The dark side of hypoferremia: does iron deficiency disable innate immunity in humans?

Project No: #OxKEN-2023/19
Intake: OxKEN 2023

project overview

During infection iron is sequestered from plasma via the regulatory hormone hepcidin. Classically, this is a beneficial 'nutritional immune' mechanism that protects against extracellular siderophilic bacterial pathogens. However, it is now recognised there is a 'dark side' to hypoferremia. Prolonged reduction in serum iron not only reduces erythropoiesis, causing the anaemia of chronic disease, but additionally impacts on leukocyte function.

Observational studies in individuals with hypoferremia have repeatedly highlighted functional deficits in both the innate and adaptive immune systems with consequent susceptibility to infection and impairment of responses to immunisation. Furthermore, individuals possessing mutations in the TFRC gene that encodes transferrin receptors experience a severe combined immune deficiency, with recurrent infection, neutropenia and hypogammaglobulinaemia.

Recent work in the Drakesmith lab (see Frost et al 2022) has dissected the pivotal role of serum iron in neutrophil production and function. Experimentally induced hypoferremia in mice caused a specific reduction in both baseline and inflammation-induced granulopoesis, highlighting sensitivity to iron availability. Those neutrophils that were released displayed reduced reactive oxygen species generation, impaired phagocytosis of Gram positive and negative organisms, attenuated cytokine release, altered NETosis and reduced bacterial killing: all consistent with clinical susceptibility to infection and immunopathology. This data supports hypoferremia as a key and, most importantly, clinically modifiable modulator of innate immune function.

In this project we seek to translate this work using human experimental challenge paradigms. Self-reported healthy volunteers (18-40) will have their iron status screened for occult iron deficiency (ID) and cohorts of those with confirmed ID (transferrin saturation [TSAT] <10%), borderline ID (TSAT 10-20%) and normal iron status (TSAT >20%) recruited. Ex-vivo functional assays using granulocytes and peripheral blood mononuclear cells (PBMC) obtained from blood will be performed to explore specific neutrophil, monocyte and lymphocyte functional deficits between groups. Recapitulation of observed deficits in those with hypoferremia in vitro via deprivation of iron to effector cells will be evaluated, as will therapeutic rescue with replenishment.

The three groups will subsequently be challenged with intradermal injection (ID) of lipopolysaccharide (LPS, see Buters et al 2022 and Figure) to elicit a transient local inflammatory response in the skin, akin to cellulitis. The clinical response will be quantified via multispectral imaging (erythema, oedema) and laser Doppler (vascular reactivity) prior to formation and aspiration of a blister over the site (via negative pressure, see Figure) at multiple time points (24h, 48h, D7). Flow cytometric and transcriptomic analysis of the cellular component alongside elucidation of the humoral cytokine profile will determine if functional deficits observed in vitro are replicated in vivo. Within subject repetition of the challenge in those with ID post therapeutic replacement will enable incontrovertible proof of the link between iron status and innate immune deficiency.

This novel work is expected to have significant impact, not only on our understanding of how iron mechanistically regulates protection from infection in the common contexts of iron deficiency and inflammatory disease in humans, but on the management of the circa ~1.5 billion individuals worldwide who are ID.

Left Panel (Buters et al 202, DOI: 10.1111/bcp.149992: Timecourse of intradermal (ID) LPS injection. Right Panel (Akbar et al 2013, DOI: 10.1111/cei.12107): Formation and aspiration of aspiration of a suction blister over an ID challenge

keywords

Iron; inflammation; experimental medicine; neutrophils; innate immunity

training opportunities

The successful student will train in a truly translational environment, being mentored by an experienced supervisory team with complementary interdisciplinary skills in human and mouse immunology, experimental medicine and clinical pharmacology and therapeutics.

Placement in the Oxford Centre for Clinical Therapeutics (OCCT, Fullerton) will afford full exposure to the design, initiation and conduct of early phase clinical trials. In addition, via working alongside exisiting clinical and non-clinical post docs and DPhil students, the accrual of a unique skillset in both the practical conduct of human challenge studies and their interpretation will be facillitated. Studies will principally be conducted in the new NIHR Experimental Medicine Clinical Research Facility (https://www.ndorms.ox.ac.uk/oxford-emcrf) with access to laboratories at the Botnar Research Centre and Kennedy Institute of Rheumatology for sample processing. The student will additionally be expected to participate in and contribute to OCCT meetings and events where the evaluation of new and exisiting medicinal compounds are discussed, shaping their therapeutic development.

Through the MRC Human Immunology Unit (Drakesmith) the student will be trained in standard immunological techniques for evaluating the systemic and localised immune responses, including the functional assessment of innate immune responses (e.g. neutrophil chemotaxis, phagocytosis and reactive burst). To complement these, flow and mass cytometry, imaging, bioinformatics and ‘omics approaches, will be employed to quantify and qualitatively describe the immunological response, linking in vitro and ex vivo observations to those made in vivo.

Oxford graduate training additionally includes core workshops, seminars, career events and online resources to enable the development of intellectual and technical research capabilities, capacity for independent and team-work, and skills to effectively communicate research to the broader scientific community and general public.

We encourage anyone interested in applying to make contact with us.

key publications

Frost et al, Plasma iron controls neutrophil production and function. Science Advances, 2022. https://doi.org/10.1126/sciadv.abq5384
Bonnadonna et al, Iron regulatory protein (IRP)–mediated iron homeostasis is critical for neutrophil development and differentiation in the bone marrow. Science Advances, 2022. https://doi.org/10.1126/sciadv.abq4469
Buters et al, Intradermal lipopolysaccharide challenge as an acute in vivo inflammatory model in healthy volunteers. Br J of Clinical Pharmacology, 2022 https://doi.org/10.1111/bcp.14999
Frost et al, Hepcidin-Mediated Hypoferremia Disrupts Immune Responses to Vaccination and Infection. Med, 2020. https://doi.org/10.1016/j.medj.2020.10.004
Maini et al, A Comparison of Human Neutrophils Acquired from Four Experimental Models of Inflammation. PLoS One, 2016. https://doi.org/10.1371/journal.pone.0165502