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Hepcidin-Ferroportin Interaction Controls Systemic Iron Homeostasis

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Despite its abundance in the environment, iron is poorly bioavailable and subject to strict conservation and internal recycling by most organisms. In vertebrates, the stability of iron concentration in plasma and extracellular fluid, and the total body iron content are maintained by the interaction of the iron-regulatory peptide hormone hepcidin with its receptor and cellular iron exporter ferroportin (SLC40a1). Ferroportin exports iron from duodenal enterocytes that absorb dietary iron, from iron-recycling macrophages in the spleen and the liver, and from iron-storing hepatocytes. Hepcidin blocks iron export through ferroportin, causing hypoferremia. During iron deficiency or after hemorrhage, hepcidin decreases to allow iron delivery to plasma through ferroportin, thus promoting compensatory erythropoiesis. As a host defense mediator, hepcidin increases in response to infection and inflammation, blocking iron delivery through ferroportin to blood plasma, thus limiting iron availability to invading microbes. Genetic diseases that decrease hepcidin synthesis or disrupt hepcidin binding to ferroportin cause the iron overload disorder hereditary hemochromatosis. The opposite phenotype, iron restriction or iron deficiency, can result from genetic or inflammatory overproduction of hepcidin.
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International Journal of
Molecular Sciences
Review
Hepcidin-Ferroportin Interaction Controls Systemic
Iron Homeostasis
Elizabeta Nemeth 1and Tomas Ganz 2, *


Citation: Nemeth, E.; Ganz, T.
Hepcidin-Ferroportin Interaction
Controls Systemic Iron Homeostasis.
Int. J. Mol. Sci. 2021,22, 6493.
https://doi.org/10.3390/ijms22126493
Academic Editor: Wojciech Bal
Received: 2 June 2021
Accepted: 14 June 2021
Published: 17 June 2021
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1Department of Medicine, University of California, Los Angeles, CA 90095, USA; enemeth@mednet.ucla.edu
2Departments of Medicine and Pathology, University of California, Los Angeles, CA 90095, USA
*Correspondence: tganz@mednet.ucla.edu
Abstract:
Despite its abundance in the environment, iron is poorly bioavailable and subject to
strict conservation and internal recycling by most organisms. In vertebrates, the stability of iron
concentration in plasma and extracellular fluid, and the total body iron content are maintained by
the interaction of the iron-regulatory peptide hormone hepcidin with its receptor and cellular iron
exporter ferroportin (SLC40a1). Ferroportin exports iron from duodenal enterocytes that absorb
dietary iron, from iron-recycling macrophages in the spleen and the liver, and from iron-storing
hepatocytes. Hepcidin blocks iron export through ferroportin, causing hypoferremia. During
iron deficiency or after hemorrhage, hepcidin decreases to allow iron delivery to plasma through
ferroportin, thus promoting compensatory erythropoiesis. As a host defense mediator, hepcidin
increases in response to infection and inflammation, blocking iron delivery through ferroportin to
blood plasma, thus limiting iron availability to invading microbes. Genetic diseases that decrease
hepcidin synthesis or disrupt hepcidin binding to ferroportin cause the iron overload disorder
hereditary hemochromatosis. The opposite phenotype, iron restriction or iron deficiency, can result
from genetic or inflammatory overproduction of hepcidin.
Keywords: iron deficiency; iron overload; hemochromatosis; anemia; metal transport
1. The Roles of Iron
Iron is an essential trace element for nearly all living organisms. Even though iron
is one of the most abundant elements in the Earth’s crust, ferric iron, an oxidized form
common in the oxygen-rich environment on the surface of the Earth, is poorly soluble
and difficult to access by most life forms. Accordingly, biological organisms have evolved
mechanisms that conserve iron and recycle it internally. In adult humans, total body iron
content is approximately 3–4 g, whereas normal daily losses are only 1–2 mg. To remain in
iron balance, healthy humans must absorb a similar amount of iron from their diets.
The ability of iron to donate or accept an electron in cellular and extracellular envi-
ronments is what makes it a versatile catalytic component of many enzymes involved in
energy-producing bioreactions and critical biosynthetic pathways, as well as in enzymes
that generate reactive oxygen species for host defense. Iron also coordinates oxygen in
hemoglobin and myoglobin, molecules involved in oxygen transport and its cellular storage.
In biological systems, iron carries out its function in association with three common types
of moieties: iron coordinated by protein side chains, iron complexed within the porphyrin
ring of heme, and iron within iron-sulfur clusters. Outside of these controlled chemical
environments, iron displays promiscuous reactivity that can damage cells and tissues.
2. Iron Compartments and Flows
Biological organisms closely regulate intracellular and extracellular iron concentra-
tions, navigating between the twin threats of inadequate iron supply that would limit
critical functions, and uncontrolled iron excess that could be toxic to the organism. Sys-
temic iron homeostasis, best understood in humans and laboratory rodents, is maintained
Int. J. Mol. Sci. 2021,22, 6493. https://doi.org/10.3390/ijms22126493 https://www.mdpi.com/journal/ijms
Int. J. Mol. Sci. 2021,22, 6493 2 of 13
by regulating intestinal iron absorption, the concentration of iron in blood plasma and
extracellular fluid, the distribution of iron among organs and tissues, and the amount
of iron kept in stores. Iron dyshomeostasis can manifest as total body iron deficit (iron
deficiency) or excess (iron overload), as well as iron maldistribution among tissues in
which individual tissues or organs may become iron-deficient or iron-overloaded. Such
iron disorders may be caused by genetic lesions that directly impair iron regulation, or
conditions that impact iron regulation indirectly.
Although all cells in a multicellular organism contain iron, systemic iron homeostasis
is primarily affected by the following compartments: the erythron (red blood cells and
their precursors in erythropoietic organs), two types of stores (hepatocytes of the liver
and macrophages of the spleen and the liver), blood plasma which moves iron between
tissues and organs, and absorptive enterocytes in the duodenum through which iron
enters the body, ordinarily to replace small losses from the body caused by shedding of
iron-containing cells (Figure 1).
Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 2 of 13
2. Iron Compartments and Flows
Biological organisms closely regulate intracellular and extracellular iron concentra-
tions, navigating between the twin threats of inadequate iron supply that would limit crit-
ical functions, and uncontrolled iron excess that could be toxic to the organism. Systemic
iron homeostasis, best understood in humans and laboratory rodents, is maintained by
regulating intestinal iron absorption, the concentration of iron in blood plasma and extra-
cellular fluid, the distribution of iron among organs and tissues, and the amount of iron
kept in stores. Iron dyshomeostasis can manifest as total body iron deficit (iron deficiency)
or excess (iron overload), as well as iron maldistribution among tissues in which individ-
ual tissues or organs may become iron-deficient or iron-overloaded. Such iron disorders
may be caused by genetic lesions that directly impair iron regulation, or conditions that
impact iron regulation indirectly.
Although all cells in a multicellular organism contain iron, systemic iron homeostasis
is primarily affected by the following compartments: the erythron (red blood cells and
their precursors in erythropoietic organs), two types of stores (hepatocytes of the liver and
macrophages of the spleen and the liver), blood plasma which moves iron between tissues
and organs, and absorptive enterocytes in the duodenum through which iron enters the
body, ordinarily to replace small losses from the body caused by shedding of iron-con-
taining cells (Figure 1).
Figure 1. The key iron flows and compartments.
The largest of these iron compartments is the erythron (erythrocytes and their pre-
cursors), which contains approximately 2–3 g of iron, representing about 2/3–3/4 of the
total body iron in adult humans. Erythroid iron is almost entirely contained within hemo-
globin, at the concentration of 1 g of iron per liter of packed erythrocytes. The second
largest compartment are the hepatocyte stores, where up to 1 g of iron is contained in
cytoplasmic ferritin of hepatocytes. The stores are highly variable and can be nearly de-
pleted in many women of reproductive age because of menstrual blood loss combined
with low dietary intake. The recycling macrophages in the spleen, liver and marrow func-
tion as a rapid turnover compartment [1]. The spleen, which represents a substantial por-
tion of the macrophage storage compartment, normally contains only about 0.05 g of iron
but has the capacity for much larger amounts [2]. Only about 0.3 g of iron are contained
in the other tissues, in myoglobin of muscles and in iron-containing enzymes.
Figure 1. The key iron flows and compartments.
The largest of these iron compartments is the erythron (erythrocytes and their precur-
sors), which contains approximately 2–3 g of iron, representing about 2/3–3/4 of the total
body iron in adult humans. Erythroid iron is almost entirely contained within hemoglobin,
at the concentration of 1 g of iron per liter of packed erythrocytes. The second largest
compartment are the hepatocyte stores, where up to 1 g of iron is contained in cytoplas-
mic ferritin of hepatocytes. The stores are highly variable and can be nearly depleted in
many women of reproductive age because of menstrual blood loss combined with low
dietary intake. The recycling macrophages in the spleen, liver and marrow function as a
rapid turnover compartment [
1
]. The spleen, which represents a substantial portion of the
macrophage storage compartment, normally contains only about 0.05 g of iron but has the
capacity for much larger amounts [
2
]. Only about 0.3 g of iron are contained in the other
tissues, in myoglobin of muscles and in iron-containing enzymes.
Iron is transported around the body on blood plasma carrier protein transferrin, whose
two iron-binding sites are normally 20–45% occupied. Each cell in the body has transferrin
receptors (TfR1) that are endocytosed into acidified endosomes where iron dissociates from
the TfR1-transferrin complex and is transported across the endosomal membrane into the
cytoplasm. The transferrin compartment in blood plasma holds only 2–3 mg of iron but
delivers to target tissues 20–25 mg/day so its iron content turns over every few hours.
Despite changes in dietary iron content and tissue demand for iron, the concentration of
iron in the blood plasma in healthy humans is remarkably stable, as is the total body iron
content. Moreover, duodenal iron absorption increases several-fold after blood loss or
Int. J. Mol. Sci. 2021,22, 6493 3 of 13
exposure to hypoxia, or in response to iron deficiency. These observations provided early
evidence that the absorption and tissue distribution of iron must be subject to endocrine
regulation [3].
3. Molecular Basis of Systemic Iron Homeostasis
The absorption and tissue distribution of iron is principally controlled by the in-
teraction of the hepatic hormone hepcidin with ferroportin. Ferroportin is expressed in
iron-storing and iron-transporting tissues [
4
] and functions both as the hepcidin recep-
tor and the sole known cellular exporter of elemental iron in multicellular organisms
(
Figure 2
). The 25 amino-acid hepcidin peptide (MW 2.8 kD) is synthesized by hepatocytes
and secreted into blood plasma, with concentrations in healthy humans ranging from
approximately 2–20 nM [
5
,
6
], around hundred-fold higher than the concentration of the
similarly-sized peptide hormones insulin, glucagon or parathyroid hormone.
Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 3 of 13
Iron is transported around the body on blood plasma carrier protein transferrin,
whose two iron-binding sites are normally 20–45% occupied. Each cell in the body has
transferrin receptors (TfR1) that are endocytosed into acidified endosomes where iron dis-
sociates from the TfR1-transferrin complex and is transported across the endosomal mem-
brane into the cytoplasm. The transferrin compartment in blood plasma holds only 2–3
mg of iron but delivers to target tissues 20–25 mg/day so its iron content turns over every
few hours. Despite changes in dietary iron content and tissue demand for iron, the con-
centration of iron in the blood plasma in healthy humans is remarkably stable, as is the
total body iron content. Moreover, duodenal iron absorption increases several-fold after
blood loss or exposure to hypoxia, or in response to iron deficiency. These observations
provided early evidence that the absorption and tissue distribution of iron must be subject
to endocrine regulation [3].
3. Molecular Basis of Systemic Iron Homeostasis
The absorption and tissue distribution of iron is principally controlled by the inter-
action of the hepatic hormone hepcidin with ferroportin. Ferroportin is expressed in iron-
storing and iron-transporting tissues [4] and functions both as the hepcidin receptor and
the sole known cellular exporter of elemental iron in multicellular organisms (Figure 2).
The 25 amino-acid hepcidin peptide (MW 2.8 kD) is synthesized by hepatocytes and se-
creted into blood plasma, with concentrations in healthy humans ranging from approxi-
mately 2–20 nM [5,6], around hundred-fold higher than the concentration of the similarly-
sized peptide hormones insulin, glucagon or parathyroid hormone.
Figure 2. The interaction of hepcidin with ferroportin controls iron flows into plasma.
4. Ferroportin Structure
Ferroportin, a member of the solute carrier family, is systematically named SLC40A1.
The human protein has 571 amino acids for a molecular weight of around 65–70 kD [7–9].
The variation in molecular mass is likely caused by tissue-specific glycosylation but the
functional consequences of the glycosylation differences between the forms purified from
the duodenal enterocytes, hepatocytes and macrophages are not yet understood [10].
Structurally, ferroportin consists of two 6-transmembrane-helix bundles (N-lobe and C-
lobe) joined by a cytoplasmic loop, with both C- and N-termini located in the cytoplasm
(Figure 3). The two helix bundles enclose a cavity through which iron is thought to exit
the cell. The shape and similarity to more completely characterized family members sug-
gest that ferroportin exports cellular iron by an alternating access mechanism, wherein
Figure 2. The interaction of hepcidin with ferroportin controls iron flows into plasma.
4. Ferroportin Structure
Ferroportin, a member of the solute carrier family, is systematically named SLC40A1.
The human protein has 571 amino acids for a molecular weight of around 65–70 kD
[79]
.
The variation in molecular mass is likely caused by tissue-specific glycosylation but the
functional consequences of the glycosylation differences between the forms purified from
the duodenal enterocytes, hepatocytes and macrophages are not yet understood [
10
].
Structurally, ferroportin consists of two 6-transmembrane-helix bundles (N-lobe and C-
lobe) joined by a cytoplasmic loop, with both C- and N-termini located in the cytoplasm
(
Figure 3
). The two helix bundles enclose a cavity through which iron is thought to exit
the cell. The shape and similarity to more completely characterized family members sug-
gest that ferroportin exports cellular iron by an alternating access mechanism, wherein
ferroportin alternates between an open-inward conformation which binds intracellular
iron, and an open-outward conformation which releases the iron to the extracellular space
(
Figure 3
). The remarkable feat of structural characterization of a ferroportin from the bac-
terium Bdellovibrio bacteriovorus in both open-outward and open-inward conformations [
11
]
represents strong support for such a model. The exported species is almost certainly ferrous
iron [
12
] but the energetics of iron export is not yet well characterized. Because iron is
exported against the force exerted by the transmembrane electric field, and there is no
evidence of direct coupling of the transport to ATP hydrolysis, the export of iron is likely
facilitated by the coupled “downhill” transport of another ion or small molecule [13].
Int. J. Mol. Sci. 2021,22, 6493 4 of 13
Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 4 of 13
ferroportin alternates between an open-inward conformation which binds intracellular
iron, and an open-outward conformation which releases the iron to the extracellular space
(Figure 3). The remarkable feat of structural characterization of a ferroportin from the bac-
terium Bdellovibrio bacteriovorus in both open-outward and open-inward conformations
[11] represents strong support for such a model. The exported species is almost certainly
ferrous iron [12] but the energetics of iron export is not yet well characterized. Because
iron is exported against the force exerted by the transmembrane electric field, and there is
no evidence of direct coupling of the transport to ATP hydrolysis, the export of iron is
likely facilitated by the coupled “downhill” transport of another ion or small molecule
[13].
Structural analyses of ferroportin indicate that there are two divalent metal-binding
sites, one in each lobe, facing the internal cavity of ferroportin (Figure 3) [11,13,14]. How
these sites mediate the export of iron is not yet clear. The site in the N-lobe can bind cal-
cium which may have a modulatory role on iron transport [13]. Although calcium itself is
not transported, it is required for human ferroportin transport activity, and is thought to
directly bind to ferroportin and facilitate a conformational change critical to the transport
cycle. Another area of interest is how cytoplasmic iron destined for export reaches ferro-
portin. Current evidence indicates that cytoplasmic iron is predominantly present as a
complex of ferrous iron with reduced glutathione (summarized in [15]), and that the com-
plex of ferrous iron and reduced glutathione may be delivered to ferroportin via the cyto-
plasmic iron chaperone PCBP2 [16].
Figure 3. A current model of hepcidin (orange) interaction with ferroportin (blue) in which bind-
ing is dependent on iron (green). The framework of helices that make up ferroportin are depicted
as cylinders connected by extracellular and intracellular disordered loops. The side chains that
make up the binding sites for iron are shown. Modified from [14].
5. Ferroportin Interactions with Hepcidin
The flow of iron out of the cells is controlled by hepcidin through two known mech-
anisms: occlusion of the open-outward conformation of ferroportin by hepcidin [17], and
hepcidin-induced endocytosis and degradation of ferroportin [4]. The occlusion mecha-
nism would be effective at hepcidin concentrations where most ferroportin molecules re-
main occluded by hepcidin most of the time, and would be rapidly reversible when hep-
cidin concentrations are decreased. The endocytosis mechanism could be initiated by even
transient binding of hepcidin to ferroportin, and would cause the permanent removal of
ferroportin from the cell surface that would require resynthesis of ferroportin for the re-
covery of iron transport. The second mechanism would therefore be expected to occur at
lower concentrations of hepcidin, and to have a prolonged effect, even if hepcidin concen-
trations subsequently decrease. Although hepcidin peptide injected into mice is cleared
from blood circulation within minutes, its plasma iron lowering effect lasts for 24–48 h,
supporting the importance of the endocytic mechanism for ferroportin physiology [18].
Hepcidin regulation of ferroportin by the endocytic mechanism resembles the ge-
neric ligand-induced receptor endocytosis. It appears to require hepcidin-induced confor-
mational change in ferroportin that triggers the ubiquitination of the lysine-rich cytoplas-
Figure 3.
A current model of hepcidin (orange) interaction with ferroportin (blue) in which binding
is dependent on iron (green). The framework of helices that make up ferroportin are depicted as
cylinders connected by extracellular and intracellular disordered loops. The side chains that make up
the binding sites for iron are shown. Modified from [14].
Structural analyses of ferroportin indicate that there are two divalent metal-binding
sites, one in each lobe, facing the internal cavity of ferroportin (Figure 3) [
11
,
13
,
14
]. How
these sites mediate the export of iron is not yet clear. The site in the N-lobe can bind
calcium which may have a modulatory role on iron transport [
13
]. Although calcium
itself is not transported, it is required for human ferroportin transport activity, and is
thought to directly bind to ferroportin and facilitate a conformational change critical to
the transport cycle. Another area of interest is how cytoplasmic iron destined for export
reaches ferroportin. Current evidence indicates that cytoplasmic iron is predominantly
present as a complex of ferrous iron with reduced glutathione (summarized in [
15
]), and
that the complex of ferrous iron and reduced glutathione may be delivered to ferroportin
via the cytoplasmic iron chaperone PCBP2 [16].
5. Ferroportin Interactions with Hepcidin
The flow of iron out of the cells is controlled by hepcidin through two known mecha-
nisms: occlusion of the open-outward conformation of ferroportin by hepcidin [
17
], and
hepcidin-induced endocytosis and degradation of ferroportin [
4
]. The occlusion mech-
anism would be effective at hepcidin concentrations where most ferroportin molecules
remain occluded by hepcidin most of the time, and would be rapidly reversible when
hepcidin concentrations are decreased. The endocytosis mechanism could be initiated by
even transient binding of hepcidin to ferroportin, and would cause the permanent removal
of ferroportin from the cell surface that would require resynthesis of ferroportin for the
recovery of iron transport. The second mechanism would therefore be expected to occur at
lower concentrations of hepcidin, and to have a prolonged effect, even if hepcidin concen-
trations subsequently decrease. Although hepcidin peptide injected into mice is cleared
from blood circulation within minutes, its plasma iron lowering effect lasts for 24–48 h,
supporting the importance of the endocytic mechanism for ferroportin physiology [18].
Hepcidin regulation of ferroportin by the endocytic mechanism resembles the generic
ligand-induced receptor endocytosis. It appears to require hepcidin-induced conforma-
tional change in ferroportin that triggers the ubiquitination of the lysine-rich cytoplasmic
segment connecting the two 6-helix domains of ferroportin [
19
,
20
]. Ubiquitinated ferro-
portin is then targeted to lysosomes and proteasomes for degradation. A recent study [
21
]
provide convincing evidence that Rnf217 is an important E3 ubiquitin ligase that triggers
the degradation of ferroportin in response to hepcidin binding.
Recent advances in structural understanding of the hepcidin-ferroportin interaction
confirmed the simple models generated by targeted mutagenesis of hepcidin and ferro-
portin but also provided surprising new details [
14
]. In a nanodisc membrane model,
hepcidin is seen to bind to the C-lobe of ferroportin (Figure 3), with the highly variable
hepcidin loop largely extracellular, and the relatively conserved C- and N-termini of hep-
cidin deeply buried in the central cavity. The model also revealed an important unexpected
feature, the indication that the suspected hepcidin-binding site in the C-lobe, centered on
the critically important C325 thiol cysteine, utilizes an iron atom to coordinate hepcidin
Int. J. Mol. Sci. 2021,22, 6493 5 of 13
binding [
14
]. It is not yet clear how this contributes to hepcidin physiology but it can be
anticipated that the mechanism could provide selectivity for hepcidin to bind to ferroportin
molecules actively transporting iron as opposed to those that may be in a resting state. One
indication that this mechanism may have a physiological effect is the recently reported
inhibition of hepcidin-induced ferroportin degradation by the intracellular iron chelator
deferiprone but not by the extracellularly acting chelator deferoxamine [
22
]. How the
new structural model accommodates other hepcidin agonists ranging from minihepcidin
peptides [23] to small molecules [24] remains to be determined.
6. Hepcidin Synthesis and Elimination
Hepcidin production is principally transcriptionally regulated. The main systemic
regulators of hepcidin include plasma iron concentrations (mainly through the interaction
of diferric transferrin with transferrin receptors TFR1 and TFR2 in the liver), hepatic
iron stores, systemic inflammation mainly communicated to hepatocytes by IL-6, and
erythroid activity conveyed by the concentrations of the erythroid hormone erythroferrone
(
Figure 4
). The mechanisms were reviewed in recent publications [
25
,
26
] and will not be
discussed here.
Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 5 of 13
mic segment connecting the two 6-helix domains of ferroportin [19,20]. Ubiquitinated fer-
roportin is then targeted to lysosomes and proteasomes for degradation. A recent study
[21] provide convincing evidence that Rnf217 is an important E3 ubiquitin ligase that trig-
gers the degradation of ferroportin in response to hepcidin binding.
Recent advances in structural understanding of the hepcidin-ferroportin interaction
confirmed the simple models generated by targeted mutagenesis of hepcidin and ferro-
portin but also provided surprising new details [14]. In a nanodisc membrane model, hep-
cidin is seen to bind to the C-lobe of ferroportin (Figure 3), with the highly variable hep-
cidin loop largely extracellular, and the relatively conserved C- and N-termini of hepcidin
deeply buried in the central cavity. The model also revealed an important unexpected
feature, the indication that the suspected hepcidin-binding site in the C-lobe, centered on
the critically important C325 thiol cysteine, utilizes an iron atom to coordinate hepcidin
binding [14]. It is not yet clear how this contributes to hepcidin physiology but it can be
anticipated that the mechanism could provide selectivity for hepcidin to bind to ferro-
portin molecules actively transporting iron as opposed to those that may be in a resting
state. One indication that this mechanism may have a physiological effect is the recently
reported inhibition of hepcidin-induced ferroportin degradation by the intracellular iron
chelator deferiprone but not by the extracellularly acting chelator deferoxamine [22]. How
the new structural model accommodates other hepcidin agonists ranging from minihep-
cidin peptides [23] to small molecules [24] remains to be determined.
6. Hepcidin Synthesis and Elimination
Hepcidin production is principally transcriptionally regulated. The main systemic
regulators of hepcidin include plasma iron concentrations (mainly through the interaction
of diferric transferrin with transferrin receptors TFR1 and TFR2 in the liver), hepatic iron
stores, systemic inflammation mainly communicated to hepatocytes by IL-6, and
erythroid activity conveyed by the concentrations of the erythroid hormone erythrofer-
rone (Figure 4). The mechanisms were reviewed in recent publications [25,26] and will not
be discussed here.
Figure 4. Iron, erythropoiesis and inflammation regulate hepcidin transcription through their ef-
fects on hepatocytes and by modulating the paracrine signaling between hepatic sinusoidal endo-
thelium and hepatocytes.
The size of hepcidin and its mechanism of action suggest two catabolic mechanisms.
One is efficient filtration of hepcidin as a small peptide through the renal glomerular
Figure 4.
Iron, erythropoiesis and inflammation regulate hepcidin transcription through their effects
on hepatocytes and by modulating the paracrine signaling between hepatic sinusoidal endothelium
and hepatocytes.
The size of hepcidin and its mechanism of action suggest two catabolic mechanisms.
One is efficient filtration of hepcidin as a small peptide through the renal glomerular
membrane [
27
], followed by reuptake and degradation of filtered hepcidin in the proximal
tubule, utilizing a generic mechanism for recycling the amino acid content of filtered
proteins. Efficient filtration of hepcidin in the glomerulus is facilitated by the relatively
low fraction of hepcidin that is bound by plasma protein, estimated at about 40% [
28
]. A
small percentage of filtered hepcidin survives the uptake and degradation in the proximal
tubule and can be detected in urine, similarly to other peptide hormones. In support of the
important role of the kidneys in hepcidin removal, serum hepcidin concentrations are very
high in patients with chronic renal failure, but are effectively lowered by hemodialysis [
29
].
This effect of hemodialysis is brief, as hepatic hepcidin synthesis is sufficiently rapid
to restore hepcidin concentrations within hours. The second mechanism of hepcidin
clearance depends on the endocytosis of the hepcidin-ferroportin complexes by the cells
that express high concentrations of ferroportin [
30
,
31
]. The quantitative contribution of the
two mechanisms as well as any possible alternative mechanisms of hepcidin catabolism
are not known.
Int. J. Mol. Sci. 2021,22, 6493 6 of 13
7. Evidence for Exclusivity of the Hepcidin-Ferroportin Ligand-Receptor Dyad
Ferroportin is evolutionarily ancient and can be found in plants and multicellular
animals as simple as hydra. By contrast, hepcidin is a vertebrate hormone, with no known
antecedent before fish. Remarkably, the essential conserved amino acid at the ferroportin
binding site for hepcidin, C326 (human numbering), first appears in cartilaginous fish
(Figure 5) some of which have a second ferroportin gene that has another amino acid in
this position. C326 is strictly conserved in all vertebrates that have hepcidin but another
amino acid is in this position in invertebrates, indicating that hepcidin and its binding site
on ferroportin co-evolved. A second hepcidin gene is found in some vertebrates, mostly
fish, but is not involved in iron regulation, and is hypothesized to have another function,
perhaps as an antimicrobial peptide [32].
Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 6 of 13
membrane [27], followed by reuptake and degradation of filtered hepcidin in the proximal
tubule, utilizing a generic mechanism for recycling the amino acid content of filtered pro-
teins. Efficient filtration of hepcidin in the glomerulus is facilitated by the relatively low
fraction of hepcidin that is bound by plasma protein, estimated at about 40% [28]. A small
percentage of filtered hepcidin survives the uptake and degradation in the proximal tu-
bule and can be detected in urine, similarly to other peptide hormones. In support of the
important role of the kidneys in hepcidin removal, serum hepcidin concentrations are
very high in patients with chronic renal failure, but are effectively lowered by hemodial-
ysis [29]. This effect of hemodialysis is brief, as hepatic hepcidin synthesis is sufficiently
rapid to restore hepcidin concentrations within hours. The second mechanism of hepcidin
clearance depends on the endocytosis of the hepcidin-ferroportin complexes by the cells
that express high concentrations of ferroportin [30,31]. The quantitative contribution of
the two mechanisms as well as any possible alternative mechanisms of hepcidin catabo-
lism are not known.
7. Evidence for Exclusivity of the Hepcidin-Ferroportin Ligand-Receptor Dyad
Ferroportin is evolutionarily ancient and can be found in plants and multicellular
animals as simple as hydra. By contrast, hepcidin is a vertebrate hormone, with no known
antecedent before fish. Remarkably, the essential conserved amino acid at the ferroportin
binding site for hepcidin, C326 (human numbering), first appears in cartilaginous fish
(Figure 5) some of which have a second ferroportin gene that has another amino acid in
this position. C326 is strictly conserved in all vertebrates that have hepcidin but another
amino acid is in this position in invertebrates, indicating that hepcidin and its binding site
on ferroportin co-evolved. A second hepcidin gene is found in some vertebrates, mostly
fish, but is not involved in iron regulation, and is hypothesized to have another function,
perhaps as an antimicrobial peptide [32].
Figure 5. Although ferroportin is evolutionarily ancient, the thiol cysteine required for hepcidin
binding by ferroportin appears first in early vertebrate ferroportins. Cartilaginous fish ferroportin
that contains the C326-equivalent is denoted with species name in red. Second ferroportin in carti-
laginous fish that has another amino acid in this position is denoted with species name in blue.
Invertebrate ferroportin lacks C326 and is denoted with species name in black.
Consistent with the dyadic relationship between hepcidin and ferroportin, the iso-
steric C326S mutation in mice and humans, which causes complete loss of binding of hep-
cidin to ferroportin, phenocopies severe hepcidin deficiency [33]. It is therefore likely that
Figure 5.
Although ferroportin is evolutionarily ancient, the thiol cysteine required for hepcidin
binding by ferroportin appears first in early vertebrate ferroportins. Cartilaginous fish ferroportin that
contains the C326-equivalent is denoted with species name in red. Second ferroportin in cartilaginous
fish that has another amino acid in this position is denoted with species name in blue. Invertebrate
ferroportin lacks C326 and is denoted with species name in black.
Consistent with the dyadic relationship between hepcidin and ferroportin, the isosteric
C326S mutation in mice and humans, which causes complete loss of binding of hepcidin to
ferroportin, phenocopies severe hepcidin deficiency [
33
]. It is therefore likely that hepcidin
has no other nonredundant function than to bind to ferroportin and regulate its ability to
transport iron.
8. Regulation of Ferroportin by Intracellular Signals
In addition to systemic control of ferroportin by circulating hepcidin, ferroportin
is also regulated by intracellular conditions. These local regulatory mechanisms may
decrease toxicity from excessive cellular iron, preserve cellular iron under conditions of
systemic iron deficiency, or amplify the effect of systemic hepcidin changes. In addition,
inflammatory stimuli in the form of Toll-like receptor ligands can directly suppress cellular
ferroportin mRNA levels and decrease cellular iron export to blood plasma [
34
]. Table 1
summarizes the local regulators of ferroportin and the setting in which their activities have
been characterized.
Int. J. Mol. Sci. 2021,22, 6493 7 of 13
Table 1. Hepcidin-independent mechanisms that regulate ferroportin.
Mechanism Mode Cell Type Cellular Effect
on FPN Note
IRE-IRP [35] translational macrophage cellular iron =
FPN
This mechanism is
bypassed by
ferroportin transcripts
lacking the 50IRE, in
erythroid cells and
duodenal enterocytes
Iron via Nrf2 [36] transcriptional macrophage cellular iron =
FPN
Requires high iron
concentrations
Heme via BACH
and Nrf2 [36]transcriptional macrophage
cellular heme =
FPN
Induces ferroportin in
macrophages that
recycle iron from
hemoglobin and heme
miR-485–3p [37] translational multiple cell types cellular iron =
FPN
Physiological role is
uncertain
TLR [34] transcriptional macrophage Ligands FPN
Unclear how much this
mechanism contributes
to responses to
infections
HIF2α[38] transcriptional enterocyte cellular iron =
FPN
May induce
ferroportin mRNA in
enterocytes during low
hepcidin states
miR-20a and
miR-20b [39,40]translational enterocyte, lung miRNA = FPN Physiological role is
uncertain
The IRE-IRP system functions in most cells to coordinate ferroportin translation with
intracellular iron levels. Ferroportin mRNA contains 5
0
IRE. Under conditions of iron
deficiency, binding of IRP1/2 to 5
0
IRE inhibits ferroportin translation, thus limiting iron
export and conserving cellular iron. In duodenal enterocytes, however, the existence
of ferroportin transcripts that lack the 5
0
IRE would make these short-lived cells less
responsive to their own cellular iron levels, and by implication more responsive to systemic
iron requirements communicated to enterocytes by the concentration of hepcidin in blood
plasma [
35
]. Low hepcidin levels during iron deficiency and anemia allow continuous iron
export from enterocytes, resulting in activation of the enterocytes HIF system (specifically
HIF2
α
), which further increases ferroportin transcription [
38
]. HIF2
α
also mediates increase
in other mRNAs encoding proteins involved in the apical absorption of dietary iron (DCYTB
and DMT1) thus leading to a coordinated increase in apical and basolateral iron transport
and overall increase in duodenal iron absorption.
The 5
0
IRE-lacking form of ferroportin mRNA is also found in erythroblasts, and the
erythroid cells supply essential iron to other organs during iron deficiency [
35
]. Here
ferroportin may also function to mitigate iron toxicity by releasing excess iron not used for
hemoglobin synthesis.
Iron-recycling macrophages, found in the spleen (but also in the liver and the marrow),
are intermittently confronted with large boluses of hemoglobin that must be digested to
release heme, which is then degraded via heme oxygenase to release iron. This process
is dependent on the derepression of multiple genes involved in this process, including
ferroportin that is required to export the recycled iron. The primary stimulus for the dere-
pression of these genes is heme, sensed by the transcriptional repressor Bach1 (Btb And Cnc
Homology 1) that binds heme and dissociates from its partner the small Maf protein bound
to the promoter element MARE/ARE (Maf-recognition element/antioxidant response
element). Bach1 is replaced by the transcriptional inducer Nrf2 which is stabilized by heme,
translocated into the nucleus, and stimulates the transcription of the ferroportin gene and
other genes involved in the recycling of heme to iron [
36
,
41
]. At high concentrations of
Int. J. Mol. Sci. 2021,22, 6493 8 of 13
iron, ferroportin mRNA and protein are increased, presumably because of oxidative stress
causing Nrf2-mediated induction of ferroportin [42,43].
There is evidence that microRNAs such as miR-485-3p, miR-20a and miR-20b, can
posttranscriptionally downregulate ferroportin but the physiological role of this mechanism
is uncertain [37,39,40].
9. Local Functions of Ferroportin and Its Autocrine/Paracrine Regulation by Hepcidin
The “professional” iron exporting cell types, i.e., duodenal enterocytes, iron-recycling
macrophages and iron-storing hepatocytes, all deliver iron to blood plasma for use in
hemoglobin synthesis by erythroblasts as well as for other uses in iron-consuming tis-
sues. However, ferroportin is also found in other tissues where it appears to serve a
cell-autonomous function of releasing unneeded and potentially toxic iron.
Ferroportin is expressed at a remarkably high level in erythroblasts and mature
erythrocytes. In erythroblasts, it may release unobligated iron and here it is subject to
hepcidin regulation [
44
]. In mature erythrocytes, which lack an endocytic mechanism and
may therefore be less sensitive to inhibition of iron export by systemic hepcidin, ferroportin
may serve as a safety valve to release labile iron leaking from hemoglobin molecules
damaged by repeated cycling between high and low oxygen states. The importance of these
mechanism is illustrated by the adverse consequences of erythroid-specific disruption of
the ferroportin gene in mice, which suffer from mild hemolytic anemia caused by increased
sensitivity of erythrocytes to hemolytic stress [45,46].
Ferroportin is required for normal functioning of cardiac myocytes [
47
], presumably
to remove excess iron. Cardiac myocytes also produce hepcidin but this does not contribute
significantly to systemic hepcidin levels. Rather, hepcidin acts as an autocrine or paracrine
regulator of ferroportin that may protect tissues from loss of iron through ferroportin and
extreme iron deficiency when systemic hepcidin concentrations are very low [47].
Macrophage ferroportin also plays an important role in iron recycling after muscle
injury [
48
]. Selective loss of macrophage ferroportin in mice with muscle injury lim-
ited local iron recycling, impaired muscle healing and resulted in smaller myofibers and
fat accumulation.
In a mouse model of colitis, the hepcidin-ferroportin interaction in the intestine was
shown to be important for mucosal healing [
49
]. Absence of local hepcidin production
or insensitivity of macrophage ferroportin to hepcidin is thought to result in higher local
extracellular iron concentrations, affecting the growth of luminal and tissue-infiltrating
bacteria and worsening DSS-induced colitis.
During pregnancy, ferroportin is very highly expressed on the basal surface of syncy-
tiotrophoblast in the placenta [
7
,
50
], where it plays an indispensable role in iron transfer to
the fetus. Placental ferroportin appears to be strongly regulated by the trophoblast intracel-
lular iron concentrations, via the IRE-IRP system [
50
]. Interestingly, in normal pregnancy,
fetal hepcidin concentrations are very low in mice [50] and do not appear to contribute to
baseline iron homeostasis or affect iron transport through placental ferroportin. However,
fetal hepatic production of hepcidin may be sufficient to act on fetal hepatic ferroportin in
an autocrine/paracrine manner to retain iron in the fetal liver [
51
] where it is used for fetal
erythropoiesis until erythropoiesis transitions to the marrow, close to birth.
10. Functions of the Hepcidin-Ferroportin Axis in Host Defense
Infection and inflammation induce hepcidin, predominantly through the effects of the
cytokine interleukin-6 (IL-6) [
52
], and increased concentrations of hepcidin then inhibit
ferroportin activity, leading to depletion of iron in plasma and extracellular fluid (hypofer-
remia). Independently of hepcidin, microbial molecules that stimulate toll-like receptors
suppress cellular ferroportin mRNA transcription [
34
] sufficiently to cause hypoferremia
but it is not clear how much this effect contributes to hypoferremia generated during
infections
in vivo
. As ferroportin is much older on the evolutionary timeline than hepcidin,
Int. J. Mol. Sci. 2021,22, 6493 9 of 13
studies of the hypoferremic effects of infection and inflammation in invertebrates, which
lack hepcidin, should be illuminating.
Hypoferremia has long been suspected of having a host defense function but direct
evidence for this is quite recent [
53
]. Since transferrin-bound iron is poorly accessible to
most microbes, the most important iron species that supports microbial growth is non-
transferrin bound iron (NTBI) [
54
]. During infections, increased cytokine concentrations
inhibit erythropoiesis and activate the production of myeloid cells important for host
defense [
55
]. The inhibition of erythropoiesis decreases the consumption of plasma iron, just
as increased recycling of damaged erythrocytes and other cells during infections increases
iron delivery to plasma. The imbalance would raise the saturation of transferrin by iron
and enhance the production of NTBI, thereby increasing the risk of microbial outgrowth.
However, inflammatory increase of hepcidin and the effects of inflammation on ferroportin
transcription prevent the generation of NTBI by sequestering iron, predominantly inside
macrophages. This form of “nutritional immunity” is particularly important in infections
with gram negative bacteria and some fungi ([
54
,
56
58
]. Hepcidin can also kill microbes on
contact
in vitro
[
59
,
60
] but the concentrations required for this direct microbicidal activity
exceeded its concentrations in extracellular fluid, even in inflamed models.
11. Disorders of the Hepcidin-Ferroportin System
The hepcidin-ferroportin system, in analogy to other endocrine systems, can be dis-
rupted by mutations that affect the quantity or functionality of the ligand hepcidin or
its receptor ferroportin. Loss-of-function mutations in positive regulators of hepcidin or
the hepcidin gene itself (Table 2) cause hereditary hemochromatosis [
61
], a set of diseases
characterized by increased absorption of dietary iron, often causing massively increased
body iron content, and high plasma iron concentrations that saturate the iron-binding
capacity of transferrin, leading to the appearance of non-transferrin-bound iron (NTBI).
In affected patients who are not treated, NTBI is taken up avidly through alternative iron
transporters [
62
] in the liver, pancreas, heart and endocrine glands, causing progressive
tissue injury, organ failure, and even premature death. Two forms of hemochromatosis
define its clinical spectrum. At one extreme is the severe juvenile form that can cause severe
heart disease, endocrinopathy and death in young adults and at the other extreme are much
less severe adult forms which can even be clinically silent and not cause organ damage.
Loss-of-function ferroportin mutations cause iron overload that is usually localized to
macrophages, cells that use ferroportin to export iron recycled from senescent erythrocytes,
and causes increased serum ferritin but not clinically significant disease.
Table 2. Forms of hereditary hemochromatosis.
Mutated Gene Function Hepcidin Tissue Iron Clinical Severity
HFE Loss Low Liver, heart, endocrine Moderate, adult form
TFR2 Loss Low Liver, heart, endocrine Moderate, adult form
Hemojuvelin Loss Very low Heart, endocrine, liver Severe, juvenile form
Hepcidin Loss Very low to absent Heart, endocrine, liver Severe, juvenile form
Ferroportin Gain High Liver, heart, endocrine Variable, some severe
Ferroportin Loss Normal or low Spleen macrophages Mild
Loss-of-function mutations in the negative regulator of hepcidin transcription, the
transmembrane serine protease TMPRSS6 (also called matriptase 2), in mice or humans
cause a disease “iron-refractory iron deficiency anemia” (IRIDA) where hepcidin is ex-
cessive despite severe iron deficiency [
63
,
64
]. Elevated hepcidin inhibits intestinal iron
absorption and the release of stored and recycled iron, leading to low plasma iron concen-
trations and a microcytic anemia.
Hepcidin concentrations can also become dysregulated as a consequence of diseases
that affect iron absorption from the diet or cause a loss of blood from the body (each 1 mL of
packed erythrocytes represents about 1 mg of iron), produce systemic inflammation, make
Int. J. Mol. Sci. 2021,22, 6493 10 of 13
erythropoiesis ineffective (lots of erythrocyte precursors but few mature) or cause anemia
requiring erythrocyte transfusions without matching blood loss. These disease pathways
are summarized in Table 3. Iron deficiency, iron deficiency anemia [
65
] and anemia of
inflammation [66] are very common disorders affecting a large portion of the population.
Table 3. Disorders with dysregulated hepcidin production.
Disease State Serum Hepcidin Plasma Iron Body Iron Common Phenotypes
Iron deficiency Low Low Low Fatigue, anemia
Systemic inflammation High Low Normal Fatigue, anemia
Ineffective erythropoiesis Low High Increased Like hemochromatosis
Erythrocyte transfusions High High Increased Like hemochromatosis
12. Summary
The hepcidin-ferroportin system evolved to maintain stable body iron content and
plasma iron concentrations. The system also plays an important role in innate immunity,
particularly in host defense against gram-negative bacteria and some fungi. Dysregulation
of hepcidin or ferroportin production or their interaction underlies the pathogenesis of
a spectrum of iron disorders, from iron restrictive anemias to iron overload conditions.
Thus, plasma hepcidin measurements may be useful for diagnosing iron disorders, and
therapeutic targeting of the hepcidin-ferroportin system is a promising new direction to
develop improved treatments for iron disorders.
Author Contributions:
Conceptualization, T.G. and E.N. Writing, T.G. and E.N. All authors have
read and agreed to the published version of the manuscript.
Funding: This research received no external funding.
Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.
Data Availability Statement: Not applicable.
Conflicts of Interest:
E.N. and T.G. are shareholders of and scientific advisors for Intrinsic Life-
Sciences and Silarus Therapeutics and are consultants for Ionis Pharmaceuticals, Protagonist Thera-
peutics, Akebia, Vifor Pharma, and Disc Medicine.
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... Nevertheless, these approaches have notable limitations, for example, frequent phlebotomy can result in anemia [15], while long-term use of iron chelating agents may lead to complications such as infections, gastrointestinal disorders, and skin damage [16]. As the key hormone of iron homeostasis, hepcidin has emerged as a crucial therapeutic target for managing iron-related disorders such as anemia and iron overload [17,18]. The ability of hepcidin analogs, such as mini-hepcidins, to alleviate iron overload has been investigated for hepcidin knockout mice [19]. ...
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Iron overload disease is characterized by the excessive accumulation of iron in the body. To better alleviate iron overload, there is an urgent need for safe and effective small molecule compounds. Rubiadin, the active ingredient derived from the Chinese herb Prismatomeris tetrandra, possesses notable anti-inflammatory and hepatoprotective properties. Nevertheless, its impact on iron metabolism remains largely unexplored. To determine the role of rubiadin on iron metabolism, Western blot analysis, real-time PCR analysis, and the measurement of serum iron were performed. Herein, we discovered that rubiadin significantly downregulated the expression of transferrin receptor 1, ferroportin 1, and ferritin light chain in ferric-ammonium-citrate-treated or -untreated HepG2 cells. Moreover, intraperitoneal administration of rubiadin remarkably decreased serum iron and duodenal iron content and upregulated expression of hepcidin mRNA in the livers of high-iron-fed mice. Mechanistically, bone morphogenetic protein 6 (BMP6) inhibitor LDN-193189 completely reversed the hepcidin upregulation and suppressor of mother against decapentaplegic 1/5/9 (SMAD1/5/9) phosphorylation induced by rubiadin. These results suggested that rubiadin increased hepcidin expression through the BMP6/SMAD1/5/9-signaling pathway. Collectively, our findings uncover a crucial mechanism through which rubiadin modulates iron metabolism and highlight it as a potential natural compound for alleviating iron-overload-related diseases.
... Iron is the most abundant essential trace element in the human body, primarily existing in the form of ferritin, and is regulated by hepcidin [7,8]. Kowdley et al. [9] suggested that elevated serum ferritin levels, hepatic iron deposition and iron overload are closely correlated with NAFLD. ...
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Background Non-alcoholic fatty liver disease (NAFLD) is a spectrum of chronic liver diseases characterized by hepatic steatosis exceeding 5% in the absence of alcohol and other liver-damaging factors. Clinical studies have identified a potential link between abnormal iron metabolism and the high incidence of NAFLD; however, the results from clinical trials remain inconsistent. This meta-analysis aims to compare serum hepcidin levels and the hepcidin/ferritin ratio between adults with NAFLD and those without to explore their potential relationship with NAFLD. Methods A systematic search was conducted across the Web of Science platform, Cochrane, Scopus, Embase, and PubMed databases from their inception until December 18, 2024. The analysis primarily focused on serum hepcidin levels and the hepcidin/ferritin ratio. Observational studies comparing serum hepcidin levels and the hepcidin/ferritin ratio between individuals with NAFLD and control groups were included. A random-effects model was employed to calculate effect estimates, and outcomes were reported as standardized mean differences (SMD) with 95% confidence intervals (95% CI). Results Following the systematic review, a total of 19 studies, comprising 2216 patients and 2125 controls, were included. The findings revealed a statistically significant difference in both hepcidin levels (SMD = 1.03, 95% CI: 0.49 to 1.56, p < 0.001) and the hepcidin/ferritin ratio (SMD = -1.13, 95% CI: -1.79 to -0.46, p < 0.001) between NAFLD and controls. Significant heterogeneity was observed across studies for both hepcidin (I² = 98.2%) and the hepcidin/ferritin ratio (I² = 93.3%), and the limited number of studies on hepcidin/ferritin were acknowledged as key limitations. Subgroup analysis revealed that patients with obesity exhibited higher levels of hepcidin (SMD = 1.12, 95% CI: 0.40 to 1.97) than overweight (SMD = 0.88, 95% CI: 0.05 to 1.72). Meta-regression analysis identified the hepcidin measurement method (p < 0.01), male-to-female ratio (p < 0.01), and study quality (p < 0.01) as significant moderators of the observed heterogeneity. Conclusion This meta-analysis revealed a significant association between hepcidin levels, the hepcidin/ferritin ratio and NAFLD in adults. Further investigations are needed to fully elucidate the role of these variables in iron metabolism and their potential impact on the diagnosis, prevention, and management of NAFLD.
... alone exposure. The imbalance between Fe uptake and efflux resulted in a significant increment of Fe bioaccumulation (Nemeth and Ganz, 2021). ...
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This study explores the effects of norfloxacin (NOR) on oxidative damage, iron (Fe) transport, energy metabolism, and immunotoxicity in the intestine of large yellow croaker under Fe stress. The fish were subjected to Fe (180 μg/L), low-dose NOR (1.8 μg/L, LNOR), high-dose NOR (180 μg/L, HNOR), Fe plus LNOR, and Fe plus HNOR for 60 days. These results demonstrated that Fe alone exposure increased malondialdehyde (MDA), protein carboxylation (PC), and mortality rate, and impaired intestinal tissue, which was related to the increment of Fe accumulation. Compared to Fe alone exposure, Fe plus LNOR exposure decreased MDA, PC, and mortality rate, and alleviated intestinal malformations by improving Fe transport, energy metabolism, anti-inflammatory response, and protein folding protective effect, and reducing pro-inflammatory response, indicating that LNOR had an antagonistic effect on Fe toxicity. Compared to Fe alone exposure, Fe plus HNOR exposure elevated MDA, PC, and mortality rate, and deteriorated intestinal malformations by inhibiting Fe excretion, energy metabolism, anti-inflammatory response, and protein folding protective effect, and enhancing pro-inflammatory response, indicating a synergetic effect between HNOR and Fe stress. These findings suggested that NOR had a dose-dependent effect on Fe-toxicity to large yellow croaker, which contributes to revealing the molecular mechanisms behind their interaction and its ecological implications.
... Crianças com sobrepeso e obesidade estão mais propensas a terem deficiência de ferro, pois a obesidade predispõe à anemia ferropriva. A relação entre as duas patologias, se deve ao aumento inflamatório do tecido adiposo, o qual aumenta a produção de hepcidina (hormônio regulador chave do metabolismo do ferro no organismo), que é produzida principalmente pelo fígado, agindo como um mediador de defesa (Nemeth & Ganz, 2021). Esse hormônio desempenha um papel crucial no controle da quantidade de ferro disponível na circulação sanguínea, regulando a absorção desse mineral no intestino. ...
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... Thalassemia (32). This is in agreement with previous studies (30,31), who have found that the human iron absorption is regulated by a combination of factors including tissue oxygenation, the body's iron store, and the erythropoietic requirement for iron because thalassaemic subjects exhibited lower levels of hepcidin than healthy ones, this could directly explain our findings. This is in agreement with previous studies (33,34), which have found that hepcidin mRNA expression was discovered to be downregulated in the Hbbth3/1 mice, the murine model of human thalassaemia. ...
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Article history: Background: α-thalassaemia is mostly caused by deletions in the alpha-globin gene complex, which result in either reduced or absent α-globin chain synthesis. β-globin chains are either absent or decreased in ß-thalassemia. Analyzing the regulatory genes (foxO1 and hepcidin) for iron and ferritin in thalassemic patients was the goal of this study. Methods: Fifty individuals were selected for this study. Ten participants (5 females and 5 males) were healthy when they were recruited from the general community and visited Al-Hadba'a Hospital (Mosul City) for blood withdrawal, while the forty patients (20 females and 20 males) have thalassaemia and range in age from 8 to 17. Results: Male participants had higher levels of iron and ferritin than female participants. Furthermore, there were notable differences between male and female thalassaemic subjects. Additionally, ferritin and HbF were directly correlated with iron level and sex. Hepcidin expression analysis in healthy rather than thalassaemic participants found both down-and up-regulation; forkhead box O1 (foxO1) expression analysis demonstrated the reverse hemoglobin type pattern. Conclusion: In both thalassaemic and healthy subjects, gender was associated with the serum levels of iron and ferritin and the genes that regulate them. Hepcidin and foxO1 synchronized with iron and ferritin in the human body.
... Liver is the main site of hepcidin production; thus, HCC has been strongly associated with hepcidin dysregulation [157]. The normal range of serum hepcidin in the human body is between 2 and 20 nm, while hepcidin expression is mostly elevated in many types of cancer, including prostate cancer, multiple myeloma, breast cancer, and non-lymphoma Hodgkin's disease [158,159]. Iron homeostasis is triggered by mutations in tumor cells or tissues and is further dysregulated by altered expression of hepcidin [143]. In cases of increased hepcidin levels, iron transfer from enterocytes and macrophages into the circulation is inhibited, while in cases of decreased hepcidin levels, plasma iron levels are upregulated, resulting in various levels of toxicity [160]. ...
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Hepatocellular carcinoma (HCC), the most common form of primary liver cancer, is rising in global incidence and mortality. Metabolic dysfunction-associated steatotic liver disease (MASLD), one of the leading causes of chronic liver disease, is strongly linked to metabolic conditions that can progress to liver cirrhosis and HCC. Iron overload (IO), whether inherited or acquired, results in abnormal iron hepatic deposition, significantly impacting MASLD development and progression to HCC. While the pathophysiological connections between hepatic IO, MASLD, and HCC are not fully understood, dysregulation of glucose and lipid metabolism and IO-induced oxidative stress are being investigated as the primary drivers. Genomic analyses of inherited IO conditions reveal inconsistencies in the association of certain mutations with liver malignancies. Moreover, hepatic IO is also associated with hepcidin dysregulation and activation of ferroptosis, representing promising targets for HCC risk assessment and therapeutic intervention. Understanding the relationship between hepatic IO, MASLD, and HCC is essential for advancing clinical strategies against liver disease progression, particularly with recent IO-targeted therapies showing potential at improving liver biochemistry and insulin sensitivity. In this review, we summarize the current evidence on the pathophysiological association between hepatic IO and the progression of MASLD to HCC, underscoring the importance of early diagnosis, risk stratification, and targeted treatment for these interconnected conditions.
... Further optimization of paraffin-embedded tissue synchrotron imaging could provide a more sensitive visual indicator of zinc status to corroborate molecular analyses. For iron, ferroportin in duodenal enterocytes is more sensitive to systemic than cellular iron levels [70]. Since BSGE did not change ferroportin expression (Figure 5d) or iron localization and concentration (Figure 4), BSGE likely did not change systemic levels of iron. ...
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Upcycling brewers’ spent grain (BSG) into poultry feed needs to be optimized. Since broiler chickens inefficiently digest fiber, we created a water-soluble BSG extract (BSGE) to explore this fraction’s potential nutritional benefits. We utilized intra-amniotic administration (in ovo) to target the gastrointestinal tract of broiler embryos. BSGE increased villus surface area and goblet cell quantity and size, implying improved duodenal development. The extract also changed cecal Escherichia coli (E. coli) and Clostridium abundances. Synchrotron X-ray fluorescence microscopy, along with zinc and iron transporter relative expression, did not reveal significant changes by BSGE. These findings highlight the potential for BSGE to be a functional feed component, underscoring the potential value of upcycling this byproduct. This pilot study supports future work exploring the impact of BSGE within feed and its effects over long-term consumption.
Chapter
Iron is an essential micronutrient for the survival and growth of living matter. In the context of host-pathogen interactions, they both require iron as a cofactor in proteins involved in energy production, DNA replication, and gene transcription, among many metabolic processes. Although iron is essential for life, excess iron is harmful because it generates reactive oxygen species by Fenton chemistry, which leads to oxidative stress, lipid peroxidation, and cell damage and death. Moreover, normal body physiology requires ferrous iron in minimal amounts. These considerations have served as the driving force behind the evolution of intricate mechanisms that tightly maintain iron homeostasis in mammals. Microbial infections trigger an inflammatory response that leads to host iron sequestration and hypoferremia, often called nutritional immunity. The goal of this innate response, along with other measures that help to sequester host iron, is to limit iron availability to the pathogen as a means of inhibiting pathogen survival, growth, and colonization. To counter host-imposed iron sequestration measures, the pathogen initiates a concerted effort to acquire iron rapidly from different sources within the host using several ingenious strategies. This chapter discusses the epic fight between the host as it attempts to sequester iron and the pathogen as it tries to acquire it.
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Ferroportin (FPN), the body's sole iron exporter, is essential for maintaining systemic iron homeostasis. In response to either increased iron or inflammation, hepatocyte-secreted hepcidin binds to FPN, inducing its internalization and subsequent degradation. However, the E3 ubiquitin ligase that underlies FPN degradation has not been identified. Here, we report the identification and characterization of a novel mechanism involving the RNF217-mediated degradation of FPN. A combination of two different E3 screens revealed that the Rnf217 gene is a target of Tet1, mediating the ubiquitination and subsequent degradation of FPN. Interestingly, loss of Tet1 expression causes an accumulation of FPN and an impaired response to iron overload, manifested by increased iron accumulation in the liver together with decreased iron in the spleen and duodenum. Moreover, we found that the degradation and ubiquitination of FPN could be attenuated by mutating RNF217. Finally, using two conditional knockout mouse lines, we found that knocking out Rnf217 in macrophages increases splenic iron export by stabilizing FPN, whereas knocking out Rnf217 in intestinal cells appears to increase iron absorption. These findings suggest that the Tet1-RNF217-FPN axis regulates iron homeostasis, revealing new therapeutic targets for FPN-related diseases.
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The serum iron level in humans is tightly controlled by the action of the hormone hepcidin on the iron efflux transporter ferroportin. Hepcidin regulates iron absorption and recycling by inducing ferroportin internalization and degradation1. Aberrant ferroportin activity can lead to diseases of iron overload, such as hemochromatosis, or iron limitation anemias2. Here, we determined cryogenic electron microscopy (cryo-EM) structures of ferroportin in lipid nanodiscs, both in the apo state and in complex with cobalt, an iron mimetic, and hepcidin. These structures and accompanying molecular dynamics simulations identify two metal binding sites within the N- and C-domains of ferroportin. Hepcidin binds ferroportin in an outward-open conformation and completely occludes the iron efflux pathway to inhibit transport. The carboxy-terminus of hepcidin directly contacts the divalent metal in the ferroportin C-domain. We further show that hepcidin binding to ferroportin is coupled to iron binding, with an 80-fold increase in hepcidin affinity in the presence of iron. These results suggest a model for hepcidin regulation of ferroportin, where only iron loaded ferroportin molecules are targeted for degradation. More broadly, our structural and functional insights are likely to enable more targeted manipulation of the hepcidin-ferroportin axis in disorders of iron homeostasis.
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β-thalassemia is a genetic anemia caused by partial or complete loss of β-globin synthesis leading to ineffective erythropoiesis and RBCs with short life-span. Currently, there is no efficacious oral medication modifying anemia for patients with beta-thalassemia. The inappropriately low levels of the iron regulatory hormone hepcidin enable excessive iron absorption by ferroportin, the unique cellular iron exporter in mammals, leading to organ iron overload and associated morbidities. Correction of unbalanced iron absorption and recycling by induction of hepcidin synthesis or treatment with hepcidin mimetics ameliorates β-thalassemia. However, hepcidin modulation or replacement strategies currently in clinical development all require parenteral drug administration. We identified oral ferroportin inhibitors by screening a library of small molecular weight compounds for modulators of ferroportin internalization. Restricting iron availability by VIT-2763, the first clinical stage oral ferroportin inhibitor, ameliorated anemia and the dysregulated iron homeostasis in the Hbbth3/+ mouse model of beta-thalassemia intermedia. VIT-2763 not only improved erythropoiesis but also corrected the proportions of myeloid precursors in spleens of Hbbth3/+ mice. VIT-2763 is currently developed as an oral drug targeting ferroportin for the treatment of β-thalassemia.
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Erythroferrone (ERFE) is the main erythroid regulator of hepcidin, the homeostatic hormone controlling plasma iron levels and total body iron. When the release of erythropoietin from the kidney stimulates the production of new red blood cells, it also increases the synthesis of ERFE in bone marrow erythroblasts. Increased ERFE then suppresses hepcidin synthesis, thereby mobilizing cellular iron stores for use in heme and hemoglobin synthesis. Recent mechanistic studies have shown that ERFE suppresses hepcidin transcription by inhibiting bone morphogenetic protein signaling in hepatocytes. In ineffective erythropoiesis, pathological overproduction of ERFE by an expanded population of erythroblasts suppresses hepcidin and causes iron overload, even in non‐transfused patients. ERFE may be a useful biomarker of ineffective erythropoiesis and an attractive target for treating its systemic effects.
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Background Iron export via the transport protein ferroportin (Fpn) plays a critical role in the regulation of dietary iron absorption and iron recycling in macrophages. Fpn plasma membrane expression is controlled by the hepatic iron-regulated hormone hepcidin in response to high iron availability and inflammation. Hepcidin binds to the central cavity of the Fpn transporter to block iron export either directly or by inducing Fpn internalization and lysosomal degradation. Here, we investigated whether iron deficiency affects Fpn protein turnover. Methods We ectopically expressed Fpn in HeLa cells and used cycloheximide chase experiments to study basal and hepcidin-induced Fpn degradation under extracellular and intracellular iron deficiency. Conclusions/General significance We show that iron deficiency does not affect basal Fpn turnover but causes a significant delay in hepcidin-induced degradation when cytosolic iron levels are low. These data have important mechanistic implications supporting the hypothesis that iron export is required for efficient targeting of Fpn by hepcidin. Additionally, we show that Fpn degradation is not involved in protecting cells from intracellular iron deficiency.
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Ironing out the details of mucosal healing Anemia is a frequent complication of disorders such as inflammatory bowel disease, occurring in part as a result of increased bleeding into the intestine. Bessman et al. show that the peptide hormone hepcidin, which regulates systemic iron homeostasis, is required for intestinal repair in a mouse model of inflammatory bowel disease (see the Perspective by Rescigno). This effect was independent of hepatocyte-produced hepcidin and systemic iron levels. Instead, production of hepcidin by conventional dendritic cells was necessary and sufficient to promote local iron sequestration by macrophages, which in turn modulated the makeup of the gut microbiota to one with a more beneficial distribution of species. Science , this issue p. 186 ; see also p. 129
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Iron deficiency is common worldwide and is associated with adverse pregnancy outcomes. The increasing prevalence of indiscriminate iron supplementation during pregnancy also raises concerns about the potential adverse effects of iron excess. We examined how maternal iron status affects the delivery of iron to the placenta and fetus. Using mouse models, we documented maternal homeostatic mechanisms which protect the placenta and fetus from maternal iron excess. We determined that under physiological conditions or in iron deficiency, fetal and placental hepcidin does not regulate fetal iron endowment. With maternal iron deficiency, critical transporters mediating placental iron uptake (transferrin receptor 1, TFR1) and export (ferroportin, FPN) were strongly regulated. In mice, not only was TFR1 increased but FPN was surprisingly decreased to preserve placental iron, in the face of fetal iron deficiency. In human placentas from pregnancies with mild iron deficiency, TFR1 was increased but without a change in FPN. However, induction of more severe iron deficiency in human trophoblast in vitro resulted in the regulation of both TFR1 and FPN, similarly to the mouse model. This placental adaptation prioritizing placental iron is mediated by the iron-regulatory protein 1 and is important for the maintenance of mitochondrial respiration, thus ultimately protecting the fetus from the potentially dire consequences of generalized placental dysfunction.