Iron homeostasis and monocyte differentiation in patients with chronic kidney disease

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PROJECT SUMMARY Chronic kidney disease (CKD) affects 10% of the population worldwide. More than 37 million people are estimated to have CKD in the US, and 2 in every 1000 Americans need dialysis or a kidney transplant to survive. CKD has numerous systemic complications including anemia and dysfunctional systemic iron homeostasis. Kidney fibrosis is the final mechanism common for all progressive kidney disorders. Unfortunately, very few therapies are available to slow the progression of kidney fibrosis in patients with CKD. Kidney macrophages are one of the key cells implicated in the pathophysiology of kidney fibrosis. The majority of kidney macrophages originate from circulating monocytes. In his K08-funded project Dr. Akchurin elucidated the pathologic role of depletion of labile iron pool in kidney macrophages in propagation of kidney fibrosis. This R03 proposal will enhance his capabilities of deciphering the role of iron in pathologic behavior of myeloid cells in CKD by now, in addition to kidney macrophages focusing on their precursors, circulating monocytes. Based on the preliminary data, the central hypothesis of this proposal is that in patients with CKD, circulating monocytes, similar to kidney macrophages, exhibit pathologically depleted intracellular labile iron pool LIP, which alters their phenotype by inducing CX3CR1 and other receptors facilitating chemotaxis and tissue invasion. Repletion of this intracellular monocyte labile iron pool through correction of systemic iron imbalance will be associated with reduction of this pro-inflammatory differentiation. This hypothesis will be tested in two specific aims: (1) determine the intracellular iron status of classical, intermediate, and non-classical circulating monocytes in patients with CKD and (2) elucidate the role of systemic and intracellular iron status in the differentiation of monocytes in patients with CKD. To test this hypothesis, the applicant will use the existing repository of peripheral blood mononuclear cells collected from children with CKD with and without functional or absolute iron deficiency and from healthy control children. He will evaluate these cells using single cell transcriptomic and multicolor flow cytometry approaches with the focus on delineating the phenotypic features responsible for subsequent tissue invasion, such as expression relevant chemokine receptors, including CX3CR1. Furthermore, he will perform ex vivo evaluation of human monocytes collected from CKD patients to directly assess their relevant functional characteristics in the presence and absence of iron stimulation. Monocyte functional characteristics will be analyzed I the context of clinical parameters, levels of circulating and urine-excreted chemokines, and systemic parameters of iron homeostasis. Results techniques an kidney from t he outlined experiments will provide preliminary data, as well as experience in for the functional ex-vivo testing of uman monocytes, to support the K08 awardee's application for R01 proposal to evaluate the impact of iron metabolism and iron therapy on myeloid cells in the context of fibrosis and CKD progression. h