Autoimmunity Reviews 11 (2012) 883–889

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Autoimmunity Reviews

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Review

Iron levels in polarized macrophages: Regulation of immunity and autoimmunity

Stefania Recalcati a,⁎, Massimo Locati b,c, Elena Gammella a, Pietro Invernizzi d, Gaetano Cairo a

a Department of Human Morphology and Biomedical Sciences “Città Studi”, University of Milan, Milan, Italyb Department of Translational Medicine, University of Milan, Milan, Italyc Laboratory for Leukocyte Biology, IRCCS Istituto Clinico Humanitas, Rozzano, Italyd Center for Autoimmune Liver Diseases, IRCCS Istituto Clinico Humanitas, Rozzano, Italy

Abbreviations: TAM, tumor associatedmacrophages; Lchronic diseases; BMP, bone morphogenic protein; HJV, h⁎ Corresponding author at: University of Milan Schoo

Italy. Tel.: +39 0250315330; fax: +39 0250315338.E-mail address: stefania.recalcati@unimi.it (S. Recalc

1568-9972/$ – see front matter © 2012 Elsevier B.V. Alldoi:10.1016/j.autrev.2012.03.003

a b s t r a c t

a r t i c l e i n f o

Article history:Received 2 March 2012Accepted 8 March 2012Available online 16 March 2012

Keywords:IronPolarized macrophagesInnate immunityFerroportinAnemia of chronic diseaseInflammation

Although the hallmark of autoimmune diseases remains the generation of autoantigen-specific lynfocytic cellresponse, growing evidence is showing a key role for macrophages in a number of autoimmune diseases.Macrophages are characterized by phenotypical and functional heterogeneity. Different immunological sig-nals, coming from systemic blood circulation or from microenvironment, polarize macrophages to classical(M1) or alternative (M2) phenotypes. Iron accumulation in M1 macrophages is a well known bacteriostaticmechanism and one of the mechanisms at the basis of anemia associated to chronic inflammation. Moreover,some recent data suggest that iron accumulation in macrophages can directly activate macrophages to pro-inflammatory M1 phenotype, highlighting a putative role of macrophage iron retention in the pathogenesisof chronic inflammatory and autoimmune diseases. Conversely, iron content is low in M2 macrophages, prin-cipally due to increased iron release, and increased availability of iron in the extracellular milieu supported byM2 macrophages could influence the growth rate of adjacent cell and thus play an important role in tumorgrowth and tissue remodeling.In this review we summarize the molecular mechanisms sustaining differential iron metabolism in polarizedmacrophages, discuss the relevance of this metabolic signature in chronic inflammatory and autoimmunediseases, and finally focus on potential therapeutic implications rising from a better understanding of under-lying molecular mechanisms.

© 2012 Elsevier B.V. All rights reserved.

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8832. Heterogeneity in macrophage differentiation and polarization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8843. Iron retention in M1 macrophages . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 884

3.1. Local effects: pathogenic role in chronic inflammation and autoimmune diseases . . . . . . . . . . . . . . . . . . . . . . . . . . 8843.2. Systemic consequences: anemia of chronic diseases, a common condition in autoimmune diseases . . . . . . . . . . . . . . . . . . 885

4. M2 iron releasing-macrophages: consequences on microenvironment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8855. Therapeutic implication for chronic inflammation and autoimmune diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8876. Concluding remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 887Take-home messages . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 887Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 887References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 887

884

cn2, lipocalin 2; DMT1, divalentmetal transporter 1; Ft, ferritin; IRP-2, iron regulatory protein 2; Fpn, ferroportin; ACD, anemia ofemojuvelin; TfRs, soluble transferrin receptor; Hb, hemoglobin; ESA, erythropoietin-stimulating agents; EpoR, Epo receptor.l of Medicine, Department of Human Morphology and Biomedical Sciences “Città Studi”, Via Mangiagalli 31, 20133, Milano,

ati).

rights reserved.

884 S. Recalcati et al. / Autoimmunity Reviews 11 (2012) 883–889

1. Introduction

For long time, infections have been postulated to play a role in thepromotion of autoimmune diseases [1]. Infection may indeed damagetissue as well as provide the adjuvant stimulus needed for the devel-opment of autoimmune diseases [2–4]. The innate immune responseplays a central role in the immunological response to infection also bydirecting adaptive immunity. Thus, understanding the pathophysio-logical mechanism underlyning the modulation of the immune re-sponse to infection can shed light on the pathogenic mechanismsinvolved in autoimmune diseases.

Although the hallmark of autoimmune diseases is the generationof autoantigen-specific T and B cell response [5–8], macrophagesare important for immune responses and are highly present inthe acute or chronic inflammatory milieu. New data suggest a directrole of these cells in the pathogenesis of autoimmune diseases [9–12].Macrophages can be differently polarized in response to differenttypes of infections or different kinds of cytokines present in the micro-enviroment, and could thus profoundly influence immune responsesand tissue homeostasis [13,14], but little is known about the state ofmacrophage activation in autoimmunity [8,15,16]. Iron metabolismalso plays a key role in the context of innate immune response:i) microbes need iron for proliferation and pathogenicity; ii) iron affectscell-mediated immune function and thus host response towardpathogens; iii) cytokines and acute-phase proteins regulate ironmetabolism genes under inflammatory conditions. Moreover, we haveto underline that on the one hand the polarization of macrophagescan have important effects on iron metabolism, but on the other handiron can influence directly macrophage polarization.

The aim of this review is to highlight the role of iron in shaping thepolarized macrophage phenotype, and the effect of macrophagepolarization on iron metabolism. Our understanding of these mecha-nisms has implication for new possible pathogenic mechanisms in-volved in autoimmune diseases.

2. Heterogeneity in macrophage differentiation and polarization

Resident macrophages are present in organs constitutively, in theabsence of overt inflammation, and perform trophic as well as ho-meostatic roles in the removal of apoptotic cells, serving as sentinelsof injury and infection. Tissue macrophages can replicate locally, butare terminally differentiated, turning over at different rates, depend-ing on particular tissue environmental stimuli that function throughspecific growth and differentiation factors, their receptors, signalpathways, and transcriptional factors [13].

Macrophages migrate to local sites of injury and infection contrib-uting to acute and chronic inflammation, locally and systemically. Inaddition they acquire enhanced cytotoxic, antimicrobial, and in-hibitory activities, initiate repair, and resolve inflammation [17]. Tis-sue macrophages originate from peripheral blood monocytes andreveal a substantial heterogeneity of phenotypes and specialization[13,18,19]. An important element shaping macrophage heterogeneityis the microenvironment, both under homeostatic conditions, inwhich the hosting tissue profoundly influences macrophage differen-tiation, and in the contest of inflammatory or immune response, inwhich different kinds of cytokines can lead to a wide range of macro-phage polarized states [13,14,19]. In particular macrophages activat-ed with IFN-gamma alone or in combination with LPS, (whichinduce TNF-alfa) have been classified as classically activated (M1)macrophages characterized by high microbicidal capacity and secre-tion of pro-inflammatory cytokines. Other immune stimuli can leadto profoundly different functional phenotypes. These include thealternatively activated macrophages (M2) as a consequence of stimu-lation with IL-4 and IL-13. M2 macrophages show more phagocyticactivity, high expression of scavenging, mannose and galactosereceptors, and a phenotype characterized by low expression of IL-12

and high expression of IL-10. In general, these cells participate inpolarized Th2 responses, help with parasite clearance, dampeninflammation, promote tissue remodeling and have immuno-regulatory functions [13,19,20]. Furthermore, in several tumors, thetumor associatedmacrophages (TAM) that are involved in tumor pro-gression show a number of features typical of M2 macrophages[21,22].

3. Iron retention in M1 macrophages

3.1. Local effects: pathogenic role in chronic inflammation andautoimmune diseases

It is well known that M1 macrophages have enhanced microbici-dal capacity, secrete high levels of pro-inflammatory cytokines, andproduce great amount of oxygen and nitrogen radicals to increasetheir killing activity, as required in the early phases of tissue repair[17]. Moreover, inflammatory macrophages are characterized by in-creased iron retention, which is generally viewed as a bacteriostaticmechanism. Mammals and microorganisms have evolved a varietyof molecules to bind different forms of iron, with the purpose ofboth acquiring this essential metal, and at the same time, to limitiron to the opponent. The control of this strategic resource is animportant element of the host–pathogen interaction and of theso-called nutritional immunity. A largely studied example of thesemolecules is lipocalin 2 (Lcn 2), a member of the lipocalin familythat binds bacterial siderophores, thus impeding bacterial ironsequestration and hence acting as a potent bacteriostatic agentagainst various Gram-negative microorganisms [23]. In chronicinflammation iron is acquired by macrophages most prominentlythrough erythrophagocytosis [24] and the transmembrane import offerrous iron by the protein divalent metal transporter 1 (DMT1)[25]. In previous studies, an increase of iron retention in M1 inflam-matory macrophages has been demonstrated by showing a stronginduction of the iron storage protein ferritin (Ft) [26–29] that wasmainly caused by an accompanying decrease in the activity of theiron regulatory protein 2 (IRP-2), a sensor of intracellular iron pool[26,27]. However, during inflammation the reticuloendothelial ironsequestration seems to be principally due to a decrease of iron releasefrom these cells. Inflammatory stimuli trigger macrophage ironretention also by down-regulating the expression of iron exporterferroportin (Fpn), thus blocking the release of iron from these cells[30,31]. The transmembrane protein Fpn, which is an essential com-ponent of systemic iron homeostasis [32], is the major or only ex-porter of non-heme iron [33–35] and is expressed at high levels onduodenal enterocytes, placenta, hepatocytes and macrophages. Fpn ex-pression is controlled at multiple levels: some studies showed that inmacrophages Fpn was negatively regulated at transcriptional level byinflammatory mediators [36,37], but it is known that this protein isalso regulated at post-transcriptional level by the IRP system (see [38]for recent review). Moreover, the blockade of macrophage iron releaseseems to bemainly due to the interaction between the acute phase pro-tein hepcidin and Fpn [39–42]. The increase in circulating hepcidintriggered by inflammatory cytokines causes the internalization anddegradation of Fpn [43], thus blocking iron release from macrophages.The inhibitory effect of systemic hepcidin on Fpn expression (seebelow) is further aggravated in inflammatory monocytes/macrophagesby the autocrine formation of hepcidin stimulated by cytokines [44].

In pathogen-free condition of chronic inflammation and also inautoimmune diseases, macrophage iron retention can have a patho-genetic role because macrophages could produce minute amountsof hepcidin to control cellular iron egress in an autocrine fashion[45]. In this regard, a recent work suggested that the increased irondeposition in macrophages found in human chronic venous leg ulcersinduces an unrestrained pro-inflammatory M1 macrophage popula-tion, thereby contributing to chronic inflammation [46]. In these

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lesions iron accumulation in macrophages is the consequence oferythrophagocytosis of extravasated erythrocytes and seems torepresent one of the local factors that influence the contribution ofmacrophages toward either wound healing or chronic inflammation.High intracellular iron levels lead to persistence of an unrestrainedpro-inflammatory M1 macrophage population with an incompleteswitch to anti-inflammatory M2 macrophages that eventually leadsto tissue breakdown and impaired tissue restoration. Moreover, ithas been shown that the increased intra-macrophage iron retentioncaused by hepcidin-mediated Fpn downregulation could be consid-ered as a pro-inflammatory signal that facilitates expression of cyto-kines such as TNF-alfa [47]. Accordingly, a recent paper showed thatiron retention in macrophages lacking Fpn stimulates the expressionof pro-inflammatory cytokines and the innate immune answer toresponse in vivo [48]. These observations are consistent with earlierreports demonstrating that increased iron levels can enhance signalingthrough theNF-kBpathway [49]. Other results favor the hypothesis thatintracellular iron can activatemacrophages and promote the underlyinginflammatory diseases. In particular, it has been shown that in athero-sclerosis systemic and local inflammation could increase the local pro-duction of hepcidin. In the atherosclerotic plaque, increased hepcidinmay cause iron accumulation in macrophages that ingest erythrocytesand apoptotic/necrotic cells. The hepcidin-mediated accumulation ofiron in plaque macrophages and the resulting inflammation couldconstitute a self-amplifying process that promotes atherosclerosis(Kautz L., BioIron, Vancouver 2011).

3.2. Systemic consequences: anemia of chronic diseases, a commoncondition in autoimmune diseases

The development of a dysregulation of iron homeostasis, with in-creased retention of iron within cells of the reticuloendothelial sys-tem, is one of the molecular mechanisms at the basis of the anemiaof chronic diseases (ACD), a condition often present in patients withinfections, tumors, but also auto-immune disorders [50]. ACD seemto be the result of pro-inflammatory cytokine-mediated effects on dif-ferent targets that lead to impairment of erythrocyte progenitor for-mation, inadequate synthesis and function of erythropoietin andiron retention within the reticuloendothelial system, the latter seem-ingly having a key role [50,51]. The inappropriate storage of iron intothe reticuloendothelial system causes a subsequent limitation of theavailability of the metal for erythroid progenitor cells which, in asso-ciation with inadequate function of erythropoietin, eventually leadsto iron-restricted erythropoiesis.

The identification of hepcidin provided a better understanding ofthe relationship between the immune response and iron homeostasisand ACD. As mentioned, hepcidin binds to Fpn inducing its internali-zation and degradation and thus regulates the flow of iron, mainlyfrom duodenum, reticuloendothelial cells and liver, into plasma. Byso doing, hepcidin regulates the distribution of iron in the body, andalso the availability of iron for the bone marrow [42,52]. Hepcidintranscription is normally stimulated by increased serum iron, in partvia the bone morphogenic protein (BMP) signaling pathway [52].BMPs, members of the TGF-beta superfamily, bind BMP receptorswith coreceptors such as hemojuvelin (HJV). Loss of function of HJVin humans and mice [53,54] and of BMP6 [55,56] in mice, leads to in-appropriately low hepcidin production and consequent iron overload.Recently, the membrane bound serine protease matriptase 2 has beenshown to degrade membrane anchored HJV thereby reducing hepci-din expression [57]. However, hepcidin expression is known to be in-duced also by inflammatory mediators, including LPS and IL6 [58],which activate hepcidin transcription through the STAT3 pathway[59]. Under inflammatory conditions, high hepcidin levels inhibitFpn-dependent iron release from macrophages and iron absorptionfrom duodenum thus restricting iron availability for hemoglobin syn-thesis and causing anemia. Actually, transgenic overexpression of

hepcidin resulted in severe iron deficiency anemia in mice, as a resultof inhibition of duodenal iron absorption and iron export frommacro-phages [60], providing the proof of principle of the role of hepcidin inthe pathogenesis of ACD.

From a clinical point of view ACD is characterized by a moderatenormocytic, normochromic anemia that develops in patients with a cel-lular immune activation; in particular it can be associatedwith differentautoimmune diseases, including rheumatoid arthritis, vasculitis, sys-temic lupus erythematosus, and inflammatory bowel diseases. Regard-ing the underlying dysregulation of ironmetabolism, it should be notedthat pro-inflammatory cytokines induce local iron sequestration inmacrophages as a bacteriostatic mechanism (see above), but circulatingcytokines induce iron sequestration also in spleen macrophages andKupffer cells (Fig. 1). Therefore, reticuloendothelial cells are probablythe main responsible for systemic hypoferremia from a quantitativeviewpoint. Understanding of the molecular mechanism at the basis ofACD is very important in clinical setting for a correct diagnosis andthe development of new therapies. From a diagnostic point of view,the major goal is to differentiate ACD from iron deficiency anemia(IDA) (see Table 1). The most important iron parameter for discrimina-tion between these two forms of anemia is the serum Ft, which is usu-ally very low in IDA but is not reduced or even increased in ACD [50].Cytokines have a role in this activation because they have been shownto activate the transcription of Ft [28,29]. Another important iron pa-rameter is the soluble transferrin receptor (TfRs), a truncated fragmentof the TfR that increases in iron deficiency, when iron availability forerythropoiesis is low. In ACD TfRs levels are normal because its expres-sion is inversely related to cytokines [50]. Evaluation of serum Ft andTfRs allows the clinician to distinguish between patients with ACDalone (normal or high Ft and low TfR) from thosewith ACD and iron de-ficiency (low Ft, high TfR). The correct diagnosis of these different formsof anemia is very important for the choice of an appropriate therapy.

4. M2 iron releasing-macrophages: consequenceson microenvironment

A role in regulating adaptive immunity and in the control of cellgrowth and tissue repair has been suggested for alternative polarizedM2 macrophages. These cells dampen pro-inflammatory cytokinelevels, secrete components of the extracellular matrix, and may be es-sential for late phase of tissue repair [17]. Concerning iron metabo-lism, it has been shown that M2 macrophages have lower ironlevels than M1 macrophages [31] and it has been demonstrated thatit is mainly due to the upregulation of Fpn that leads to more efficientFpn-dependent iron export [31,61]. Low intracellular iron levels, asalready mentioned above, can influence macrophage populations interms of cytokine production. A study of inflammatory responses ina murine model of hemochromatosis has shown that low intracellulariron levels in macrophages of HFE mutant mice impair the translationof IL-6, TNFα, and consequently attenuate the inflammatory response[62].

InM2macrophages an induction of hemeoxygenase (HO-1) expres-sion that seems to play a role in the iron release-prone phenotype ofthese cells has been also found [31]. This is in linewith findings showingthat HO-1 favors iron release [63], and thatmassive iron overload is pre-sent inHO-1 knock-outmice [64]. HO-1 has also a role in the catalysis ofheme oxidative degradation. In turn, heme by-products seem to havepositive effects on macrophages and their microenvironment: heme,which has a pro-oxidant and pro-inflammatory activity, is transformedin biliverdin and subsequently converted into bilirubin, endowed withanti-inflammatory and anti-oxidant properties, and CO, which pro-motes vasodilation and angiogenesis via several mechanisms [65].Moreover a recent study showed that monocytes polarized toward aM2 phenotype by exposure to glucocorticoids are characterized by en-hanced expression of the hemoglobin (Hb) scavenger receptor CD163,increased Hb uptake, increased heme-degradation and export of

he

MicroorganismAutoimmune disregulation

hepcidin

Iron sequestration in M1

Anemia of chronic disease

Persistent iron –dependent activationof M1 macrophages

Impaired restoration in chronic disease

LPSIFN-γIL-6IL-1

Ferritin(iron storage)

Duodenal iron absorption

Ferroportin(iron exporter)

erythrophagocytosis

Fe2+

labile iron pool

liver

M1machrophage

DMT1(iron importer)

Defence against invadingpathogen

hepcidin

Fig. 1. Iron sequestration in M1 polarized macrophages.Different pro-inflammatory signals (in red) can stimulate iron retention in macrophages. In particular pro-inflammatorycytokines stimulate a) macrophage erythrophagocytosis, b) iron uptake through the iron transporter DMT1, c) liver production of acute-phase protein hepcidin, that inhibits fer-roportin expression and iron excretion, and d) production of autocrine hepcidin frommacrophages. Increased iron levels into macrophages lead to different consequences (in blue):a) anemia of chronic diseases (ACD), b) defense against invading pathogen, and c) impaired restoration in chronic diseases.

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heme-derived iron [66]. In particular, the interactive effect of glucocor-ticoids and Hb seems to shift the monocyte phenotype toward en-hanced iron-recycling and antioxidant activity [66]. These datasupport novel activities of glucocorticoid therapy in autoimmune dis-eases, such as autoimmune hemolytic anemia in which a large quantityof freeHb is present into circulation, that could support adaptive protec-tive response to extracellular Hb exposure independent of commonanti-inflammatory and immunosuppressive mechanisms. On the otherhand, M2 anti-inflammatory cytokines, like IL-10, present in the latephases of inflammatory process, are responsible for the release of ironfrom themacrophages to the extracellular space [30] and could improvefibroblast proliferation and tissue repair.

It has been shown that M2 macrophages are necessary for the re-generation of acutely injured skeletal muscle [67] a condition inwhich large amounts of the metal are essential to build new,

Table 1Iron parameters for diagnosis of anemia of chronic diseases (ACD) and iron deficiencyanemia (IDA).

Iron parameters ACD IDA

Serum iron concentration Reduced to normal ReducedTranferrin levels Reduced to normal IncreasedTransferrin saturation Reduced to normal ReducedFerritin Normal to increased ReducedSerum transferrin receptor Normal IncreasedCytokines (TNF, IL-1, IL-6) Increased Normal

functional myofibers. In this regard, Corna et al. suggested that ironrelease fromM2macrophages could play a relevant role in muscle re-pair [61]. Moreover, the greater iron releasing capacity of M2 macro-phages may affect other cells in the microenvironment. In particular,TAMs that infiltrate tumors, and are involved in tumor growth, pro-gression and invasion, also acquire a polarized M2-like phenotype.Iron is an essential component of many proteins involved in cellgrowth and replication, and neoplastic cells require more iron thannormal cells because they generally proliferate more rapidly, as indi-cated by their higher expression of TfR1 and greater transferrin-bound iron uptake. Therefore, in an in vivo situation of tumor growth,a high level of iron release from TAM may provide an unrestrictedsource of iron for the multiplication of tumor cells and may thus rep-resent a further and previously unknown mechanism underlyingtheir tumor-promoting activity.

As it regards the role of iron in the tumormicroenvironment, a possi-ble role in tumor growth for the bacterial siderophore binding proteinLcn2 has been recently shown (see above) [68]. Recent reports providedevidence that mammalian cells also produce siderophore-like moleculesthat circulate bound to Lcn 2 [69]while other studies showed an increaseof Lcn2 expression in cancer [70]. Moreover, of critical importance arethe observations showing that blocking this molecule's expression inseveral types of cancer delays or even abrogates tumorigenesis [71].The fact that Lcn2 transcription may be regulated by TNFα suggests theexistence of autocrine signaling in the production of Lcn2 in the tumormicroenvironment, which can partially explain why Lcn2 is stronglyupregulated as tumor progresses. Lcn2 is a versatile molecule with twoapparent roles: a beneficial one in innate immunity that protects against

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bacterial pathogens (see above), and a detrimental one when co-optedby cancer cells into a tumor-promoting function.

Recent finding in animal models suggests that M2 polarized mac-rophages can exacerbate autoimmune diseases by presenting self an-tigens taken up by scavenger receptors in response to tissue damageand by contributing to fibrosis and immunocomplex-mediated pa-thology [72]. Evidence is beginning to emerge that M2 may be pa-thogenetic in autoimmune encephalomyelitis, arthritis, and lupus[72]. However, few studies addressing autoimmune disease have spe-cifically examined the role of M2 in the pathogenesis of the disease.

5. Therapeutic implication for chronic inflammation andautoimmune diseases

Up to now, considering the role of intracellular iron for macro-phages activation, it seems that blocking hepcidin expression mayrepresent a novel approach to control inflammation. Iron chelationtherapy with agents such as desferrioxamine has been tried previous-ly for chronic inflammatory conditions, such as rheumatoid arthritis,and limited success has been reported in some studies [73]. Inhibitionof hepcidin expression or function could have advantages over thenonspecific reduction of iron levels afforded by chelating drugssince its effects would be confined largely to cells with high expres-sion of Fpn, such as macrophages. In addition, the increase in Fpn ex-pression caused by a reduction in circulating hepcidin levels wouldincrease iron availability and be helpful to correct the anemia associ-ated with many longstanding inflammatory states [74]. As EPO hasbeen demonstrated to reduce the expression of the principal iron reg-ulator, hepcidin [40,75], it will be of interest to verify whether it canhave an effect on IL-6 inducible production of hepcidin in macro-phages [76].

It is obvious that the best therapy for ACD is the treatment or cure ofthe underlying inflammatory disease. Alternative therapeutic measuresare aimed to increase Hb levels in ACD patients but the impact of suchinterventions on iron overload in the reticuloendothelial system, immu-nity, and most importantly the underlying disease, is largely unknown.Blood transfusion can be readily used for rapid correction of severe ane-mia. Iron administration is a possible choice for the therapy of lesssevere ACD, but it should be remembered that iron is very poorlyabsorbed in ACD, mainly because of hepcidin-mediated down-regulation of duodenal Fpn (see above); therefore, i.v. iron administra-tion is a better strategy for iron therapy. Moreover, it is important tonote that iron should not be used if infections or cancer is the cause ofACD or in presence of serum Ft levels>100 ng/mL [77]. In these cases,iron may have adverse effects, such as favoring proliferation of patho-gens or impairing immune function. Additionally, the metal wouldprobably be sequestered into the reticuloendothelial system, thus notreaching erythroid cells, and/or it may cause tissue damage via forma-tion of toxic radicals by the Fenton reaction [50]. However, in autoim-mune diseases, iron may inhibit pro-inflammatory immune effectorpathways, thus reducing disease activity [77].

For the choice of iron therapy, it is important to formulate a correct di-agnosis of ACD alone or ACD associated with IDA (see Table 1) becausethese patients need contrasting therapies; in particular iron is needed inACD/IDA but not in patient with ACD alone [77]. Another therapeuticstrategy effective in increasing hemoglobin levels in ACD patients withcancer, infections, and autoimmune disorders is the utilization oferythropoietin-stimulating agents (ESA). In this case the response rateto treatment depends on underlying disease, stage, immune activation,and iron availability. Combination therapy with ESA and iron seems tobe a good choice for conditions with ACD/IDA [78,79]. At present, someuncertainties remain on the safety of this therapeutic approach, basedon recent studies indicating increased mortality in certain patient groupsin associationwith ESA therapy; in this context, the biologic role of Epo re-ceptor (EpoR) on tumor cells remains undefined [78,79].

In the future, in consideration of the pathogenesis of this disease, themajor emerging therapies seem to be: iron chelation, (anti)-cytokineadministration, EpoR modulation, and hepcidin/Fpn agonist/antagonisttherapy. In this regard, it should be considered that since EPO per se in-hibits promptly and remarkably serum hepcidin levels [40,75], ESAtreatment could have also collateral beneficial effects on iron absorptionin these patients.

6. Concluding remarks

In this review we showed the role of iron in the modulation of im-mune system and its possible involvement in the pathogenesis of au-toimmune diseases. In particular, regarding the relationship betweeniron and macrophage polarization, evidence suggests that in chronicand autoimmune diseases the strong production of cytokines couldlead to the persistence of an unrestrained pro-inflammatory M1 mac-rophage population with an incomplete switch to anti-inflammatoryM2 macrophages. In this way, macrophages accumulate iron leadingto maintenance of an active pro-inflammatory profile that can accom-pany autoimmune diseases. On the other hand, M2 polarized macro-phages can exacerbate autoimmune disease especially by releasinghigh amount of iron and contributing to fibrosis. Moreover, from asystemic point of view iron retention in reticuloendothelial cells isthe principal pathogenetic cause of ACD, a condition often associatedto autoimmune diseases. A better understanding of the role of iron inmacrophage polarization and autoimmunity may pave the way to thedevelopment of innovative therapeutic strategies.

Take-home messages

• Iron retention in pro-inflammatory M1 macrophages has bacterio-static effect.

• Iron accumulation in M1 macrophages has pathogenetic role inchronic inflammatory diseases.

• Anemia associated to inflammatory diseases is due to iron retentionin M1 macrophages.

• Iron release from M2 macrophages has effect on cells in themicroenviroment.

• Understanding of the role of iron in macrophages leads to potentialtherapeutic implications.

Acknowledgments

The authors are thankful to Prof. A. Mantovani for insightful com-ments and suggestions.

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Endoplasmic reticulum mammalian chaperones are potential autolupus erythematosus

Cytosolic and endoplasmic reticulum (ER) resident chaperones, includinin recognition and binding of non-native misfolded proteins, under physheavy chain binding protein (BiP), glucose-regulated protein 94 (Grp94)autoimmune diseases.In this paper, Weber et al. (Rheumatology 2010;49:2255-63) establishetiters to BiP, Grp94 and calnexin in patients with various systemic autoimserved that autoantibodies to ER resident chaperones are significantly inpus erythematosus (SLE), present early during the disease course, and csuch as rheumatoid factor or anti-CCP antibodies, nor with HLA-DR1/DRmune response against ER chaperones is detectable and may have a patAnna Ghirardello

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antigens in patients with rheumatoid arthritis and systemic

g heat shock proteins, are intracellular scavenger molecules involvediological or stress conditions. Some of them, such as immunoglobulinand calnexin have been evoked as potential neoantigens in systemic

d analytically specific ELISA tests for measuring serum autoantibodymnue diseases and healthy individuals, as controls. The authors ob-creased in patients with rheumatoid arthritis (RA) and systemic lu-orrelated one to each other but not with RA-specific autoantibodies,4 alleles. In conclusion, the present findings suggest that an autoim-hogenetic role in RA and SLE.