Palazzo della Gran Guardia

Verona, 6 dicembre 2019

Fabiana Busti

Dipartimento di Medicina, Università di Verona

Centro di Riferimento per i Disordini del Metabolismo del Ferro

www.gimferverona.org

4° Congresso Nazionale AMGe

Geriatria e Dintorni

Un viaggio di incontri

Summary

• Basi fisiopatologiche dell’omeostasi del ferro

- Il ruolo essenziale del ferro

- La necessità di una fine regolazione

• Implicazioni della carenza di ferro nell’anziano

• Dalla fisiopatologia alla pratica clinica, verso una migliore

comprensione ed un uso più razionale della terapia marziale

Andrews, Ann Rev Physiol 2007

Il ferro è un micronutriente essenziale, ma potenzialmente tossico

Trasporto ed accumulo

di O2

Produzione di energia

Funzione di enzimi e

citocromi

facile scambio elettroni

Fe3+ Fe2+

utili proprietà redox

formazione radicali liberi O2

(Fe2++ H202 Fe3+ + OH- + OH•)

Proliferazione cellulare

ed eritropoiesi

Stress ossidativo

Morte cellulare/Apoptosi

Danno di DNA, proteine

e lipidi

Normal

Fe 4 g Fe

(toxic)

HH

Cirrhosis

HCC

diabetes

heart

failure

Skin

pigmentation

hypogonadism

Fe

(anemia)

CNS dysfunction

heart

dysfunction

Muscle

weaknessRestless

legs Synd.

Pica

IRON DEFICIENCY NORMAL IRON OVERLOAD

Il ferro è un micronutriente essenziale, ma potenzialmente tossico

La maggior parte del ferro viene riciclata ogni giorno

3-4 g

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewofironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

2 g

300 mg

1 g

3-4 mg

600 mg

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewofironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

Eritrociti

Macrofagi splenici

Precursori eritroidimidollari

L’omeostasi del ferro necessita di una stretta regolazione

Pool plasmatico

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

Epatociti

Low Fe-TF

High Fe-TF

FB (7) Verona 31.05.2019

3-4 g Fe

Equilibrio mediante

regolazione di

assorbimento e riciclo

del ferro

Assenza di

meccanismi per

l’eliminazione attiva

del ferro in eccesso

20-25 mg/die

Ferro necessario

per eritropoiesi

20-25 mg/die

Ricliclo del ferro

dai GR senescenti

1-2 mg/die

Assorbimento

duodenale

Altri

utilizzatori

Perdite fisiol.

1-2 mg/die

Camaschella C, New Eng J Med 2015

The «iron economy»

Il contenuto di ferro dell’organismo deve essere finemente regolato

3-4 g

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewofironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

2 g

300 mg

1 g

3-4 mg

600 mg

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewofironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

Eritrociti

Macrofagi splenici

Precursori eritroidimidollari

L’omeostasi del ferro necessita di una stretta regolazione

Pool plasmatico

mutant zebrafish.17 Interestingly, mutations in mitoferrin result in a

clinical disorder that is very similar to erythropoietic protoporphy-

ria caused by ferrochelatase mutations.17

Although many tissues express TFR1 at low levels, relatively

few cell types are strictly dependent on the transferrin cycle for iron

uptake. Targeted disruption of the Tfr1 gene in mice demonstrated

that most tissues develop normally without Tfr1, but erythroid

precursors, early lymphoid cells, and neuroepithelial cells require

Tfr1 for differentiation.18,19 The likely role of TFR1 in erythropoi-

esis is obvious—the transferrin cycle serves to concentrate iron in

the vicinity of DMT1 to maximize iron assimilation for hemoglo-

bin production. However, it is less clear why lymphopoiesis and

neurodevelopment should require TFR1.

In the past it was assumed that iron assimilated by erythroid

precursors was incorporated into hemoglobin, remaining within the

cells until erythrocyte senescence. Recently, however, Quigley and

colleagues20 have described a heme exporter, FLVCR, which

appears to be necessary for normal erythroid development. They

hypothesize that erythroblasts need to have a pop-off valve for

extra heme to avoid its toxicity. Targeted disruption of the mouse

gene encoding FLVCR demonstrated the importance of this protein

in vivo.21 FLVCR-null mice had a failure of definitive erythropoi-

esis, resulting in fetal demise. Interestingly, the fetuses had

craniofacial and limb deformities suggestive of Diamond-Blackfan

anemia. When the FLVCR gene was inactivated after birth the

animals developed severe, macrocytic anemia, implying that heme

export is important for normal erythropoiesis.

Regulationof intracellular ironhomeostasis

Intracellular iron homeostasis is maintained, at least in part,

through a very elegant posttranscriptional regulatory mechanism.

In 1987, investigators observed that conserved sequences in the

5 untranslated regions (UTRs) of both H- and L-ferritin mRNAs

were needed to control a ready but quiescent pool of ferritin mRNA

in the cell, which could quickly be mobilized to produce ferritin

protein when iron was abundant.22,23 Thermodynamic predictions

indicated that the UTR sequences could form stable RNA hairpins

with a characteristic secondary structure, termed iron responsive

(or regulatory) elements (IREs).24 Soon afterward it was shown that

cytoplasmic proteins, now known as iron regulatory proteins (IRPs,

formerly IREBPs), recognize and bind to the IREs.25-28

The 2 known IRPs share sequence homology but have distinc-

tive properties. At the time of its discovery, IRP1 was recognized to

bear strong similarity to aconitase, a mitochondrial enzyme of the

tricarboxylic acid cycle. Remarkably, IRP1 also has aconitase

activity, making it a prime candidate for a previously described

cytoplasmic aconitase.29,30 But the aconitase and IRE-binding

activities are mutually exclusive, providing a clue to a clever

regulatory switch. Similar to a number of other iron-containing

proteins, IRP1 incorporates an iron-sulfur cluster (4Fe•4S). The

iron-sulfur cluster forms when iron is abundant, but disassembles

when iron is scarce. Haile and Rouault showed that the aconitase

activity of IRP1 is present only when the iron-sulfur cluster is

complete; when it is not, IRP1 acts as an RNA binding protein,

recognizing IREs.31 IRP2, on the other hand, does not incorporate

an iron-sulfur cluster. Rather, its activity is regulated at the level of

protein stability. Under low iron conditions IRP2 accumulates, but

when iron is abundant it triggers IRP2 degradation.32-36 It is still not

entirely clear why it is necessary to have 2 IRPs, but recent observations

suggest that the 2 may respond differently over the physiologically

relevant range of oxygen tensions.37 They may also have somewhat

different target selectivity among IRE-containing mRNAs.

The ferritin IRE is located just upstream of the start codon for

protein translation. Muckenthaler and colleagues showed that IRP

binding sterically blocks recruitment of the small ribosomal

subunit to the initiation complex, thus preventing translation.38 As a

result, ferritin protein production is abrogated under low iron

circumstances when the small amount of intracellular iron is

needed for cellular functions. On the other hand, when iron is

abundant, translational repression is relieved and newly made

ferritin subunits assemble to provide iron storage capacity.

Figure1.Overviewof ironhomeostasis. The central

portion of the figure depicts the flow of iron into the body

(through the small intestine), to transferrin (Tf), to the

major site of utilization (the erythroid bone marrow), to

circulating erythrocytes, to tissue macrophages that

phagocytose senescent erythrocytes and recycle iron

(spleen), to storage in hepatocytes, and back to TF

through mobilization of iron stores. Cellular iron trans-

port is described in detail in the text and shown in

schematic form on the outside edges of this figure.

(A) Nonheme iron transport across an intestinal entero-

cyte. (B) Erythrophagocytosis and iron recycling in a

tissue macrophage. The aqua oval in the cytoplasm

represents a storage depot for ferroportin protein within

the cell. (C) Hepatocyte iron transport, with arrows

indicating that neither import nor export is well under-

stood. (D) Iron uptake through the transferrin cycle in

the erythoblast. Illustration by Kenneth Probst.

220 ANDREWS BLOOD, 15 JULY 2008 VOLUME 112, NUMBER 2

For personal use only. at SWETS INFORMATION SERVICES on July 8, 2008. www.bloodjournal.orgFrom

Epatociti

Low Fe-TF

High Fe-TF

FB (7) Verona 31.05.2019

3-4 g Fe

Camaschella C, New Eng J Med 2015

L’epcidina controlla assorbimento e riciclo del ferro

Equilibrio mediante

regolazione di

assorbimento e riciclo

del ferro

Assenza di

meccanismi per

l’eliminazione attiva

del ferro in eccesso

20-25 mg/die

Ferro necessario

per eritropoiesi

20-25 mg/die

Ricliclo del ferro

dai GR senescenti

1-2 mg/die

Assorbimento

duodenale

Altri

utilizzatori

Perdite fisiol.

1-2 mg/die

HEP-(atic) CIDIN (antimicrobial)

Hepcidin Ferroportin

Epcidina è l’ormone regolatore chiave dell’omeostasi marziale

• piccolo (25 aa) peptide prevalentemente prodotto dal fegato

• defensin-like (peptidi dell’immunità innata con attività antimicrobica)

• interagisce col suo recettore (ferroportina, l’unico esportatore cellulare di ferro,

ubiquitario ma altamente espresso in cellule duodenali, macrofagi ed epatociti)

Kushner JP, Blood 2010

FPN localized on cell membrane

Iron export

FPN internalized and degraded

control

Il ruolo principale di epcidina è controllare l’espressione di FPN

Il legame con epcidina determina internalizzazione di FPN e la sua degradazione

Nemeth E, Science 2004

FPN internalized and degraded

Iron export blocked

+ 1 g/ml hepcidin

Il ruolo principale di epcidina è controllare l’espressione di FPN

Il legame con epcidina determina internalizzazione di FPN e la sua degradazione

Nemeth E, Science 2004

FPN localized on cell membrane

Iron export

control

Milza

Fegato

Duodeno

Hepcidin

Fpn

Fpn

Fpn

Plasma

Fe-Tf

Midollo osseo e

altri utilizzatori

di ferro

L’asse epcidina-ferroportina regola il flusso di ferro nel plasma

I livelli di epcidina influenzano la distribuzione del ferro nell’organismo

Fpn

Macrofago

Fpn

Enterocita

Epcidina è regolata da molteplici fattori

Girelli D et al., Blood 2016

Prodotta in risposta all’ del Fe, per regolare negativamente la sideremia

EPCIDINA

Summary

• Basi fisiopatologiche dell’omeostasi del ferro

- Il ruolo essenziale del ferro

- La necessità di una fine regolazione

• Implicazioni della carenza di ferro nell’anziano

• Dalla fisiopatologia alla pratica clinica, verso una migliore

comprensione ed un uso più razionale della terapia marziale

Impatto clinico della carenza di ferro (ID) nell’anziano

• Alta prevalenza (10-40%), causa frequente di anemia nell’anziano

• Cut-off diagnostici più elevati: ferritina <45-50 ng/ml

• Complicanze: decadimento cognitivo, depressione, rischio di caduta, malattie CV,

ospedalizzazione, morte

• Associata ad outcomes avversi anche in assenza di anemia

• Trattamento migliora outcomes

Busti F et al., Front Pharmacol 2014

Girelli D et al, HemaSphere 2018

Perdite GI alte

Non infrequentemente carenza di ferro assoluta e funzionale coesistono

La carenza di ferro può essere «assoluta» (AID) e/o «funzionale (FID)

Malassorbimento

Perdite GI basse

Inadeguata assunzione

Scompenso cardiaco

Malattie renali croniche

Disordini autoimmuni

± fattori iatrogeni (es. PPI, antitrombotici/anticoagulanti)

=depositi di ferro “bloccati"

Infezioni

AID =depositi di ferro esauriti XFID

EPCIDINA

Summary

• Basi fisiopatologiche dell’omeostasi del ferro

- Il ruolo essenziale del ferro

- La necessità di una fine regolazione

• Implicazioni della carenza di ferro nell’anziano

• Dalla fisiopatologia alla pratica clinica, verso una migliore

comprensione ed un uso più razionale della terapia marziale

La terapia orale è semplice, facilmente accessibile e sicura

Modificato da Veuthey T et al., Front Pharmacol 2014

Lume

intestinale

Sangue

• Per molto tempo

considerata terapia di 1°

scelta nei soggetti con

IDA lieve-moderata

(Hb >8 g/dL)

• Sali di ferro e

complessi

polisaccaridici, in

forma ferrosa (2+) o

ferrica (3+)

Assorbimento attraverso la via del

ferro non-eme (scarsamente

efficiente), mediante trasportatore

specializzato (DMT1)

Assorbimento favorito da ambiente

acido (potenzialmente ↑ da

vitamina C 250-500 mg/die, ↓ da

PPI)

• I preparati maggiormente utilizzati sono il ferro

gluconato, il ferro fumarato e il ferro solfato

Attenzione al

contenuto di

ferro elementare

(80-100 mg)

Modificato da Parmanand BA et al, J Nutr Biochem, 2019

Ferro orale

80-90% ferro non

assorbito raggiunge

il colon

Tolkien et al., 2015

Danno direttoNausea, vomito, dolore

addominale, diarrea o stipsi

nel 30-70% dei soggetti

Modificazioni microbioma

intestinale ↓ Lactobacilli e Bifidobacteria

↑Enterobacteria

= Tempi di assunzione

prolungata (almeno 3-6

mesi) per ottenere non

solo normalizzazione

dell’Hb, ma anche

replezione dei depositi

(ferritina >100 ng/ml)

Solo 10-20% viene

assorbito

Gli effetti avversi, soprattutto GI, compromettono la compliance alla terapia

Ferro orale

Molteplici fattori contribuiscono alla refrattarietà alla terapia orale

• Sospensione prematura trattamento

• Mancata compliance

• Gastriti atrofica, autoimmune, HP-correlata

• Chirurgia bariatrica

• Malattia celiaca (o altro malassorbimento)

• (Iron Refractory Iron Deficiency Anemia)

• Condizioni infiammatorie, anche

«subcliniche» (es. scompenso cardiaco,

CKD)

• Perdite troppo abbondanti

Mancata compliance

Inadeguata acidità gastrica

Mucosa danneggiata

Refrattarietà alla terapia orale =

incremento di Hb < 1 g/dl dopo 3

settimane di trattamento

Tf plasmatica

Fpn

Enterocita

xBlocco assorbimento

causato da ↑ epcidina

50 donne pre-menopausa

ID (ferritina < 20 ng/ml), non anemiche

Dose crescenti (40-60-80-120-240 mg)

Ferro per os per 2 giorni consecutivi

La terapia con ferro influenza i livelli di epcidina

EPCIDINA

Replete iron stores

1. Incremento acuto di epcidina

persistente per 48 ore

+ 9 h picco

epcidina

+ 4 h

picco TSAT

+24 h

+48 h

normalizzazione

epcidina

50 donne pre-menopausa

ID (ferritina < 20 ng/ml), non anemiche

Dose crescenti (40-60-80-120-240 mg)

Ferro per os per 2 giorni consecutivi

TSAT= saturazione

transferrina

Il picco di epcidina indotto dalla terapia con ferro ne riduce l’assorbimento

1. Incremento acuto di epcidina

persistente per 48 ore

2. Ridotto assorbimento della

seconda dose (-35-45%)

2° Dose 60 mg

Assorbimento -36%

50 donne pre-menopausa

ID (ferritina < 20 ng/ml), non anemiche

Dose crescenti (40-60-80-120-240 mg)

Ferro per os per 2 giorni consecutivi

Il picco di epcidina indotto dalla terapia con ferro ne riduce l’assorbimento

Tutti i giorni per

14 giorni

Dì alterni per

28 giornip

Frazione ferro assorbita (%) 16.3 21.8 0.0013

Quantità ferro assorbita (mg) 131 mg 175 mg 0.0010

Ferritina 13.8→28.3 13.8→23.6 0.058

Frequenza effetti avversi GI → 33% più bassa gg alterni (p=0.57)

20 donne

ID, non anemiche

Ferro orale 60 mg

Somministrazione per 14 giorni consecutivi

oppure per 28 giorni a dì alterni

La somministrazione a dì alterni sembra ottimizzare l’assorbimento di ferro

Necessità di

conferma in

pazienti

anemici

Mancano dati

sugli esiti a

lungo termine

Ferritina umana

1947: Fe Saccaridico

1954: Fe-Destrano (HMW)

1991: Fe-Destrano (LMW)

1999: Fe-Gluconato

2000: Fe-Sucrosio

>2009: Ferumoxytol

Fe-isomaltoside

Fe-carboximaltosio

“Preparati di

III generazione”

8-2

5 n

m

Fe3+

oxyhydroxide(CORE)

Carboidrati (GUSCIO)

Caratteristica unica di

ciascun composto, ne

influenza:

• Immunogenicità

• Stabilità

Moderate risk

High risk

Very low risk

1932: FeOH

Modificato da Girelli D et al., Int J Hematol 2018

I più recenti preparati di ferro EV sono efficaci e sicuri

Ferro EV <1:200000

AEs gravi

Trasfusioni 1:21000

Avni T et al., Mayo Clin Proc 2015

3° generazione

FCM

Si basa su

peso e

livelli di Hb

Max 1

g/settimana

Hb Peso

g/dl <35 kg 35-70 kg >70 kg

<10 500 mg 1500 mg 2000 mg

10-14 500 mg 1000 mg 1500 mg

>14 500 mg 500 mg 500 mg

Determinazione della dose (schema semplificato)

Modificato da Girelli D et al., Int J Hematol 2018

La farmacocinetica del ferro EV è diversa da quella del ferro orale

Terapia orale Terapia EV

Integrità

mucosa

gastro-

duodenale

Assorbimento

bloccato da

epcidina

Va direttamente

in circolo,

captata dai

macrofagi

Meno

influenzata dai

livelli di

epcidina

Modificato da Girelli D et al.,

Int J Hematol 2018

Principali indicazioni alla terapia marziale EV

Modificato da Girelli D et al., Int J Hematol 2018

Take home messages:

• La carenza di ferro nell’anziano è un problema rilevante

che non andrebbe trascurato

• Terapia orale presenta molteplici limitazioni, che

potrebbero essere superate da schemi di trattamento più

«fisiologici» e meglio tollerati (dì alterni?)

• I «moderni» preparati di ferro EV permettono correzione dell’ID in 1-2 dosi

con ottima tollerabilità ed efficacia (anche in condizioni infiammatorie)

• Reazioni avverse al ferro EV sono estremamente rare e solo eccezionalmente

pericolose (molto inferiori a quelle trasfusionali)