Elsevier

Clinical Immunology

Volume 145, Issue 3, December 2012, Pages 201-208
Clinical Immunology

Fli-1 transcription factor affects glomerulonephritis development by regulating expression of monocyte chemoattractant protein-1 in endothelial cells in the kidney

https://doi.org/10.1016/j.clim.2012.09.006Get rights and content

Abstract

Expression of transcription factor Fli-1 is implicated in the development of glomerulonephritis. Fli-1 heterozygous knockout (Fli1+/−) NZM2410 mice, a murine model of lupus, had significantly improved survival and reduced glomerulonephritis. In this study, we found that infiltrated inflammatory cells were significantly decreased in the kidneys from Fli-1+/− NZM2410 mice. The expression of monocyte chemoattractant protein-1 (MCP-1) was significantly decreased in kidneys from Fli-1+/− NZM2410 mice. The primary endothelial cells isolated from the kidneys of Fli-1+/− NZM2410 mice produced significantly less MCP-1. In endothelial cells transfected with specific Fli-1 siRNA the production of MCP-1 was significantly reduced compared to cells transfected with negative control siRNA. By Chromatin Immunoprecipitation (ChIP) assay, we further demonstrated that Fli-1 directly binds to the promoter of the MCP-1 gene. Our data indicate that Fli-1 impacts glomerulonephritis development by regulating expression of inflammatory chemokine MCP-1 and inflammatory cell infiltration in the kidneys in the NZM2410 mice.

Highlights

► Fli-1 regulates expression of MCP-1 in the endothelial cells in kidneys. ► Less inflammatory cells infiltrate in the kidneys with reduced expression of Fli-1. ► Fli-1 directly binds to the promoter of the MCP-1 gene. ► Fli-1 impacts glomerulonephritis development by regulating expression of MCP-1.

Introduction

Systemic lupus erythematosus (SLE) is an autoimmune disease with a wide spectrum of clinical and immunological abnormalities [1], [2]. Glomerulonephritis is a major cause of death in both SLE human patients and animal models [1], [2], [3], [4], [5]. It has been well demonstrated that infiltration of inflammatory cells, including dendritic cells, macrophages, T cells and B cells, into the kidneys plays a critical role in the development of glomerulonephritis [5], [6], [7]. Proinflammatory cytokines and chemokines such as MCP-1 produced in kidneys have been shown to play a critical role in the infiltration of inflammatory cells into the kidneys and glomerulonephritis development [8]. MCP-1, a member of the chemokine family of inflammatory mediators and also known as chemokine (C–C motif) ligand 2 (CCL2), recruits macrophages, B cells, and T cells into the kidney and is a key mediator of glomerulonephritis in murine models of SLE [6], [8], [9]. MCP-1 deficient MRL/MpJ-Faslpr (MRL/lpr) mice, a murine model of SLE, have dramatically reduced macrophage and T cell infiltration in the kidney, decreased proteinuria, limited renal disease, and significantly prolonged survival [8]. Furthermore, pharmacologically blocking MCP-1 significantly decreases renal disease in murine lupus [10]. Several reports have demonstrated that following an immune complex deposition and before inflammatory cell infiltration, proinflammatory cytokine and chemokine production was upregulated in the kidneys in murine models of SLE, MRL/lpr mice as well as NZB/W F1 mice [6], [7]. Specifically within glomerular mesangium and in sub-endothelial areas, MCP-1 in MRL/lpr mice was up-regulated prior to inflammatory cell infiltration into kidneys [6], [7]. In addition, recent studies also suggest a key role for MCP-1 because its urinary excretion was increased in human lupus nephritis development [11], [12], [13].

The Fli-1 gene was first characterized in 1991 and belongs to the Ets gene family of transcription factors [14], [15]. Members of the Ets gene family are widely conserved in genomes of diverse organisms, including Drosophila, Xenopus, sea urchin, chicken, mouse, and human [16], [17], [18]. Expression of Fli-1 was found in endothelial cells, fibroblasts and immune cells, including T cells and B cells. Several reports have demonstrated that expression of Fli-1 protein is implicated in SLE development in both human patients and murine models [19], [20], [21], [22]. In humans, overexpression of the Fli-1 gene has been detected in peripheral blood lymphocytes of SLE patients compared to normal healthy controls, and the level of Fli-1 expression paralleled clinical activity measures of SLE [19]. Furthermore, Fli-1 transgenic mice developed a lupus-like disease [20]. We have generated Fli-1 heterozygous knockout (Fli-1+/−) MRL/lpr mice and NZM2410 mice with decreased expression of Fli-1 protein to study the role of Fli-1 in SLE development [21], [22]. Both MRL/lpr mice and NZM2410 mice share many clinical manifestations found in human SLE [4], [5]. Compared to littermate wild-type mice, we have found that both Fli-1+/− MRL/lpr and NZM2410 mice had significantly lower serum autoantibodies, lower proteinuria, reduced pathologic renal disease and markedly prolonged survival [21], [22]. In this report, we found that expression of proinflammatory chemokine MCP-1 was significantly lower in the kidneys from Fli-1+/− NZM2410 mice compared to wild-type littermate controls at the pre-disease stage. In addition, endothelial cells isolated from the kidneys of Fli-1+/− NZM2410 mice produced significantly less MCP-1 compared to renal endothelial cells from wild-type littermate controls. We further demonstrated that Fli-1 directly binds the promoter region of MCP-1 in endothelial cells, and inhibition of expression of Fli-1 with Small interfering RNA (siRNA) resulted in significantly reduced production of MCP-1 in endothelial cells. Thus, Fli-1 transcription factor affects glomerulonephritis development by regulating expression of inflammatory chemokine in endothelial cells in the kidneys of NZM2410 mice.

Section snippets

Mice

NZM2410 mice were purchased from The Jackson Laboratory (Bar Harbor, ME). Fli-1+/− NZM2410, and wild-type littermates mice used in the study were generated by backcrossing with Fli-1+/− B6 mice for 12 generations as previously reported [22]. All mice were housed under pathogen-free conditions at the animal facility of the Ralph H. Johnson Veterans Affairs Medical Center.

Genotyping of the mice by PCR

For genotyping of the mice, PCR was used to detect fragments of wild-type and Fli-1+/− alleles as previously reported [21].

Significantly reduced infiltration of inflammatory cells in kidneys from Fli-1+/− NZM2410 mice

We found that decreased expression of Fli-1 resulted in significant prolonged survival and reduced renal pathological scores in Fli-1+/− NZM2410 mice compared to wild-type littermate controls [22]. Since infiltration of inflammatory cells into kidneys plays an important role in glomerulonephritis development, we quantitated and compared the inflammatory cells in kidneys from Fli-1+/− NZM2410 mice and wild-type littermates. 18 Fli-1+/− NZM2410 mice and 21 wild-type littermate controls were

Discussion

We have demonstrated that decreased expression of Fli-1 significantly reduced renal pathology and markedly prolonged survival in murine models of lupus including NZM2410 mice [22]. In this report, we found that one of the molecular mechanisms that the expression of Fli-1 impacts on disease is the regulation of the proinflammatory chemokine MCP-1 in the endothelial cells in kidneys.

Infiltration of inflammatory cells plays a critical role in lupus nephritis development and renal injury [4], [5],

Conflict of interest statement

The authors have no financial conflicts of interest.

Acknowledgments

This study was supported in part by National Institutes of Health grants (R01AR056670 to X.K.Z.) and the Medical Research Service, Department of Veterans Affairs (to G.G. and X.K. Z.). We thank Mr. Jeremy Mathenia and Mr. Emanuel Reyes-Cortes for their excellent technical supports.

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