Regulation of iron balance

INTRODUCTION — A tight regulation of iron balance is essential to avoid both iron deficiency and overload. The regulation of iron metabolism involves the interaction of a number of specific proteins, as well as the interplay between iron absorption, utilization, recycling, and loss. This topic review will discuss these factors [1].

Clinical implications are discussed separately:

●Iron deficiency – (See "Iron deficiency in infants and children <12 years: Treatment" and "Iron requirements and iron deficiency in adolescents" and "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults".)

●Iron overload – (See "Approach to the patient with suspected iron overload" and "Clinical manifestations and diagnosis of hereditary hemochromatosis" and "HFE and other hemochromatosis genes" and "Gene test interpretation: HFE (hereditary hemochromatosis gene)".)

ROLE OF SPECIFIC PROTEINS — Our understanding of iron metabolism and the hallmarks of iron deficiency and iron excess is based upon the biology of a number of critical proteins, including but not limited to the following (figure 1) [2]:

●Transferrin (Tf), the plasma iron transport protein.

●Transferrin receptor (TfR), the cellular receptor for iron-bound transferrin.

●Ferritin (Ft), the cellular iron storage protein.

●Iron regulatory protein 1 and 2 (IRP1 and IRP2), the cellular iron-sensing proteins.

●Divalent metal transporter 1 (DMT1, also called Nramp2, DCT1, and solute carrier family 11, member 2 [SLC11A2]), the duodenal iron importer at the luminal surface.

●Duodenal cytochrome B (DyctB, also called DcytB), an iron reductase at the luminal site of the enterocyte enabling reduction of iron for absorption.

●STEAP3, the intracellular ferric reductase essential for transcellular iron transport.

●Ferroportin (Ireg1, SLC40A1, Mtp1), the cellular iron exporter. Pathogenic variants in the SLC40A1 (also called FPN1) gene, which encodes ferroportin, cause rare forms of iron overload.

●Hephaestin, which likely cooperates with ferroportin for exporting iron from enterocytes to transferrin.

●Ceruloplasmin, a plasma metalloprotein required by ferroportin to export iron from macrophages, hepatocytes, and glial cells.

●Poly(rC) binding proteins 1 and 2 (PCBP1 and PCBP2), the intracellular iron chaperones that bind to cytosines in RNA.

●Hepcidin, the key negative regulator of intestinal iron absorption as well as macrophage iron release. Mutations of hepcidin cause a rare form of juvenile hemochromatosis.

●HFE, mutations of which are responsible for the common form of hereditary hemochromatosis.

●TFR2, mutations of which are responsible for a rare form of hereditary hemochromatosis.

●Hemojuvelin, a BMP coreceptor and hepcidin regulator, mutations of which are responsible for the common form of juvenile hemochromatosis.

●Bone morphogenetic proteins (BMPs; cytokines produced by endothelial cells) such as BMP6 and BMP2, which activate hepcidin by binding to BMP receptors and signaling through SMAD proteins.

●Matriptase 2/TMPRSS6, the liver hepcidin inhibitor with a major role in iron deficiency.

●Erythroferrone, a hormone produced by erythroblasts stimulated by erythropoietin, which downregulates hepcidin in response to erythropoiesis expansion.

●Fibrinogen-like protein 1, a hepatokine that contributes to suppression of hepcidin in hypoxia.

●NCOA4, the nuclear receptor coactivator 4, cargo receptor for intracellular ferritin degradation and cellular iron mobilization.

Transferrin — The gene for apo-transferrin is on the long arm of chromosome 3. It codes for a protein (molecular weight 80 kDa) that tightly binds one or two ferric (Fe³⁺) iron molecules and is the major transporter for iron trafficking through the plasma. Most of the Tf, which has a half-life of eight days, is made in the liver, where its synthesis is considerably increased in states of iron deficiency or reduced upon iron loading or inflammation by unknown mechanisms [1,3].

ApoTf acquires iron from ferroportin, becoming monoferric or (when iron is abundant) diferric. A study reported that Tf binds iron at the N-terminal and C-terminal lobes with different affinities; binding to the C-terminal lobe favors the N-terminal lobe binding and the formation of diferric transferrin [4]. Diferric transferrin is the true ligand of both transferrin receptors (TfR1 and TfR2), while monoferric Tfs likely have a regulatory function [4]. (See 'Transferrin receptor' below.)

●Tf can be measured in the plasma using an enzyme-linked immunosorbent assay (ELISA) or turbidimetric method to determine the mg of the transferrin protein/dL of plasma.

●The total iron binding capacity of Tf (ie, TIBC) can be measured directly using iron binding methodology (ie, mg of iron binding capacity/dL of plasma), or it can be calculated by multiplying the results of the chemical or immunologic method by a conversion factor calculated by the individual laboratory, as follows:

Complete lack of transferrin is most likely incompatible with life. Hypotransferrinemia is a rare autosomal recessive disorder associated with low transferrin levels (<10 mg/dL), severe iron deficiency anemia with hypochromic, microcytic red cells, and iron overloading of the liver and other parenchymal organs [5].

High transferrin levels, as observed in higher altitude or in association with iron deficiency, is linked to an increased risk of thromboembolic events that may reflect a potentiating effect of transferrin on the activity of procoagulant factors thrombin and Factor XIIa [6].

Transferrin receptor — The gene for the TfR (TFRC) is located on the long arm of chromosome 3 (as is the gene for transferrin). TFRC codes for a homodimeric transmembrane protein (mol wt 94 kDa) that is found on most cells, most densely on erythroid precursors and placental cells.

There is a binding site for the transcription factor Stat5 in the first intron of the gene that encodes TfR [7]. Lethally irradiated mice that received a transplant of Stat5a/b^-/- liver cells developed microcytic, hypochromic anemia with reduced TfR gene expression [7]. In addition, TfR expression is induced by hypoxia via a hypoxia-inducible factor 1 (HIF1) binding site within the promoter [8].

Each TfR molecule can bind two diferric Tf molecules (four Fe³⁺ atoms), which it endocytoses after clustering on clathrin-coated pits. The iron is offloaded in acidified vacuoles and the apotransferrin-TfR complex is recycled to the cell surface, where apoTf is discharged and released back into the circulation. TfR can undergo proteasomal degradation by ubiquitin ligases, thereby controlling cellular iron accumulation [9]. The regulation of TfR expression via iron availability is mainly exerted by posttranscriptional mRNA stabilization. TfR mRNA has five 3' iron-response elements (IREs). As occurs in other iron genes, these IREs are targeted by iron-response proteins (IRPs), being stabilized in iron deficiency and degraded in iron overload. (See 'Systemic iron homeostasis' below.)

Important roles in several tissues have been illustrated in mouse models:

●Germline inactivation of TfR in mice is embryonically lethal due to severe anemia and abnormal central nervous system development. Haploinsufficiency of TfR causes microcytosis and a reduction of total body iron [10].

●Conditional heart inactivation of TfR in mice causes cardiomegaly, decreased cardiac function, failure of mitochondrial respiration, and early death [11].

●Conditional inactivation of TfR in the epithelial cells of the mouse intestine causes disruption of the epithelial barrier and early death. Since the phenotype was unresponsive to parenteral iron treatment, the authors suggested that TfR is implicated in the maintenance of the intestinal epithelium [12].

●Conditional inactivation of TfR in the liver indicates that TfR is not essential for basal hepatocellular iron supply but is indispensable for the fine-tuning of hepcidin expression in response to hepatocyte iron loading [13]. Another study on conditional inactivation of TfR in the liver showed that the most important function of TfR in hepatocytes is to sequester HFE in iron deficiency and to block hepcidin upregulation [14]. (See 'HFE' below.)

The only disease associated with pathogenic variants in TfR causes only mild anemia and leads to a type of combined immunodeficiency. It is due to homozygosity for a variant that disrupts the TfR internalization signal, strongly impairing endocytosis and underscoring the essential role of iron in B and T lymphocyte development and differentiation [15].

Serum (soluble) TfR (sTfR) is a product of membrane TfR that is released into the circulation by membrane proteases when TfR is not associated with its ligand, diferric transferrin, as occurs in iron deficiency [16]. Levels of sTfR measured in serum correlate directly with erythropoietic expansion [17]. sTfR levels can be a measure of iron deficiency and the bone marrow erythroid progenitors' need for iron. (See "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults", section on 'Iron studies (list of available tests)'.)

TfR is a cellular entry receptor for arenavirus and Plasmodium vivax, but it ameliorates cell entry by other viruses [18]. TfR recycling is utilized by influenza viruses to invade host target cells [19]. The same TfR ectodomain used by these pathogens to enter human cells interacts with the extracellular domain of H ferritin (ferritin heavy chain), as demonstrated by cryo-electron microscopy [20]. (See 'Ferritin' below.)

Ferritin — Ferritin is the cellular storage protein for iron. It is a huge, 24-subunit protein (mol wt 440 kDa) consisting of light chains (L ferritin, 20 kd, gene on chromosome 19) and heavy chains (H ferritin, 21 kd, gene on chromosome 11) that can store up to 4500 atoms of iron within its spherical cavity [21]. H ferritin possesses ferroxidase activity necessary for iron uptake by the ferritin molecule. L ferritin stabilizes the multimeric ferritin shell. A poly (rC) binding protein (PCBP1) appears to be required as a cytosolic chaperone to deliver iron to ferritin [22]. PCBP1 gene deletion in mice causes microcytic anemia [23]. PCBP1 and PCBP2 form homodimeric and heteromeric complexes with high iron binding affinity and distinct functions in cellular iron trafficking, with PCBP2 thought to affect iron delivery from DMT1 and to FPN1 [24,25].

Ferritin synthesis is subject to different levels of control, including DNA transcription via its promoter and mRNA translation via interactions with iron regulatory proteins with a single 5' IRE; this leads to increased ferritin translation in iron overload [26]. (See 'Iron regulatory proteins or iron-responsive element binding protein' below.)

Ferritin is an acute phase reactant, and, along with transferrin and the transferrin receptor, is a member of the protein family that orchestrates cellular defense against oxidative stress and inflammation [26,27]. The gene for H ferritin is activated by oxidative stress via an upstream enhancer antioxidant-responsive element (ARE) and by several proinflammatory cytokines, such as interleukin (IL)-1, IL-6, or tumor necrosis factor (TNF) alpha. (See "Acute phase reactants".)

Mice lacking H ferritin die early in gestation [28]. Mice heterozygous for H ferritin have slightly elevated tissue L ferritin and 7- to 10-fold more serum L ferritin than controls, although they do not have tissue iron overload [29]. These observations suggest that reduced H ferritin expression in humans might be responsible for cases in which high serum L ferritin is present in the absence of iron overload [29]. Conditional inactivation of H ferritin in duodenal cells in mice causes iron overload, indicating that ferritin in the gut is important in the control of iron absorption [30].

Much of the iron stored in ferritin is accessible for metabolic needs. Ferritin is degraded by lysosomes through a process called ferritinophagy, which requires the cargo protein NCOA4 [31,32]. This process is important for recovering intracellular iron when needed. Inactivation of NCOA4 in mice causes increased ferritin aggregates in spleen and liver macrophages and other organs [31,33]. In vitro downregulation of NCOA4 affects erythroid maturation and hemoglobin synthesis [23]. Ferritin within erythroid precursors may donate iron for heme synthesis, especially at the beginning of hemoglobin accumulation, at a time when the transferrin-transferrin receptor pathway is still insufficient [34]. However, the most important role of NCOA4 is in macrophage ferritinophagy, to recover iron stored in ferritin in iron deficiency [35] and in the gut where it contributes to iron absorption (see below).

When ferritin accumulates, it aggregates and is proteolyzed by lysosomal enzymes; it is then converted to an iron-rich, poorly characterized hemosiderin, which releases its iron slowly and is detected by the Prussian blue reaction, which is used in the common Perls staining for iron in bone marrow aspirates.

Ferritin measured clinically in plasma or serum is usually apoferritin, a noniron-containing molecule. The ferritin level generally reflects overall iron storage, with 1 ng of ferritin per mL indicating approximately 10 mg of total iron stores.

●An adult male with a ferritin level of 50 to 100 ng/mL has iron stores of approximately 500 to 1000 mg [36].

●A serum ferritin <15 ng/mL is 99 percent specific for making a diagnosis of iron deficiency, although iron deficiency diagnosis is accepted for serum ferritin levels <30 ng/mL. (See "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults", section on 'Iron studies (list of available tests)'.)

●An elevated serum ferritin in the absence of infection or inflammation may suggest the presence of an iron overload state. (See "Approach to the patient with suspected iron overload", section on 'CBC, LFTs, and iron studies'.)

●Ferritin levels may be extremely high in patients with hemophagocytic lymphohistiocytosis or certain rheumatologic disorders, as well as in acute infections with concomitant inflammation, such as in severe coronavirus disease 2019 (COVID-19) [37-39]. In such cases, the ferritin tends to be less glycosylated than normal. (See "Clinical features and diagnosis of hemophagocytic lymphohistiocytosis", section on 'Serum ferritin levels'.)

A separate ferritin (m-ferritin) is present within mitochondria and is the product of an intronless nuclear gene [21]. Its expression is increased in tissues with high numbers of mitochondria rather than in tissues involved in iron storage. M-ferritin likely protects mitochondria from oxidative damage [21].

The role and the origin of circulating ferritin as well as the pathways of cellular ferritin secretion are largely unknown [40]. Scavenger receptor, member 5 (Scara5), is a proposed receptor for circulating L ferritin and mediates the delivery of iron bound to ferritin to the developing kidney [41].The T cell immunoglobulin-domain and mucin-domain (TIM) protein Tim-2 on B lymphocytes and liver and kidney cells is proposed as an H ferritin receptor [42]. TfR binds H ferritin (ferritin heavy chain) independently from transferrin [20]. Whether this interaction may locally provide iron to cells when transferrin saturation is low remains to be demonstrated.

Iron regulatory proteins or iron-responsive element binding protein — The expression of proteins involved in cellular iron uptake and storage is regulated by the iron status of the cell.

Iron-regulatory proteins 1 and 2 (IRP1 and IRP2) are cytosolic RNA-binding proteins that bind to iron-responsive elements (IRE), consisting of a loop configuration of nucleotides that are located in the 5' or 3' untranslated regions of specific mRNAs encoding for iron and other genes (eg, ferritin, TfR, DMT1, ferroportin, the erythroid-specific form of delta-aminolevulinic acid synthase [eALAS], hypoxia inducible factor-2 alpha [HIF-2 alpha; HIF-2a]).

Binding of IRPs to their target sequences occurs when the cell is iron deficient; this has different effects according to whether the IRE position is at the 5' or the 3' UTR:

●When IRPs bind to the 5' IRE of ferritin, ferroportin, eALAS, or HIF-2 alpha, the rates of mRNA translation and protein biosynthesis are decreased.

●When IRPs bind to the 3' end of transcripts such as TfR or DMT1, the mRNA half-life is prolonged and rates of biosynthesis are increased (figure 2).

IRP1 and IRP2 sense the availability of metabolically active iron in the cell in different ways. When cellular iron levels increase, assembly with iron sulfur clusters changes IRP1 to cytoplasmic aconitase, and its binding ability is lost. Under the same conditions of cellular iron increase, IRP2 interacts with the iron-stabilized FBXL5 protein, which recruits an E3 ligase. This causes IRP2 ubiquitination and proteasomal degradation [43]. FBXL5 has an iron- and oxygen-sensing domain, representing an example of the iron-oxygen connection.

The net effect of these IRPs is that the iron overloaded state is characterized by increased production of ferritin (to permit adequate storage), ferroportin (to export excess iron), and eALAS (for iron consumption), as well as decreased production of TfR and DMT1 (to minimize iron uptake). These changes are reversed in iron deficiency, which is characterized by reduced ferritin and ferroportin and elevated TfR synthesis (figure 2).

Targeted deletion of the gene encoding IRP1 in a mouse model leads to no evident hematologic change in adulthood, while targeted deletion of the gene encoding IRP2 causes misregulation of iron metabolism, refractory microcytic anemia, and a neurodegenerative disease due to abnormal neuronal accumulation of iron [44].

These observations establish a major role for IRP2 in the regulation of iron uptake in erythroid cells. However, it has been shown that HIF-2 alpha, which has an IRE in the 5' UTR, is specifically controlled by IRP1 [45]. This demonstrates another example of the iron-hypoxia connection. IRP1^-/- mice, when young or iron deficient, develop polycythemia, pulmonary hypertension, and sudden death from hemorrhages, due to an inability to suppress the synthesis of HIF-2 alpha that stimulates both erythropoietin in the kidney and endothelin 1 in pulmonary endothelial cells [46,47].

There are differences in tissue-specific expression and specific targets of the two IRPs. (See "Molecular pathogenesis of congenital erythrocytoses and polycythemia vera", section on 'Oxygen sensor'.)

HFE — HFE, a product of the HFE gene on the short arm of chromosome 6, is an MHC class I-like protein present ubiquitously at low levels, with high-level expression in hepatocytes. The C282Y variant in the HFE gene is responsible for the vast majority of cases of hereditary hemochromatosis in adults. Iron overload is also noted in mice with HFE gene deletion [48]. Conditional deletion of HFE in the liver produces the phenotype of hemochromatosis [49]. In contrast, conditional deletion of HFE in duodenal cells or macrophages does not alter systemic iron metabolism [50].

The HFE protein is associated in a complex with TfR as a possible iron sensor. Diferric transferrin competes with this binding, releasing HFE from TfR when iron is abundant. Free HFE increases the response of hepcidin to iron, since patients with hemochromatosis due to an HFE mutation have low hepcidin levels and show a blunted response to oral iron [51]. (See 'Hepcidin' below.)

It has been reported that when HFE is free from TfR, it binds to TfR2, encoded by a gene mutated in type 3 hereditary hemochromatosis that is a sensor of the level of transferrin saturation [52]. (See "HFE and other hemochromatosis genes".)

However, HFE-TFR2 binding has been disputed [53]. Indeed, the diseases caused by pathogenic variants affecting HFE and TfR2 are different. Simultaneous gene deletion of both HFE and TfR2 in mice results in marked dysregulation of hepcidin and more severe iron overload than loss of the single molecules [54]. These observations suggest distinct functions for HFE and TfR2. According to an in vitro study, HFE stabilizes the BMP receptor ALK3, which is degraded when HFE is inactive [55]. HFE has been shown to interact with low density lipoprotein-receptor (LDL-R) in hepatocytes, thereby affecting systemic lipid homeostasis and atherosclerosis development [56].

Transferrin receptor 2 — Transferrin receptor 2 (TfR2), encoded by a gene on chromosome 7q22, is a member of the TfR family and is homologous to TfR1 but has no IRE elements and has a lower affinity for diferric transferrin than TfR1 [57]. It displays a restricted expression pattern, being present at high levels in hepatocytes, erythroid cells, and bone cells. TfR2 may bind diferric transferrin and is considered a sensor of Tf saturation [58].

Mutations of TfR2 cause a form of hereditary hemochromatosis. Mice with either germline or liver-conditional inactivation of TfR2 develop iron overload [59,60]. (See "HFE and other hemochromatosis genes", section on 'Transferrin receptor 2 (TFR2)'.)

TfR2 is expressed in immature erythroid cells, where it interacts with the erythropoietin receptor, stabilizing it on the cell surface [61]. In mice, conditional inactivation of TfR2 in the bone marrow causes erythrocytosis with normal erythropoietin levels. It has been shown that TfR2 modulates the erythropoietin sensitivity of erythroblasts by sensing iron deficiency through decreased diferric transferrin. Through its sensor activity, TfR2 may coordinate erythropoiesis with hepcidin synthesis [62]. TfR2 is also expressed in osteoclasts and osteoblasts, where it controls bone structure by modulating the BMP pathway [63].

Hemojuvelin — Hemojuvelin (HJV) is a glycosylphosphatidylinositol (GPI)-anchored protein that regulates hepcidin production in hepatocytes. It is the product of a gene on chromosome 1q21 and is homologous to RGM (Repulsive Guidance Molecule, also known as RGMc) expressed in the central nervous system. HJV is highly expressed in liver, skeletal muscles, and heart. Mutations of the gene encoding HJV (HFE2) produce a form of juvenile hemochromatosis with extremely low hepcidin levels [64]. (See "HFE and other hemochromatosis genes", section on 'Hemojuvelin (HJV)'.)

HJV is present in a membrane-associated form, bound to a glycosylphosphatidylinositol (GPI) anchor, and also as a soluble form with opposite effects on hepcidin activation [65,66]. As other RGMs, membrane HJV is a coreceptor for bone morphogenetic proteins (BMPs) and is essential in hepcidin activation. In vitro cleavage of HJV by furin in hypoxia and iron deficiency produces soluble HJV components and serves to downregulate hepcidin [67]. Its role in vivo is uncertain. In iron deficiency, membrane HJV is cleaved by the liver serine protease TMPRSS6 to attenuate the BMP signaling and suppress hepcidin [68]. (See 'Matriptase-2/TMPRSS6' below.)

Divalent metal transporter 1 and duodenal cytochrome b — The duodenal divalent metal transporter 1 (DMT1), encoded by the SLC11A2 gene on chromosome 12p13, is the major route for the uptake of non-heme iron from the intestinal lumen (figure 1). It was identified by positional cloning as the gene responsible for a form of microcytic anemia in mice with a missense mutation that had a marked impairment in intestinal iron transport [69].

Parallel functional studies in Xenopus oocytes found a single cDNA that stimulated iron transport. This divalent metal transporter protein (DMT1) was identical to Nramp2 [70]. The transporter also transports other heavy metals such as lead, zinc, and copper by an ATP and proton-dependent process. DMT1 is widely expressed, particularly in the proximal duodenum. Expression of the isoform containing an iron-responsive element is specifically upregulated in dietary iron deficiency [69] and hypoxia through the action of HIF-2 alpha [71]. DMT1 expression is greatest at the brush border of the apical pole of the enterocytes in the apical two-thirds of the villi, the major site of iron absorption.

Pathogenic variants in SLC11A2 are extremely rare and cause lifelong microcytic anemia, increased Tf saturation, and liver iron accumulation, but with low or only moderately high serum ferritin levels [72].

In iron import, DMT1 cooperates with duodenal cytochrome b (DYCTB), a membrane reductase that facilitates iron absorption as ferrous iron from the lumen of the duodenum (figure 1) [73].

Local intestinal hypoxia has an important role in increasing the expression of genes involved in iron transport, especially the luminal proteins DMT1 and DcytB and the basolateral iron exporter ferroportin (see 'Ferroportin' below). It has been shown that HIF-2 alpha binds to the DMT1 promoter and regulates DMT1 expression in duodenal cells; tissue-specific deletion of HIF-2 alpha in mouse enterocytes decreases intestinal iron absorption as well as the expression of DMT1 in enterocytes [71].

Four different isoforms of DMT1 have been described in cells and various tissues including intestinal cells, erythroid cells, macrophages, brain cells, or kidney cells, which have different organ-specific and subcellular functions for iron trafficking, an issue that still awaits clarification [74].

HIF-1 and 2 alpha — Hypoxia-inducible factor 2 alpha (HIF-2a), encoded by the EPAS1 gene, has an IRE in the 5' UTR and is controlled by IRP1 [45]. (See 'Iron regulatory proteins or iron-responsive element binding protein' above.)

HIF-2a is an essential mediator of iron absorption that cooperates with low hepcidin to maximize absorption in iron deficiency, anemia, and hypoxia [75]. In hypoxia, HIF-2a increases the expression of key genes (DMT1, DCYTB, and ferroportin) that contribute to enhanced iron absorption [71]. (See 'Intestinal iron absorption' below.)

A number of families with gain-of-function mutations in EPAS1 have been described with autosomal dominant erythrocytosis. (See "Molecular pathogenesis of congenital erythrocytoses and polycythemia vera", section on 'EPAS1 mutations'.)

HIF-1a is regulated by the von Hippel-Lindau (VHL) gene, and its dysregulation may contribute to some of the findings in VHL disease. (See "Molecular biology and pathogenesis of von Hippel-Lindau disease", section on 'Hypoxia-inducible factor 1 and 2'.)

Importantly, HIF-2a regulates the production of erythropoietin, which subsequently reduces hepcidin expression by mechanisms detailed below [76]. The stability of HIFs is controlled by prolylhydroxylases (PHDs) that mediate the hypoxic and iron deficiency responses for the regulation of erythropoietin; PHDs are inhibited under low iron conditions or hypoxia, thereby preventing HIF degradation by PHD and subsequently leading to erythropoietin expression [77]. (See "Regulation of erythropoiesis", section on 'Hypoxia and EPO expression'.)

Ferroportin — Ferroportin-1 (Ireg1, encoded by SLC40A1, formerly called SLC11A3, Mtp1) is an iron export protein found by positional cloning in mutant zebrafish with hypochromic anemia [78]. The human ferroportin gene maps to 2q32. Human ferroportin is highly expressed in the basal portion of placental syncytiotrophoblasts, the basolateral surface of duodenal enterocytes, macrophages, hepatocytes, cardiomyocytes, erythrocytes, and other cells [79-82].

Ferroportin functions as a major exporter of iron, transporting iron from mother to fetus, transferring absorbed iron from enterocytes into the circulation, and allowing macrophages to recycle iron from damaged and senescent red cells back into the circulation (figure 1).

In animal and human in vitro models, ferroportin is posttranscriptionally regulated by the amount of available iron, due to the presence of an IRE in the 5' UTR [83-86]. An isoform of ferroportin without the 5' IRE (ferroportin B) has been identified in duodenal mucosa. It appears to function to evade iron-mediated posttranscriptional regulation in conditions of iron deficiency [85]. The same isoform is expressed in erythroid cells and may function to "fine-tune" systemic iron usage [82,87].

The most important control of ferroportin is posttranslational, since ferroportin is downregulated through its interaction with hepcidin [80,88,89]. When hepcidin levels increase, hepcidin binds to ferroportin, occludes the central cavity that exports iron, and induces ferroportin internalization and lysosomal degradation [90]. This reduces the amount of iron that is released into the circulation from duodenal cells as well as macrophages [88]. Hepcidin binding to ferroportin is only effective when ferroportin is loaded with iron [91].

Besides posttranslational control by IRPs, ferroportin expression is increased by heme independently from iron [92]. Both iron and erythrophagocytosis (through heme increase) stimulate ferroportin transcription [93]. Ferroportin transcription is reduced in inflammation via Toll-like receptors and proinflammatory cytokine signaling [94].

Pathogenic variants (loss of function mutations) in the gene for ferroportin (SLC40A1) have been found in a number of kindreds with an autosomal dominant form of iron overload with a distinct phenotype called "ferroportin disease" [95-97]. In contrast, gain of function mutations make ferroportin resistant to the effect of hepcidin, producing a phenotype that overlaps with hemochromatosis. (See "HFE and other hemochromatosis genes", section on 'Ferroportin (SLC40A1; FPN1)'.)

The hepcidin-ferroportin axis may have important tissue-specific roles (eg, in the heart). Mice with conditional inactivation of ferroportin in cardiomyocytes develop cardiac iron overload and dilated cardiomyopathy, and they have decreased survival, strengthening the concept that these cells need to maintain an intact iron export system [93].

Hephaestin and ceruloplasmin — Hephaestin and ceruloplasmin are involved in iron export as multioxidases:

●Hephaestin is the product of an X-linked gene mutated in mice with sex-linked anemia, a disorder in which enterocytes are iron-loaded but efflux of iron through the basolateral membrane and into the plasma is inhibited [98]. Hephaestin cooperates with ferroportin in iron export in enterocytes. It has significant homology to the serum protein ceruloplasmin and has ferroxidase activity as well.

●Ceruloplasmin is a copper-containing protein encoded by the CP gene on chromosome 3q24-25; it is a ferroxidase required for efficient recycling of iron in the liver, reticuloendothelial system, and glial cells. It is present as a soluble form and as a membrane form bound to a GPI anchor in the central nervous system. Mutations of ceruloplasmin lead to aceruloplasminemia. (See 'Iron release from macrophages' below.)

FLVCR — A mammalian heme export protein (FLVCR, feline leukemia virus, subgroup C, receptor) has been postulated to protect developing erythroid cells from the toxicity of unbound cytoplasmic heme. Interference with this protein results in a loss of erythroid progenitors and severe anemia in experimental animals, while its inhibition in human erythroleukemia cells decreases heme export, impairs their erythroid maturation, and leads to apoptosis [99].

The heme exporter FLVCR1 regulates expansion and differentiation of committed erythroid progenitors by controlling intracellular heme accumulation in mice [100] and plays a crucial role in maintaining intestinal heme homeostasis [101].

ZIP14 — ZIP14, a metal transporter of zinc and manganese encoded by the SLC39A1 gene, has been shown to be important for nontransferrin-bound iron (NTBI) uptake by hepatocytes, pancreatic acinar cells [102], and beta-cells of the islets, while other NTBI transporters such as L-type calcium channels, DMT1, and ZIP8 are active in the heart and anterior pituitary [103].

Hepcidin — Hepcidin (also called liver-expressed antimicrobial peptide [LEAP-1] or hepcidin antimicrobial peptide [HAMP]) is an acute phase reactant with intrinsic antimicrobial activity [104-107]. It is encoded as a propeptide by a gene on 19q13. There are two isoforms of the mature peptide; hepcidin-25 has a central role in iron homeostasis, while the function of hepcidin-20, which lacks the five amino acid sequence crucial for iron regulation, is unknown.

Hepcidin production is induced or inhibited by multiple factors and signaling mechanisms that appear to follow a certain hierarchical concept [108-110]. Serum hepcidin levels have correlated directly with serum ferritin in healthy people. Hepcidin levels are highest in inflammation and are lowest in iron deficiency anemia and hypoxia. Reference ranges for hepcidin levels in healthy controls were noted to be wide in one study, but when rigorous criteria were required to eliminate individuals with inflammation or kidney or liver disease, variability was considerably less [108].

Two large studies have measured serum hepcidin in the general population. In the first study, hepcidin levels were analyzed using a competitive enzyme-linked immunosorbent assay in 2998 well-characterized participants from the Nijmegen Biomedical Study [111]. In the second report, serum hepcidin levels were measured by mass spectrometry in 1545 individuals from an Italian cohort [112].

●Both studies observed stable values in males across ages but strong variation in females, with low values in younger and higher levels in older females [112].

●There was a trend of increasing hepcidin concentrations during the day, although there was no evidence for a primary or secondary circadian variation in hepcidin levels [111].

●Levels were strongly associated with serum ferritin levels and were less strongly associated with C-reactive protein and total iron binding capacity (TIBC) in men and with TIBC, alanine aminotransferase, and glomerular filtration rate in women [111].

Because of the lack of a standardized assay, hepcidin testing is not ready for regular clinical use, although a number of test platforms are under active investigation [108]. Serum hepcidin levels may have diagnostic value in certain iron disorders and in diagnosis of specific forms of anemia [108].

Hepcidin is produced in many tissues, but the primary site of synthesis is in the liver [113-115]. Other tissues that produce hepcidin include:

●Macrophages in inflammation, adipocytes, retinal cells [116-118]

●Skin keratinocytes [119]

●Cardiomyocytes, where hepcidin has an essential cell-autonomous role in cardiac iron homeostasis [120]

Extrahepatic hepcidin has local effects and does not contribute to systemic regulation. It is usually activated by signals opposite of those that regulate hepatic hepcidin. As an example, hypoxia increases hepcidin in the heart to maintain sufficient iron in cardiomyocytes, while iron overload decreases cardiomyocyte hepcidin to avoid toxicity from excess intracellular iron [120].

Hepcidin is rapidly excreted by the kidney and reabsorbed in the proximal tubules; its levels increase in chronic kidney disease. It serves as an important mediator in the pathogenesis of the anemia of chronic disease [114,121-123]. It may decrease in chronic liver disease [108]. Its deficiency or inappropriate production explains the pathogenesis of iron overload in all types of hemochromatosis [124-126]. (See "HFE and other hemochromatosis genes".)

Its excessive inhibition by ineffective erythropoiesis explains iron overload in iron-loading anemias [127]. Hepcidin levels are also influenced by hormones and are especially inhibited by testosterone [128]. Hepcidin levels are increased by cell specific stress signals such as endoplasmatic reticulum stress, linking iron homeostasis to cellular protein quality control [129].

The following examples demonstrate the importance of hepcidin in iron balance and indicate that hepcidin plays a major role as a negative regulator of intestinal iron absorption and iron release from macrophages:

●Hepcidin knockout (KO) mice develop iron overload [105,130]. Liver-specific KO mice fully recapitulate the severe iron overload phenotype observed in the total hepcidin KO mice, demonstrating that the hepatocyte constitutes the predominant reservoir for systemic hepcidin and that the other tissues capable of synthesizing hepcidin are unable to compensate [131].

●In humans, pathogenic variants in the gene for hepcidin causes a rare form of juvenile hemochromatosis [132]. (See "HFE and other hemochromatosis genes", section on 'Hepcidin (HAMP)'.)

●Injection of hepcidin inhibited intestinal iron absorption in mice independent of their iron status. In other experiments, injection of a synthetic hepcidin in mice was associated with a rapid and prolonged reduction in serum iron, along with accumulation of hepcidin in ferroportin-rich organs (eg, liver, spleen, proximal duodenum) [89].

●Constitutive overexpression of hepcidin leads to severe iron deficiency anemia at birth [133] and slows dietary iron absorption and cycling through macrophages, resulting in iron-restricted erythropoiesis and a failure to respond to adequate erythropoietin levels [134].

●Overexpression of hepcidin inhibits the iron accumulation normally observed in HFE-deficient mice [124] and in mouse models of beta thalassemia [135].

The BMP SMAD signaling pathway that activates hepcidin in response to iron and the IL-6 signaling pathway that increases hepcidin in inflammation are discussed below. (See 'Iron sensing and signaling pathway' below.)

Hepcidin reduces iron absorption in the intestine and releases iron from macrophages via a mechanism that involves interactions with and inactivation of the iron export protein ferroportin [88,106]. The interaction occurs only with ferroportin bound to iron [91]; this explains the strong effect on cells such as macrophages and enterocytes, which export iron to plasma. Hepcidin also appears to regulate levels of nontransferrin-bound iron (NTBI) in a mouse model of bacterial infection; this may be the key mechanism by which hepcidin exerts its antimicrobial properties against circulating "siderophilic" bacterial strains that use NTBI and thrive in an iron-rich environment [107].

A distinct example of the antimicrobial role played by hepcidin occurs in necrotizing fasciitis caused by certain strains of group A Streptococcus. Hepcidin production is induced in the skin of the infected patients and has a protective role [119]. Hepcidin loss in keratinocytes of mice infected with the same microorganism blocks the production of the CXCL1 chemokine, reducing neutrophil recruitment and enabling infection to disseminate. This outcome can be reverted by the injection of exogenous hepcidin.

Another example of the antimicrobial function of hepcidin is illustrated in the gut, where hepcidin production by dendritic cells can lead to sequestering of iron from the microbiome, allowing mucosal repair of gut inflammation, such as would occur in inflammatory bowel disease [136].

Preclinical studies have shown the possibility of altering/inhibiting the function of hepcidin and its receptor ferroportin to alleviate some disorders of iron metabolism [137]. (See "Anemia of chronic disease/anemia of inflammation", section on 'Pathogenesis'.)

Clinical trials are underway in iron overload and other disorders. As an example, a hepcidin mimetic, rusfertide, was able to significantly reduce hematocrit in patients with polycythemia vera [138]. (See "Polycythemia vera and secondary polycythemia: Treatment and prognosis", section on 'Other agents'.)

BMP and BMP receptors — Bone morphogenetic proteins (BMPs; cytokines produced by endothelial cells) such as BMP6 and BMP2 activate hepcidin binding to BMP receptors and signaling through SMAD proteins [139].

BMP6-null mice have a phenotype resembling hereditary hemochromatosis, with reduced hepcidin expression and tissue iron overload, indicating the essential role of BMP6 in iron regulation in mammals [139,140]. BMP6 is highly expressed in liver sinusoidal endothelial cells (LSEC), and conditional inactivation of BMP6 in LSEC causes iron overload [141]. BMP2, produced by the same cells, regulates hepcidin; its conditional inactivation in LSEC and in endothelial cells in mice causes iron overload, defining its role in signaling for hepcidin, for which the mechanism still needs to be clarified [142].

The phenotype of BMP6 and BMP2 double knockout mice is not more severe than the individual knockouts, suggesting that the two BMPs cooperate in hepcidin activation, possibly through the formation of BMP heterodimers [143]. Also, BMP5 may contribute to hepcidin activation and systemic iron regulation, especially in situations where BMP6 availability is limited [144].

Matriptase-2/TMPRSS6 — The most powerful inhibitor of hepcidin expression is matriptase-2, which is encoded by TMPRSS6, the liver transmembrane protease, serine 6 gene. Matriptase-2 has been shown to exert its hepcidin regulatory effects by cleaving membrane hemojuvelin, a protein that normally signals to promote hepcidin expression and likely blocks other activators of hepcidin [68,145]. (See 'Hemojuvelin' above.)

Alternatively, the effect of TMPRSS6 has been proposed to be independent of cleavage and instead due to the simple association of the protease with multiple membrane substrates, including HJV [146].

●In agreement with hemojuvelin being the substrate of matriptase-2, mice that are double knockout for both TMPRSS6 and HJV have the same iron overload phenotype as that of HJV^-/- mice [147].

●The important role of TMPRSS6 in human iron deficiency is indicated by the observation that mutations in TMPRSS6 cause a rare autosomal recessive disorder characterized by iron deficiency anemia unresponsive to treatment with oral iron, but partially responsive to parenteral iron, a condition termed iron-refractory iron deficiency anemia (IRIDA) [148]. This condition is discussed in detail separately. (See "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults", section on 'Inherited disorders/IRIDA'.)

●Single nucleotide polymorphisms in the TMPRSS6 gene described in several populations appear to affect serum iron concentrations, hemoglobin levels, erythrocyte characteristics (eg, mean corpuscular volume [MCV], mean corpuscular hemoglobin [MCH]), and, indirectly, erythropoiesis [149-153]. These findings suggest a genetic susceptibility to the development of iron deficiency.

●Anti-TMPRSS6 compounds increase hepcidin and control iron overload in animal models of iron disorders or inherited anemias, such as beta thalassemia, and these compounds are being studied in clinical trials to increase hepcidin to alleviate iron loading [154].

●TMPRSS6 has also been involved in lipid metabolism, with a protective effect on high-fat diet-induced obesity in mice [155].

Erythroferrone — Erythroferrone (ERFE) is encoded by the ERFE gene (also called FAM132B or CTRP15). The protein is a member of the tumor necrosis factor alpha (TNF-alpha) family. Erythroferrone is stimulated by erythropoietin, a circulating hormone essential for the maturation and survival of erythroid progenitor cells to raise the red blood cell count in response to hypoxia or anemia. ERFE downregulates hepcidin to ensure iron supply.

The mechanism is mediated by blocking the binding of a subgroup of BMP proteins (including BMP6 and heterodimeric BMP6/BMP2) to their receptors [156,157]. ERFE behaves as a ligand trap for BMPs, to attenuate BMP signaling and acquire iron [158]. Antibodies raised against the N-terminal domain of ERFE prevent hepcidin suppression and are proposed as a therapeutic tool for iron loading disorders due to low hepcidin [158]. (See "Regulation of erythropoiesis", section on 'Erythropoietin'.)

Serum ERFE is low/undetectable in the absence of disease. Its levels are increased by erythropoietin in hypoxia or after hemorrhage [159]. ERFE function must be transient to allow acquisition of only the amount of iron needed to compensate erythropoiesis. Chronic production of ERFE induces iron overload and developmental abnormalities in transgenic mice [160]. ERFE is responsible for transfusion-independent iron overload in patients with ineffective erythropoiesis [2,159].

ERFE knockout mice are not anemic and recover from anemia after hemorrhage, suggesting the existence of other hepcidin inhibitors, such as hypoxia-induced platelet-derived growth factor BB or growth and differentiation factor 15 (GDF15) [161,162]. (See 'Iron and erythropoiesis' below.)

ERFE is produced in bone (by both osteoblasts and osteoclasts) and plays an important role in the erythropoiesis-iron-bone circuitry [163].

ERFE expression in osteoblasts is independent of erythropoietin and contributes to liver hepcidin suppression. ERFE^-/- mice have increased bone turnover with both enhanced resorption and formation. ERFE shows an osteoprotective effect by modulating BMP signaling in osteoblasts and limiting osteoclastogenesis. It prevents excessive bone loss during expanded erythropoiesis in beta thalassemia [163].

Fibrinogen-like protein 1 — FGL1 (fibrinogen-like protein 1) is produced by the liver in hypoxia and is another inhibitor of the BMP-SMAD pathway. At variance with erythroferrone, FGL1 production is independent of erythropoietin activation. Both inhibitors act by sequestering BMPs, but the function of FGL1 is delayed compared with ERFE [164,165]. FGL1 knockout mice have no hematologic phenotype but are overweight and show reduced glucose tolerance.

SYSTEMIC IRON HOMEOSTASIS

Body iron content — The normal iron content of the body is approximately 3 to 4 grams. It exists in the following forms (figure 3):

●Hemoglobin in circulating red cells and developing erythroblasts – approximately 2.0 to 2.5 g

●Iron-containing proteins (eg, myoglobin, cytochromes, enzymes) – 300 to 400 mg

●Plasma transferrin-bound iron – 3 to 4 mg

●The remainder is storage iron in the form of ferritin or hemosiderin

Males have approximately 0.7 to 1.0 g of storage iron (mostly in liver, spleen, muscle, and bone marrow). Females have less storage iron (0.3 g), depending upon the extent of menses, pregnancies, deliveries, and iron intake. (See "Anemia in pregnancy", section on 'Iron deficiency'.)

Only a small amount of iron (1 to 2 mg) enters and leaves the body via enterocyte or keratinocyte desquamation on a daily basis. Most iron is recycled from the breakdown of old red blood cells by macrophages of the reticuloendothelial system.

To avoid iron-mediated free radical toxicity, iron is always bound to proteins in the less reactive ferric iron (Fe³⁺) form. Essentially all circulating iron is bound to transferrin. This chelation renders the iron soluble and prevents toxicity. A toxic species of iron (nontransferrin-bound iron [NTBI]) appears in the circulation in iron overload when transferrin saturation exceeds 60 to 70 percent.

Iron homeostasis is regulated strictly at the level of intestinal iron absorption and release of iron from macrophages. (See 'Intestinal iron absorption' below and 'Iron release from macrophages' below.)

Intestinal iron absorption — The gastrointestinal mucosa plays a major role in regulating iron absorption, which varies according to the form of iron in the diet.

A Western diet contains approximately 15 mg of iron per day. Some of this is heme iron, of which approximately 30 percent is promptly absorbed, likely via its own transport system. A candidate heme iron transporter, named heme carrier protein 1, has been found in the apical brush border membrane of duodenal enterocytes in mice [166]. However, the role in heme transport was subsequently dismissed [167].

The remaining non-heme iron, accounting for almost all of the iron in the diet in non-Western countries, is poorly absorbed, with less than 10 to 20 percent being taken into the mucosal cells (figure 3) [168]. Iron absorption is increased in iron-deficient states and may be different with food sources relative to oral iron supplements [169].

The table summarizes various factors that can impact iron absorption (table 1). As examples:

●Dietary sources of heme iron (fish, poultry, and meat) have a higher bioavailability than do non-heme (vegetable) sources (30 versus <10 percent).

●Intraluminal factors and genetic polymorphisms of iron metabolism genes can affect absorption.

●Ascorbic acid and meat sources enhance the absorption of nonanimal sources of iron such as cereal, breads, fruits, and vegetables, whereas tannates (teas) and bran foods rich in phosphates and phytates inhibit iron absorption [170-174].

●Certain medications can impact iron absorption, as shown for reduced iron uptake in individuals treated with proton pump inhibitors [175]. (See "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults", section on 'Medications'.)

●The composition of the gut microbiota can affect iron absorption, as some specific bacteria can produce metabolites that block enterocyte iron import [176]. (See "Treatment of iron deficiency anemia in adults", section on 'Dosing and administration (oral iron)'.)

Molecular mechanisms of intestinal heme absorption are unclear. The proposed heme carrier protein 1, which is highly expressed in the gut and stimulated by hypoxia [166], functions as a folate transporter [167]. A heme exporter, feline leukemia virus receptor 5 (FLVR5), is expressed in enterocytes, macrophages, and erythroblasts with the likely function of exporting heme excess [99]. (See 'FLVCR' above.)

Iron in food is prominently ferric (Fe³⁺), which is poorly soluble above a pH of 3 and is therefore poorly absorbed. In comparison, ferrous iron (Fe²⁺) is more soluble, even at the pH of 7 to 8 seen in the duodenum. As a result, it is more easily absorbed.

Ferrous iron is taken up at the mucosal side by the intestinal transporter, DMT1. In this process, duodenal cytochrome b (DcytB) reduces ferric iron to ferrous iron. Transcription of both DMT1 and DCYTB is stimulated by HIF-2a in the hypoxic environment of intestinal mucosa [177,178]. The microbiota-derived products 1,3-diaminopropane and reuterin reduce HIF-2a stability and reduce iron absorption, pointing to the importance of the gut microbiota for systemic iron homeostasis [176].

When iron enters the cell, it binds to cytosolic iron chaperones PCBP1 and PCBP2 The function of these peptides is essential for proper iron absorption and cellular iron release [179,180]. Iron is then transported to either ferritin or ferroportin (the duodenal basolateral membrane iron exporter), oxidized to the ferric form, and loaded onto transferrin. This oxidation process involves hephaestin, a homologue of ceruloplasmin, a known ferroxidase.

Mucosal cells, in addition to the cellular mechanisms noted above, respond to specific physiologic signals.

●In hypoxia, hepcidin is downregulated to allow increased iron export through ferroportin, while HIF-2a increases the expression of key genes (DMT1, DCYTB, and ferroportin) that enhance iron absorption [71]. (See 'Hepcidin' above.)

●The rate of iron absorption is appropriately enhanced when iron stores are reduced or absent (figure 1).

●HIF-2a is an essential mediator of iron absorption that cooperates with low hepcidin to increase absorption in iron deficiency, anemia, and hypoxia [75]. (See 'HIF-1 and 2 alpha' above.)

●Intestinal macrophages determine enterocyte iron transfer to the circulation by controlling the levels of transferrin in lamina propria cells [181]. This mechanism of iron absorption can be modulated by injection of apo-transferrin [182].

●HIF-2 alpha also controls NCOA4 and stimulates ferritinophagy as an additional mechanism to increase iron export to plasma, which contributes to iron overload in hemochromatosis [183].

●The degree of erythropoiesis has an indirect effect, as iron absorption is increased when driven by increased erythropoiesis, a process mediated by hepcidin suppression.

Iron absorption is especially increased in disorders that cause ineffective erythropoiesis, such as beta thalassemia, dyserythropoietic and sideroblastic anemia, and some myelodysplastic syndromes.

On the other hand, intestinal cells can hold onto iron in the iron-replete state; this iron is lost when the mucosal cells are sloughed.

All of these regulatory processes are mainly mediated by hepcidin through its interaction with ferroportin [1]. This function of hepcidin appears to be especially important when there are competing needs for iron, such as when anemia, iron deficiency, and infection coexist, as shown in children with malaria [184]. (See 'Hepcidin' above and 'Ferroportin' above.)

Iron absorption is decreased in conditions of iron excess through hepcidin increase, a process lost in hereditary hemochromatosis. (See "Clinical manifestations and diagnosis of hereditary hemochromatosis", section on 'Pathophysiology'.)

Transferrin saturation — Circulating transferrin is normally approximately one-third saturated with iron (ie, Fe ÷ TIBC = 1/3, when both are expressed as micrograms of iron per 100 mL of plasma) [185,186]. The formula to change transferrin levels from mg of protein to micrograms of iron binding capacity can be used if this information has not already been provided by the laboratory. (See 'Transferrin' above.)

Conditions in which transferrin saturation (TSAT) is reduced (<20 percent) include those in which the supply of iron to the plasma from the macrophage and other storage sites is reduced. These include:

●Iron deficiency anemia

●Anemia of chronic disease/anemia of inflammation

●In some patients with a specific pathogenic variant in the SLC40A1 gene, which encodes ferroportin (see 'Ferroportin' above and "HFE and other hemochromatosis genes", section on 'Ferroportin (SLC40A1; FPN1)')

Conversely, TSAT is increased (>45 to 50 percent) in conditions in which the supply of iron is excessive or is greater than the current demand. These include:

●Most cases of hereditary and acquired hemochromatosis

●Aplastic anemia, bone marrow suppression

●Sideroblastic anemias

●Ineffective erythropoiesis

●Heavily transfused patients

●Liver disease with reduced transferrin synthesis

●Monoclonal immunoglobulin with antitransferrin activity (rare) [187]

The use of TSAT to evaluate disease states is discussed separately:

●Iron deficiency – (See "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults", section on 'Iron studies (list of available tests)'.)

●Iron overload – (See "Approach to the patient with suspected iron overload", section on 'CBC, LFTs, and iron studies' and "Clinical manifestations and diagnosis of hereditary hemochromatosis", section on 'Diagnostic evaluation'.)

Iron loss — There is no physiologic mechanism of regulated iron excretion.

Iron is lost in sweat, shed skin cells, and some gastrointestinal loss at a rate of approximately 1 to 2 mg/day (figure 3).

●The Western diet of an adult male has 1 to 2 mg of heme iron and 10 to 15 mg of other iron. If 30 percent of the heme iron and 10 percent of the other iron is absorbed, then the total rate of iron absorption is 1 to 2 mg/day [185]. Thus, a male can easily stay in iron balance and even build up iron stores.

●On the other hand, a female with an additional menstrual iron loss of 1 to 2 mg/day generally has lower iron stores than a male and is always delicately poised to become iron deficient.

There is abundant expression of DMT1 in the proximal tubule and collecting ducts of the kidney [69]; these cells also express TFR1, ZIP14, and ZIP8 [188].

The iron handling by the kidney in terms of uptake and export is not fully elucidated. Iron is filtered in the glomerulus and reabsorbed in the tubules both as transferrin-bound and nontransferrin-bound iron (NTBI). The expression of ferroportin on the apical membrane of proximal tubules suggests a possible contribution of iron excretion in iron overload [189]. Increased ferroportin expression by renal tubular epithelial cells can prevent iron-mediated kidney injury by promoting iron reabsorption in a murine model of malaria infection [190].

Iron release from macrophages — Approximately 20 to 25 mg of iron are released daily from the breakdown of senescent red cells in macrophages (figure 3).

Heme from hemoglobin released from phagocytosed red cells is catabolized by microsomal heme oxygenase to biliverdin and carbon monoxide, and the resulting iron is released to the circulation through ferroportin or stored in ferritin according to the body's needs and the local hepcidin concentration [191]. Thus, hepcidin coordinates both duodenal iron absorption as well as macrophage iron release. (See 'Hepcidin' above.)

Upon its release from ferroportin, ferrous iron is oxidized to the ferric form and loaded onto transferrin. The oxidation process involves ceruloplasmin, a known copper-dependent multioxidase. This could explain the iron overload that is seen in aceruloplasminemia, an autosomal recessive disorder of iron metabolism characterized by anemia, diabetes, retinal degeneration, and neurologic symptoms. Affected patients have inherited variants in the ceruloplasmin gene (CP), along with progressive parenchymal iron accumulation in conjunction with strongly decreased circulating ceruloplasmin [192]. (See "Bradykinetic movement disorders in children", section on 'Neurodegeneration with brain iron accumulation'.)

Iron sensing and signaling pathway — Hepcidin is upregulated in response to increased circulating and body iron levels (figure 1), inflammation, infection, endotoxins, and ER-stress; it is downregulated following conditions characterized by increased levels of erythropoietin including hypoxia, anemia, iron deficiency, and ineffective erythropoiesisall , but also by steroid hormones or alcohol consumption (figure 4) [129,161,162,193-197].

Hepcidin has also been shown to be regulated by a transferrin-dependent pathway in the zebrafish embryo [198]. In humans, hepcidin increases following the absorption of amounts of iron sufficient to acutely increase transferrin saturation [113]. (See 'Transferrin saturation' above.)

This increase is blunted in patients with hereditary hemochromatosis due to HFE variants and abolished in patients with TFR2-related hemochromatosis [51].

Transcription of hepcidin in response to increased plasma or tissue iron is mediated by bone morphogenetic proteins (BMP), requires hemojuvelin (HJV) as a BMP coreceptor, and is SMAD-dependent (figure 4) [66,199]. Increased hepcidin production is seen in acute and chronic inflammation, mediated by lipopolysaccharide, interleukin (IL)-6, and IL-1-beta (figure 4) [121], and is an essential component of anemia of acute inflammation and anemia of chronic diseases [123,200].

Iron and erythropoiesis — Most serum iron, estimated at approximately 80 percent, is used in the bone marrow for red blood cell (RBC) production. The crosstalk of iron with erythropoiesis is essential to maintain iron balance [127,201,202].

Since most iron is used by maturing erythroid cells, several conditions, including iron deficiency anemia, hypoxia, and erythropoietic expansion, decrease hepcidin production to favor iron acquisition [127]. Suppression of hepcidin in hypoxia occurs indirectly through erythropoietic expansion [203].

Ferrokinetic studies provided earlier information about erythropoietic control of iron absorption. The methods consisted of intravenously injecting a tracer label of Fe-59 bound to plasma transferrin. Three measurements were then made: the disappearance rate of Fe-59 from plasma over a period of minutes provided an index of beginning erythropoiesis; the appearance of Fe-59 by scanning over the spleen, liver, and bone marrow indicated the erythropoietic sites; and the total amount of iron appearing in RBCs in the circulation 7 to 10 days later provided a measure of effective erythropoiesis.

As an example, in severe beta thalassemia, the very rapid disappearance of plasma iron indicated a massive onset of erythropoiesis, and the low incorporation of injected iron in RBCs (20 to 30 percent, versus a normal value of >80 percent) showed that the increased erythropoiesis was severely ineffective. The existence of a regulator of iron absorption produced by erythroblasts was first proposed based on ferrokinetic studies [204]. It was subsequently documented that both effective and ineffective erythropoiesis upregulate iron acquisition by suppressing hepcidin.

Erythroferrone (ERFE) produced by the erythropoietin-stimulated erythroid precursors plays a major role as erythroid regulator in hepcidin inhibition [205]. In vitro, ERFE binds and sequesters different BMPs, including BMP6 and heterodimers BMP6/BMP2, attenuating the SMAD signaling [156,157]. (See 'Erythroferrone' above.)

However, full hepcidin inhibition by ERFE requires an attenuated BMP pathway, strengthening the complex process of hepcidin inhibitory control [206]. Also, ERFE produced by bone cells contributes to modulating the BMP-SMAD signaling cascade [163]. The existence of other hepcidin inhibitor(s) was hypothesized based on the partial suppression of hepcidin in ERFE knockout mice. This observation led to the discovery of FGL1, produced by murine hepatocytes in hypoxia [164]; however, it is also in line with the systemic function of PDGF-BB and GDF-15 as negative hepcidin regulators [161,162]. (See 'Fibrinogen-like protein 1' above.)

A role for PDGF-BB as a hepcidin inhibitor in hypoxia has also been demonstrated in humans [162].

In iron sensing, an important role is played by TFR2. TFR2, expressed in both bone marrow and hepatocytes, is stabilized on the plasma membrane by binding diferric transferrin and removed from the membrane in the absence of diferric transferrin. Sensing serum iron availability, it coordinates hepcidin production with the erythropoiesis response to erythropoietin [62]. (See 'Transferrin receptor 2' above.)

In iron deficiency, the BMP pathway is attenuated. In addition to cleavage of HJV by TMPRSS6, a study has described a role for epigenetic downregulation of the pathway [207].

The iron sensing and signaling pathway involving hepcidin is complex and not fully elucidated [2,106,208]. The proposed model is shown in the figure (figure 4).

●BMP6 produced by LSEC in iron overload conditions binds and activates its own receptors (BMPRs) in the presence of the coreceptor HJV. In agreement with this model, BMP6 inactivation in mice causes severe iron overload with low hepcidin [139,140]. BMP6 signals prevalently through ALK2.

●BMP2 participates in hepcidin activation, likely setting up the basal hepcidin levels. It does not respond significantly to iron increase and signals prevalently through ALK3.

●Inactivation of BMP2 in mice causes severe iron overload that is not compensated by BMP6.

●BMPs bind to BMPRs. Conditional inactivation of the BMPR ALK2 and ALK3 in mouse liver causes iron overload of different severity [209]. Signal transduction of BMPs occurs through SMAD proteins; conditional inactivation of SMAD4 in mouse liver causes liver iron accumulation and the inability to upregulate hepcidin, findings that are similar to those seen in hemochromatosis [210].

●Whether and how HFE and TFR2 make a complex in the presence of increased diferric transferrin that cooperates with the BMP-HJV-SMAD pathway for hepcidin activation is not fully defined. In Hfe^-/- mice, the BMP pathway is indeed less active, and treatment with BMP6 appears to ameliorate iron overload [211].

●Inflammatory cytokines, especially IL-6 (and IL1-beta) activate hepcidin transcription through interaction with the IL-6 receptor and signal transduction through STAT3 [66,113,212].

●There is a crosstalk between the two pathways of hepcidin activation (inflammation- and iron-dependent) as shown by improved hepcidin control by compounds that inhibit the BMP-SMAD pathway in inflammation [213].

●In iron deficiency, TMPRSS6 seems to be the major regulator of BMP-SMAD and hepcidin suppression. When erythropoiesis is expanded by erythropoietin, ERFE plays a major role in downregulating hepcidin expression, both in effective and ineffective erythropoiesis. FGL1, PDGF-BB, and GDF-15 contribute to suppressing hepcidin in hypoxia.

IRON AND THE BRAIN — Iron is essential for neuronal function, neurotransmitter synthesis, and myelinization [214]. Iron is taken up from plasma transferrin by the endothelial cells of the blood-brain barrier, and then it is released through ferroportin to the cerebral fluid, where it is bound to local transferrin.

Iron regulation, distribution, and exchange between neurons and glial cells remain incompletely understood. Neurons express high levels of the transferrin receptor (TfR), while glial cells mainly take up nontransferrin-bound iron (NTBI) and express high levels of ferritin. Some areas of the brain, such as the globus pallidus and substantia nigra, accumulate ferritin-iron over time, perhaps as a form of iron storage.

Specific regions of the brain are susceptible to iron accumulation in rare disorders collectively called neurodegeneration with brain iron accumulation (NBIA). (See "Bradykinetic movement disorders in children", section on 'Neurodegeneration with brain iron accumulation'.)

Neurodegenerative disorders of aging, such as Parkinson disease, Alzheimer disease, Huntington disease, amyotrophic lateral sclerosis, and Friedreich ataxia have been associated with excessive intracellular free iron and ferroptosis, a form of cell death due to iron-induced lipid peroxidation [215-217].

●In Friedreich ataxia (FA), an autosomal recessive progressive neurodegenerative disorder mainly affecting the spinal cord and cerebellum, reduced expression of frataxin results in toxic iron accumulation [218]. This causes mitochondrial oxidative stress and dysfunction along with an abnormality in iron sulfur-cluster formation [219]. Nonetheless, evidence also suggests a systemic and cellular iron deficiency signature in FA patients that was positively associated with genetic disease severity, challenging the idea of iron chelation therapy [220]. (See "Friedreich ataxia".)

●In Parkinson disease and Alzheimer disease, it is still debated whether spatiotemporal accumulation of iron is a causative disease trigger or a secondary effect due to cell degradation [221]. The neurotransmitter dopamine has been identified as an iron chaperone that can shuttle iron across membranes and affect cellular iron homeostasis [222]. Dopamine deficiency likely results in iron misdistribution in the brain. (See "Epidemiology, pathogenesis, and genetics of Parkinson disease", section on 'Basal ganglia circuits' and "Epidemiology, pathogenesis, and genetics of Parkinson disease", section on 'Iron metabolism' and "Epidemiology, pathology, and pathogenesis of Alzheimer disease", section on 'Pathogenesis'.)

In Parkinson disease-related restless legs syndrome, a frequent sensorimotor disorder in which iron deficiency has been causally related to disease severity, treatment with L-dopamine results in improvement of symptoms [223,224], in parallel with improvement of mitochondrial iron deficiency and mitochondrial functionality [225]. (See "Clinical manifestations of Parkinson disease", section on 'Sleep disorders'.)

Iron accumulation has been observed in dopaminergic neurons in individuals with Parkinson disease [226]. Based on this, a 2022 study investigated the efficacy of the iron chelator deferiprone in patients with newly diagnosed Parkinson disease; however, this found worsening of symptoms, likely due to reduced cellular and mitochondrial iron in neurons [227].

An improved understanding of cellular and spatiotemporal iron fluxes in the brain and their underlying regulatory processes will be essential to estimate the true impact of abnormalities in iron regulation in neurological disorders and may offer an opportunity to develop targeted interventions. Of interest, iron uptake mechanisms by neurons or adjacent endothelial cells may be used to facilitate the transfer of therapeutics across the blood-brain barrier, as shown for the brain introduction of biomolecules for the treatment of lysosomal enzyme defect disease, mucopolysaccharidosis II, which could be shuttled via transferrin receptor-mediated transcytosis [228].

SUMMARY

●Proteins involved in iron regulation – The regulation of iron metabolism involves the interaction of a number of specific proteins, as well as the interplay between iron absorption from the gastrointestinal tract, recycling of iron from red cells at the end of their life span, release of iron stores from the monocyte-macrophage system, and iron loss from the body (figure 1 and figure 2 and figure 3). (See 'Role of specific proteins' above.)

●Intestinal iron absorption – Intestinal iron absorption is tightly controlled, since there is no physiologic means of excreting iron from the body once it is absorbed. Factors that affect absorption are summarized in the table (table 1). HIF-2a and hepcidin are the most important regulators of apical and basolateral transporters, respectively. (See 'Intestinal iron absorption' above.)

•Role of ferroportin – Ferroportin functions as a major exporter of iron, transporting iron from mother to fetus, transferring absorbed iron from enterocytes into the circulation, and allowing macrophages to recycle iron from senescent and damaged red cells back into the circulation. (See 'Ferroportin' above and 'Iron release from macrophages' above.)

•Role of hepcidin – Hepcidin occludes the iron export channel of ferroportin and downregulates ferroportin posttranslationally. This is the most important control of iron cycling and systemic iron homeostasis (figure 4). (See 'Hepcidin' above.)

●Iron loss – Iron is lost in sweat, shed skin cells, and perhaps some gastrointestinal loss at a rate of approximately 1 mg/day. Females with additional menstrual iron loss of 1 to 2 mg/day generally have lower iron stores than males or postmenopausal females and are delicately poised to become iron deficient. (See 'Iron loss' above and "Causes and diagnosis of iron deficiency and iron deficiency anemia in adults", section on 'Search for source of blood and iron loss'.)

●Iron in the brain – Iron is essential in the brain, but its regulation and distribution remain incompletely understood. Abnormal iron accumulation has been seen in Parkinson disease, Alzheimer disease, and neurodegeneration with brain iron accumulation (NBIA), but it remains unclear whether iron accumulation is a cause of neuronal dysfunction or a consequence. Greater understanding might offer the opportunity to develop targeted interventions. (See 'Iron and the brain' above.)

ACKNOWLEDGMENTS — UpToDate gratefully acknowledges Stanley L Schrier, MD, who contributed as Section Editor on earlier versions of this topic review and was a founding Editor-in-Chief for UpToDate in Hematology.

The UpToDate editorial staff also acknowledges the extensive contributions of William C Mentzer, MD, to earlier versions of this and many other topic reviews.