenal Macrophages Research Articles

Substance P Improves Renal Ischemia Reperfusion Injury Through Modulating Immune Response.

Substance P (SP), an injury-inducible messenger that mobilizes bone marrow stem cells and modulates the immune response, has been suggested as a novel target for therapeutic agents. We evaluated the role of SP as an immune cell modulator during the progression of renal ischemic/reperfusion injury (IRI). Unilateral IRI induced the transient expression of endogenous SP and the infiltration of CCR7+ M1 macrophages in injured kidneys. However, SP altered the intrarenal macrophage polarization from CCR7+ M1 macrophages to CD206+ M2 macrophages in injured kidneys. SP also modulated bone marrow-derived neutrophils and mesenchymal stromal cells after IRI. SP treatment for 4 weeks starting one week after unilateral IRI significantly preserved kidney size and length and normal tubular structures and alleviated necrotic tubules, inflammation, apoptosis, and tubulointerstitial fibrosis. The beneficial effects of SP were accompanied by attenuation of intrarenal recruitment of CD4, CD8, and CD20 cells and abnormal angiogenesis. The immunomodulatory effect of SP suggested that SP could be a promising therapeutic target for preventing the progression of acute kidney injury to chronic kidney disease.

Modulation of iron homeostasis with hepcidin ameliorates spontaneous murine lupus nephritis

Lupus nephritis is the end organ manifestation of systemic lupus erythematosus. Iron metabolism and its master regulator, hepcidin, are known to regulate cell proliferation and inflammation, but their direct role in the pathophysiology of lupus nephritis remains under-investigated. Exogenous hepcidin reduced the severity of lupus nephritis in MRL/lpr mice, a preclinical model of spontaneous systemic lupus erythematosus without worsening anemia of inflammation. Hepcidin treatment reduced renal iron accumulation, systemic and intrarenal cytokines, and renal immune cell infiltration, independent of glomerular immune complex deposits and circulating autoantibodies. Hepcidin increased renal H-ferritin (a ferroxidase), reduced expression of free iron dependent DNA synthesis enzymes, Ribonucleotide Reductase 1 and 2, and intra-renal macrophage proliferation. These findings were recapitulated invitro upon treatment of macrophages with hepcidin and murine colony stimulation factor-1. Furthermore, hepcidin-treated macrophages secreted less IL-1β and IL-6 upon stimulation with the TLR3 agonist polyinosine-polycytidylic acid. Of clinical relevance, hepcidin reduced progression and severity of nephritis in old mice with established systemic autoimmunity and overt proteinuria, highlighting its therapeutic potential. Thus, our findings provide a proof-of-concept that targeting cellular iron metabolism with hepcidin represents a promising therapeutic strategy in lupus nephritis.

IL-34-Dependent Intrarenal and Systemic Mechanisms Promote Lupus Nephritis in MRL-Faslpr Mice.

In people with SLE and in the MRL-Faslpr lupus mouse model, macrophages and autoantibodies are central to lupus nephritis. IL-34 mediates macrophage survival and proliferation, is expressed by tubular epithelial cells (TECs), and binds to the cFMS receptor on macrophages and to a newly identified second receptor, PTPRZ. To investigate whether IL-34-dependent intrarenal and systemic mechanisms promote lupus nephritis, we compared lupus nephritis and systemic illness in MRL-Faslpr mice expressing IL-34 and IL-34 knockout (KO) MRL-Faslpr mice. We also assessed expression of IL-34 and the cFMS and PTPRZ receptors in patients with lupus nephritis. Intrarenal IL-34 and its two receptors increase during lupus nephritis in MRL-Faslpr mice. In knockout mice lacking IL-34, nephritis and systemic illness are suppressed. IL-34 fosters intrarenal macrophage accumulation via monocyte proliferation in bone marrow (which increases circulating monocytes that are recruited by chemokines into the kidney) and via intrarenal macrophage proliferation. This accumulation leads to macrophage-mediated TEC apoptosis. We also found suppression of circulating autoantibodies and glomerular antibody deposits in the knockout mice. This is consistent with fewer activated and proliferating intrarenal and splenic B cells in mice lacking IL-34, and with our novel discovery that PTPRZ is expressed by macrophages, B and T cells. These findings appear translatable to human patients with lupus nephritis, whose expression of IL-34, cFMS, and PTPRZ is similar to that seen in the MRL-Faslpr lupus mouse model. Moreover, expression of IL-34 in TECs correlates with disease activity. IL-34 is a promising novel therapeutic target for patients with lupus nephritis.

Metabolic pathways and immunometabolism in rare kidney diseases

ObjectivesTo characterise renal tissue metabolic pathway gene expression in different forms of glomerulonephritis.MethodsPatients with nephrotic syndrome (NS), antineutrophil cytoplasmic antibody-associated vasculitis (AAV), systemic lupus erythematosus (SLE) and healthy living donors...

Parabiosis reveals leukocyte dynamics in the kidney

The immune cellular compartment of the kidney is involved in organ development and homeostasis, as well as in many pathological conditions. Little is known about the mechanisms that drive intrarenal immune responses in the presence of renal tubular and interstitial cell death. However, it is known that tissue-resident leukocytes have the potential to have distinct roles compared with circulating cells. We used a parabiosis model in C57BL/6 CD45 congenic and green fluorescent protein transgenic mice to better understand the dynamics of immune cells in the kidney. We found F4/80Hi intrarenal macrophages exhibit minimal exchange with the peripheral circulation in two models of parabiosis, whether mice were attached for 4 or 16 weeks. Other intrarenal inflammatory cells demonstrate near total exchange with the circulating immune cell pool in healthy kidneys, indicating that innate and adaptive immune cells extensively traffic through the kidney interstitium during normal physiology. Neutrophils, dendritic cells, F4/80Low macrophages, T cells, B cells, and NK cells are renewed from the circulating immune cell pool. However, a fraction of double-negative T (CD4- CD8-) and NKT cells are long-lived or tissue resident. This study provides direct evidence of leukocyte sub-populations that are resident in the renal tissue, cells which demonstrate minimal to no exchange with the peripheral blood. In addition, the data demonstrate continual exchange of other sub-populations through uninflamed tissue.

Immunomodulatory Molecule IRAK-M Balances Macrophage Polarization and Determines Macrophage Responses during Renal Fibrosis.

Activation of various innate immune receptors results in IL-1 receptor-associated kinase (IRAK)-1/IRAK-4-mediated signaling and secretion of proinflammatory cytokines such as IL-12, IL-6, or TNF-α, all of which are implicated in tissue injury and elevated during tissue remodeling processes. IRAK-M, also known as IRAK-3, is an inhibitor of proinflammatory cytokine and chemokine expression in intrarenal macrophages. Innate immune activation contributes to both acute kidney injury and tissue remodeling that is associated with chronic kidney disease (CKD). Our study assessed the contribution of macrophages in CKD and the role of IRAK-M in modulating disease progression. To evaluate the effect of IRAK-M in chronic renal injury in vivo, a mouse model of unilateral ureteral obstruction (UUO) was employed. The expression of IRAK-M increased within 2 d after UUO in obstructed compared with unobstructed kidneys. Mice deficient in IRAK-M were protected from fibrosis and displayed a diminished number of alternatively activated macrophages. Compared to wild-type mice, IRAK-M-deficient mice showed reduced tubular injury, leukocyte infiltration, and inflammation following renal injury as determined by light microscopy, immunohistochemistry, and intrarenal mRNA expression of proinflammatory and profibrotic mediators. Taken together, these results strongly support a role for IRAK-M in renal injury and identify IRAK-M as a possible modulator in driving an alternatively activated profibrotic macrophage phenotype in UUO-induced CKD.

Intrarenal macrophage infiltration induced by T cells is associated with podocyte injury in lupus nephritis patients.

Proteinuria is the hallmark of clinical manifestation of disease activity in lupus nephritis (LN) patients, which arises from direct or indirect podocyte injury. This study is to explore the relationship between intrarenal T cell infiltration and podocyte injury in lupus nephritis (LN), and to understand the potential mechanisms of podocyte injury induced by intrarenal T cells. Sixty renal biopsies from patients diagnosed with LN were included in the present study. Histological changes in LN patients were detected by light and electron microscopy. Podocyte-specific nephrin expression in renal tissues was detected by immunofluorescence. Infiltration of T cells (CD3+ cells), infiltration of macrophages (CD68+ cells) and the expression of osteopontin (OPN) in renal tissues were examined by immunohistochemical staining. Pearson or Spearman's tests were used to perform correlation analysis. Morphologic lesions of podocytes were more severe in LN patients than in normal control subjects. Compared with normal control subjects, nephrin expression was significantly decreased in LN patients. The expression level of nephrin was significantly lower in active LN patients than in the inactive group of patients (P < 0.05). Compared with normal control subjects, the number of infiltrated intrarenal T cells and macrophages was significantly increased in LN patients. T cells were mainly distributed in renal interstitium, with very few being in glomeruli, while macrophages were mainly located in glomeruli. The number of intrarenal infiltrated T cells and macrophages in active LN patients was more than that in the inactive group (P < 0.05). Compared with normal control subjects, OPN expression in LN patients was increased significantly. The expression level of OPN in active LN patients was significantly higher than that in the inactive group (P < 0.05). Podocyte-specific nephrin was negatively correlated with 24-hour proteinuria, intrarenal T cells infiltration and intrarenal OPN expression in LN patients (P < 0.001). Intrarenal macrophages had significantly positive correlation with intrarenal OPN expression (P < 0.001). The present study provides possible links between intrarenal T cells, OPN, macrophages with reduced podocyte-nephrin and podocytopathy in systemic lupus erythematosus. In addition, infiltration of macrophages in glomeruli induced by OPN that is induced by T cells may be a crucial mechanism for podocyte injury.

Chronicity following ischaemia-reperfusion injury depends on tubular-macrophage crosstalk involving two tubular cell-derived CSF-1R activators: CSF-1 and IL-34

Two structurally unrelated ligands activate the macrophage colony stimulating factor receptor (CSF-1R, c-fms, CD115): M-CSF/CSF-1 and interleukin-34 (IL-34). Both ligands promote macrophage proliferation, survival and differentiation. IL-34 also activates the protein-tyrosine phosphatase ζ receptor (PTP-ζ, PTPRZ1). Both receptors and cytokines are increased during acute kidney injury. While tubular cell-derived CSF-1 is required for kidney repair, Baek et al (J Clin Invest 2015; 125: 3198-3214) have now identified tubular epithelial cell-derived IL-34 as a promoter of kidney neutrophil and macrophage infiltration and tubular cell destruction during experimental kidney ischaemia-reperfusion, leading to chronic injury. IL-34 promoted proliferation of both intrarenal macrophages and bone marrow cells, increasing circulating neutrophils and monocytes and their kidney recruitment. Thus, injured tubular cells release two CSF-1R activators, one (CSF-1) that promotes tubular cell survival and kidney repair and another (IL-34) that promotes chronic kidney damage. These results hold promise for the development of IL-34-targeting strategies to prevent ischaemia-reperfusion kidney injury in contexts such as kidney transplantation. However, careful consideration should be given to the recent characterization by Bezie et al. (J Clin Invest 2015; 125: 3952-3964) of IL-34 as a T regulatory cell (Treg) cytokine that modulates macrophage responses so that IL-34-primed macrophages potentiate the immune suppressive capacity of Tregs and promote graft tolerance.

Anti-Inflammatory Role of MicroRNA-146a in the Pathogenesis of Diabetic Nephropathy.

Inflammation has a critical role in the pathogenesis of diabetic complications, including diabetic nephropathy (DN). MicroRNAs have recently emerged as important regulators of DN. However, the role of microRNAs in the regulation of inflammation during DN is poorly understood. Here, we examined the in vivo role of microRNA-146a (miR-146a), a known anti-inflammatory microRNA, in the pathogenesis of DN. In a model of streptozotocin-induced diabetes, miR-146a(-/-) mice showed significantly exacerbated proteinuria, renal macrophage infiltration, glomerular hypertrophy, and fibrosis relative to the respective levels in control wild-type mice. Diabetes-induced upregulation of proinflammatory and profibrotic genes was significantly greater in the kidneys of miR-146a(-/-) than in the kidneys of wild-type mice. Notably, miR-146a expression increased in both peritoneal and intrarenal macrophages in diabetic wild-type mice. Mechanistically, miR-146a deficiency during diabetes led to increased expression of M1 activation markers and suppression of M2 markers in macrophages. Concomitant with increased expression of proinflammatory cytokines, such as IL-1β and IL-18, markers of inflammasome activation also increased in the macrophages of diabetic miR-146a(-/-) mice. These studies suggest that in early DN, miR-146a upregulation exerts a protective effect by downregulating target inflammation-related genes, resulting in suppression of proinflammatory and inflammasome gene activation. Loss of this protective mechanism in miR-146a(-/-) mice leads to accelerated DN. Taken together, these results identify miR-146a as a novel anti-inflammatory noncoding RNA modulator of DN.

Leukocyte-derived MMP9 is crucial for the recruitment of proinflammatory macrophages in experimental glomerulonephritis

Matrix metalloproteinase 9 (MMP9) is a conditionally expressed enzyme and is upregulated in glomerulonephritis. Its function in these diseases, however, remains to be fully elucidated. The induction of nephrotoxic serum nephritis (NTN) in wild-type mice resulted in an upregulation of MMP9, followed by leukocyte infiltration, albuminuria, and subsequent renal failure. MMP9 deficiency ameliorated the course of NTN as indicated by reduced histological injury and reduced infiltration of proinflammatory macrophages. The chemotaxis of MMP9-deficient macrophages in vitro was impaired. Intrarenal macrophages isolated from the kidneys of nephritic MMP9 knockout mice still displayed the typical features of a proinflammatory phenotype and were indistinguishable from wild type-derived cells. Bone marrow transplantation restored renal tissue injury and macrophage recruitment when wild type-derived donor cells were transplanted onto MMP9-deficient mice prior to the induction of NTN. Thus, leukocyte-derived MMP9 mediates the recruitment of proinflammatory macrophages into kidneys during experimental crescentic glomerulonephritis.

Toll-Like Receptor Signaling and SIGIRR in Renal Fibrosis upon Unilateral Ureteral Obstruction

Innate immune activation via IL-1R or Toll-like receptors (TLR) contibutes to acute kidney injury but its role in tissue remodeling during chronic kidney disease is unclear. SIGIRR is an inhibitor of TLR-induced cytokine and chemokine expression in intrarenal immune cells, therefore, we hypothesized that Sigirr-deficiency would aggravate postobstructive renal fibrosis. The expression of TLRs as well as endogenous TLR agonists increased within six days after UUO in obstructed compared to unobstructed kidneys while SIGIRR itself was downregulated by day 10. However, lack of SIGIRR did not affect the intrarenal mRNA expression of proinflammatory and profibrotic mediators as well as the numbers of intrarenal macrophages and T cells or morphometric markers of tubular atrophy and interstitial fibrosis. Because SIGIRR is known to block TLR/IL-1R signaling at the level of the intracellular adaptor molecule MyD88 UUO experiments were also performed in mice deficient for either MyD88, TLR2 or TLR9. After UUO there was no significant change of tubular interstitial damage and interstitial fibrosis in neither of these mice compared to wildtype counterparts. Additional in-vitro studies with CD90+ renal fibroblasts revealed that TLR agonists induce the expression of IL-6 and MCP-1/CCL2 but not of TGF-β, collagen-1α or smooth muscle actin. Together, postobstructive renal interstitial fibrosis and tubular atrophy develop independent of SIGIRR, TLR2, TLR9, and MyD88. These data argue against a significant role of these molecules in renal fibrosis.

Mineralocorticoid receptor activation contributes to salt-induced hypertension and renal injury in prepubertal Dahl salt-sensitive rats

Excessive prepubertal salt intake permanently increases blood pressure (BP). We examined the role that the mineralocorticoid receptor (MR) plays in the salt-induced hypertension and renal damage of prepubertal Dahl salt-sensitive (SS) rats. Prepubertal (6 weeks old) and adult (10 weeks old) Dahl SS rats fed a high (8.0%) salt (HS) diet for 10 weeks were compared in terms of BP and renal function. The effect of treatment between the ages of 4 and 10 weeks with the MR antagonist eplerenone (0.125% in chow), the vasodilator hydralazine (50 mg/kg/day in drinking water) or the superoxide dismutase mimetic 4-hydroxy-2,2,6,6-tetramethyl-piperidine-N-oxyl (tempol) (0.6 mmol/kg/day in drinking water) on the BP and renal function of prepubertal rats fed a HS diet for 10 weeks was also examined. Excessive salt intake starting in prepuberty was associated with a higher BP increase and greater proteinuria than if it started in adulthood. Eplerenone moderately reduced BP and markedly improved renal injury during its administration in prepubertal rats. These effects continued after drug discontinuation. Hydralazine greatly decreased BP and reduced proteinuria, but these effects were completely lost after drug discontinuation. Excessive salt increased urinary 8-hydroxy-2'-deoxyguanosine levels, intrarenal macrophage infiltration and renal plasminogen activator inhibitor-1 and transforming growth factor-beta mRNA expression. Eplerenone, but not hydralazine, attenuated these salt-induced inflammatory reactions. Tempol improved salt-induced hypertension and renal injury, even after its discontinuation. Dahl SS rats exposed to excessive salt in prepubescence show a permanent increase in susceptibility to salt-induced hypertension and proteinuria. MR activation may promote these effects at least in part by inducing oxidation and inflammation.

Circulating CSF-1 Promotes Monocyte and Macrophage Phenotypes that Enhance Lupus Nephritis

Macrophages mediate kidney disease and are prominent in a mouse model (MRL-Fas(lpr)) of lupus nephritis. Colony stimulating factor-1 (CSF-1) is the primary growth factor for macrophages, and CSF-1 deficiency protects MRL-Fas(lpr) mice from kidney disease and systemic illness. Whether this renoprotection derives from a reduction of macrophages and whether systemic CSF-1, as opposed to intrarenal CSF-1, promotes macrophage-dependent lupus nephritis remain unclear. Here, we found that increasing systemic CSF-1 hastened the onset of lupus nephritis in MRL-Fas(lpr) mice. Using mutant MRL-Fas(lpr) strains that express high, moderate, or no systemic CSF-1, we detected a much higher tempo of kidney disease in mice with the highest level of CSF-1. Furthermore, we uncovered a multistep CSF-1-dependent systemic mechanism central to lupus nephritis. CSF-1 heightened monocyte proliferation in the bone marrow (SSC(low)CD11b(+)), and these monocytes subsequently seeded the circulation. Systemic CSF-1 skewed the frequency of monocytes toward "inflammatory" (SSC(low)CD11b(+)Ly6C(high)) and activated populations that homed to sites of inflammation, resulting in a more rapid accumulation of intrarenal macrophages (CD11b(+)CSF-1R(+) or CD68(+)) that induced apoptosis of tubular epithelial cells, damaging the kidney. In humans, we found increased levels of CSF-1 in the serum, urine, and kidneys of patients with lupus compared with healthy controls. Furthermore, serum and urine CSF-1 levels correlated with lupus activity, and intrarenal CSF-1 expression correlated with the histopathology activity index of lupus nephritis. Taken together, circulating CSF-1 is a potential therapeutic target for lupus nephritis.

Unilateral ureteral obstruction in neonatal rats leads to renal insufficiency in adulthood

Although unilateral ureteropelvic junction obstruction is the most common cause of congenital obstructive nephropathy in infants and children, management remains controversial, and follow-up after pyeloplasty is generally limited to the pediatric ages. We have developed a model of temporary unilateral ureteral obstruction (UUO) in the neonatal rat: One month following the relief of five-day UUO, the glomerular filtration rate (GFR) of the postobstructed kidney was normal despite a 40% reduction in the number of glomeruli and residual vascular, glomerular, tubular, and interstitial injury. To determine whether hyperfiltration and residual injury of remaining nephrons leads to progression of renal insufficiency in later life, 31 rats were sham operated or subjected to left UUO at one day of age, with relief of UUO five days later, and were studied at one year of age. GFR was measured by inulin clearance, and the number of glomeruli, tubular atrophy, glomerular sclerosis, and interstitial fibrosis were measured by histomorphometry in sham, obstructed (UUO), and intact opposite kidneys. Intrarenal macrophages and alpha-smooth muscle actin were identified by immunohistochemistry. Despite relief of UUO, ultimate growth of the postobstructed kidney was impaired. The number of glomeruli was reduced by 40%, and GFR was decreased by 80%. However, despite significant compensatory growth of the opposite kidney, there was no compensatory increase in GFR, and proteinuria was increased. Moreover, glomerular sclerosis, tubular atrophy, macrophage infiltration, and interstitial fibrosis were significantly increased not only in the postobstructed kidney, but also in the opposite kidney. Although GFR is initially maintained following relief of five-day UUO in the neonatal rat, there is eventual profound loss of function of the postobstructed and opposite kidneys because of progressive tubulointerstitial and glomerular damage. These findings suggest that despite normal postoperative GFR in infancy, children undergoing pyeloplasty for ureteropelvic junction obstruction should be followed into adulthood. Elucidation of the cellular response to temporary UUO may lead to improved methods to assess renal growth, injury, and functional reserve in patients with congenital obstructive nephropathy.

MR imaging of intrarenal macrophage infiltration in an experimental model of nephrotic syndrome

MR imaging of intrarenal macrophage infiltration in an experimental model of nephrotic syndrome

SELECTIVE ENHANCEMENT OF THROMBOXANE IN MACROPHAGES AND KIDNEYS IN CYCLOSPORINE-INDUCED NEPHROTOXICITY

Cyclosporine (CsA) associated renal dysfunction is related in part to renal vasoconstriction. To identify the role of cyclooxygenase metabolites in the induction of vasoconstriction, we analyzed the effect of CsA on the synthesis of thromboxane (TxA2) prostacyclin (PGI2) and prostaglandin E2 (PGE2) in the kidney and peritoneal macrophages. Groups of rats were pair-fed diets enriched with 20% fish oil (FO) or corn oil (CO) for 4 weeks and then were injected with CsA 12.5 mg/kg/day i.p. for 2 weeks. CsA induced the synthesis of TxA2 and modestly reduced PGE2 and PGI2 in renal cortex and peritoneal macrophages. Feeding rats a diet enriched in FO containing omega-3 fatty acids as compared with CO without these fatty acids suppressed the increase in TxA2 and decreased the vasodilators PGE2 and PGI2 in the kidney and peritoneal macrophages, while modifying the decrease in the glomerular filtration rate and vacuolization in proximal convoluted tubules characteristic of rodent CsA-associated nephrotoxicity. Thus, CsA-initiated renal vasoconstriction is related to an increase in the vasoconstrictive Tx molecule and the reduction in vasodilator metabolites. Intrarenal macrophages represent a likely source of this Tx production. Feeding rats diets containing omega-3 fatty acids, known to be competitive inhibitors of cyclooxygenase metabolites, prevents CsA from selectively increasing TxA2 and preserves renal function.