Background/Purpose: Fli-1 expression level, a member of the Ets family of transcription factors, is a mitigating factor in the development of nephritis in murine models of systemic lupus erythematosus. We have reported that reduced expression of Fli-1 in MRL/lpr mice, a murine model of lupus, resulted in significantly prolonged survival and reduced glomerulonephritis. Inflammatory cell infiltration into the kidney plays a critical role in lupus nephritis progression. In kidney, Fli-1 is mainly expressed in endothelial cells in the interstitial capillaries and glomerulus. The role of Fli-1 regarding immune cell infiltration into the kidney is still unclear. We hypothesized that reduced Fli-1 expression affects local chemokine expression, cytokine production and influences immune cell infiltration into the kidney. To investigate this hypothesis, we performed following experiments.

Methods: We generated MRL/lpr transgenic mice expressing green fluorescent protein (GFP), and isolated spleen cells from these mice (6 to 8 months old). One million cells were injected into 9 Fli-1 heterozygous (+/-) and 13 wild-type (WT) MRL/lpr mice (4 to 8-month-old). Mice were sacrificed 18 hours after injection, after that kidneys were removed and frozen sections were made. Inflammatory cells with GFP were counted by immunofluorescence studies. The expression of chemokines (CCL2, CCL3, CCL4, and CCL5) and IL-6 in the recipient kidney was analyzed by real-time PCR. To investigate whether the Fli-1 disruption actually reduces cytokine production in vitro, we transfected Fli-1 specific siRNA to the MS1 cells (murine endothelial cell line) and analyzed the supernatant IL-6 concentration after LPS stimulation. In addition, whether Fli-1 directly regulates IL-6 expression by binding IL-6 promoter region, we performed chromatin immunoprecipitation (ChIP) assay using anti-Fli-1 antibody in MS1 cells. Isolated ChIP DNA was amplified by real-time PCR using IL-6 promoter primers.

Results: The infiltrated GFP positive inflammatory cells were significantly decreased in kidneys of Fli-1+/- MRL/lpr mice. The number of infiltrated T cells, B cells and macrophages were also significantly decreased. The relative mRNA expression of chemokines (CCL2, 3 and 4) and IL-6 in the kidney (WT vs Fli-1+/-) showed significantly lower in Fli-1+/- MRL/lpr mice. CCL5 expression was also showed lower trends (P=0.079). Fli-1 disruption of MS1 cells using siRNA showed reduced production of IL-6 after LPS stimulation. Furthermore, ChIP assay using IL-6 promoter primers indicated that Fli-1 directly binds to IL-6 promoter regions in murine endothelial cells.

Conclusion: Fli-1 regulates lupus nephritis development by modulating expression of inflammatory chemokines, IL-6 and immune cell infiltration into the kidney.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/fli-1-regulates-immune-cell-infiltration-through-affecting-local-chemokine-expression-and-il-6-expression-in-the-kidney-of-lupus-prone-mice/