Chronic Kidney Disease (CKD) is a health epidemic that increases the risk of death due to systemic inflammation and anemia which affects up to 60% of patients by the time they reach dialysis. Early metabolic complications of CKD, such as elevated serum concentrations of bone-derived fibroblast growth factor (FGF) 23, are strongly associated with increased levels of inflammatory cytokines, such as interleukin-6 (IL-6) and C-reactive protein (CRP), and of iron regulators, including hepcidin, and with mortality. In a recent experimental study, we demonstrated that FGF23 contributes to inflammation by directly targeting hepatocytes through FGF receptor isoform 4 (FGFR4) and inducing PLC?/calcineurin/NFAT signaling and the production of IL-6 and CRP. We found that animal models with FGF23 elevations have increased hepatic and circulating levels of IL-6 and CRP, which did not occur if FGFR4 was blocked or deleted. IL-6 is a potent inducer of hepcidin production in the liver by activating IL-6 receptor (IL-6R) and signal transducer and activator of transcription 3 (STAT3) signaling in hepatocytes. Hepcidin lowers serum iron levels by reducing iron absorption in different cell types, including hepatocytes. Prolonged elevations of serum hepcidin levels, as observed in CKD, result in iron deficiency, and the reduction of hepcidin production can serve as a therapeutic approach to tackle anemia. The purpose of this proposal is to study functional interactions between FGF23, IL-6 and hepcidin. We postulate that by increasing IL-6 production, FGF23 can elevated hepatic and systemic levels of hepcidin and contribute to anima. This hypothesis is supported by our preliminary data showing that FGF23 significantly increases hepcidin expression in primary mouse hepatocytes. This effect occurs only after 48 hours of FGF23 treatment and involves the activation of STAT3.
In Aim 1, we will determine if phosphorylation and transcriptional activity of STAT3 as well as expression and release of hepcidin in FGF23-treated mouse hepatocytes is reduced by inhibitors of the PLC?/calcineurin/NFAT or the IL-6R/JAK/STAT3 signaling pathways. We will repeat the experiment in hepatocytes isolated from FGFR4 knockout (FGFR4-/-) mice or following viral knockdown of IL-6 or IL-6R.
In Aim 2, we will determine if deletion or blockade of FGFR4 in animal models with elevated FGF23 reduces hepcidin production and protects from anemia. We have reported that administration of FGF23 in mice increases hepatic and circulating levels of IL-6 within five days. Our preliminary studies indicate that Alport mice, a genetic model of CKD with high FGF23 levels, show elevated IL-6 and hepcidin expression in the liver and develop anemia. We will subject FGFR4-/- mice to chronic FGF23 infusions using osmotic minipumps, and we will administer a FGFR4-specific small molecule inhibitor to Alport mice, followed by the analysis of IL-6 and hepcidin expression in the liver and of serological iron parameters. We postulate that via the proposed mechanism, elevated FGF23 concentrations can contribute to CKD-associated anemia and that pharmacological FGFR4 blockade might serve as novel anti-anemic intervention in patients with CKD.
The majority of patients with chronic kidney disease (CKD) develops systemic inflammation and reduced iron levels (also called anemia) which can both contribute to early death. In patients with CKD, circulating levels of fibroblast growth factor 23 (FGF23), interleukin 6 (IL-6) and hepcidin are markedly elevated, and together these alterations cause an overactivation of the inflammatory response and anemia. In this proposal, we will characterize the crosstalk between FGF23, IL-6 and hepcidin in the liver and assess beneficial effects of inhibiting FGF23-mediated molecular events in liver cells as a novel therapeutic approach to block anemia in animal models of CKD.