The Iron age of host–microbe interactions
2015; Springer Nature; Volume: 16; Issue: 11 Linguagem: Inglês
10.15252/embr.201540558
ISSN1469-3178
AutoresMiguel P. Soares, Günter Weiß,
Tópico(s)Heme Oxygenase-1 and Carbon Monoxide
ResumoReview16 October 2015free access The Iron age of host–microbe interactions Miguel P Soares Corresponding Author Miguel P Soares Instituto Gulbenkian de Ciência, Oeiras, Portugal Search for more papers by this author Günter Weiss Corresponding Author Günter Weiss Department of Internal Medicine VI, Infectious Diseases, Immunology, Rheumatology, Pneumology, Medical University, Innsbruck, Austria Search for more papers by this author Miguel P Soares Corresponding Author Miguel P Soares Instituto Gulbenkian de Ciência, Oeiras, Portugal Search for more papers by this author Günter Weiss Corresponding Author Günter Weiss Department of Internal Medicine VI, Infectious Diseases, Immunology, Rheumatology, Pneumology, Medical University, Innsbruck, Austria Search for more papers by this author Author Information Miguel P Soares 1 and Günter Weiss 2 1Instituto Gulbenkian de Ciência, Oeiras, Portugal 2Department of Internal Medicine VI, Infectious Diseases, Immunology, Rheumatology, Pneumology, Medical University, Innsbruck, Austria *Corresponding author. Tel: +351 446 45 20; E-mail: [email protected] *Corresponding author. Tel: +43 512 504 23255; E-mail: [email protected] EMBO Reports (2015)16:1482-1500https://doi.org/10.15252/embr.201540558 See the Glossary for abbreviations used in this article. PDFDownload PDF of article text and main figures. ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Microbes exert a major impact on human health and disease by either promoting or disrupting homeostasis, in the latter instance leading to the development of infectious diseases. Such disparate outcomes are driven by the ever-evolving genetic diversity of microbes and the countervailing host responses that minimize their pathogenic impact. Host defense strategies that limit microbial pathogenicity include resistance mechanisms that exert a negative impact on microbes, and disease tolerance mechanisms that sustain host homeostasis without interfering directly with microbes. While genetically distinct, these host defense strategies are functionally integrated, via mechanisms that remain incompletely defined. Here, we explore the general principles via which host adaptive responses regulating iron (Fe) metabolism impact on resistance and disease tolerance to infection. Glossary AIDS acquired immunodeficiency syndrome ALA δ-aminolevulinic acid ALAS1/2 δ-aminolevulinate synthase CO carbon monoxide CPOX coproporphyrinogen oxidase 2,5-DHBA 2,5-dihydroxy-benzoic acid DMT1 divalent metal transporter-1 Fe iron FECH ferrochelatase FTH ferritin H (heavy/heart) chain FTL ferritin L (light/liver) chain FPN ferroportin-1 Epo erythropoietin Erfe erythroferrone G6PD glucose 6 phosphate dehydrogenase GDF15 growth differentiation factor 15 HAMP hepcidin gene (hepcidin antimicrobial peptide) HFE hemochromatosis protein gene HIF1α hypoxia-inducible factor 1α HIV human immunodeficiency virus HMBS hydroxylmethylbilane synthase HO-1 heme oxygenase-1 HP haptoglobin HPX hemopexin HRG1 heme-responsive gene-1 IL interleukin IFNγ interferon γ IRP Fe regulatory protein IRE Fe-responsive element JNK c-Jun N-terminal kinase Keap1 Kelch-like ECH-associated protein 1 Lcn2 lipocalin LPS lipopolysaccharide MHC major histocompatibility complex Mø monocytes/macrophages NADPH nicotinamide adenine dinucleotide phosphate NALP3 NACHT, LRR, and PYD domains-containing protein 3 NETs neutrophil extracellular traps NF-IL6 nuclear factor for IL-6 expression NFκB nuclear factor κB NK cells natural killer cells NOX2 NADPH oxidase NO nitric oxide iNOS/NOS2 nitric oxide synthase NRAMP1 natural resistance-associated Mø protein 1 Nrf2 nuclear factor erythroid 2-related factor 2 ONNO− peroxynitrate PDGF-BB platelet-derived growth factor subunit B PBBS catalyst porphobilinogen synthase PGBS catalyst porphobilinogen synthase PMN cells polymorphonuclear cells PPIX protoporphyrin IX PPOX protoporphyrin oxidase PRR pattern recognition receptor RBC red blood cell ROS reactive oxygen species Rbx1 RING-box protein 1 RING really interesting new gene RIPK1 receptor-interacting serine/threonine kinases 1 RNS reactive nitrogen species STAT signal transducers and activators of transcription TCR T-cell receptor Tf transferrin TfR ferritin and transferrin receptor TH1 T helper type 1 cells TH2 T helper type 2 cells TLR Toll-like receptor TNF tumor necrosis factor UROD uroporphyrinogen III decarboxylase UROS uroporphyrinogen III co-synthase Introduction The pathogenic outcome of host–microbe interactions is countered by a number of evolutionarily conserved host defense strategies 1. These include avoidance, a behavioral-based strategy that limits exposure of the host to potentially pathogenic microbes 2. Physical barriers in the form of different epithelia provide an additional defense strategy that prevents microbes from gaining systemic host access 3. In the event of an infection, when microbes become systemic, immune-driven resistance mechanisms exert a negative impact on microbes that limits their pathogenicity. Resistance to infection relies, in most cases, on the expression of cytotoxic molecules that target pathogens. However, there are also resistance mechanisms that rely on the expression of molecules that while not cytotoxic per se prevent pathogens from accessing metabolites and/or nutrients that are essential for their survival and/or proliferation. This defense strategy, termed nutritional immunity 4, encompasses mechanisms that restrict microbes from accessing iron (Fe) 567 as well as other trace metals such as zinc, manganese, or copper 89. That nutritional immunity is a central defense strategy against infection is supported by the realization that several "infection resistance genes" act via a mechanism controlling host Fe metabolism. While essential to confer host protection against infection, some resistance mechanisms, including those involved in nutritional immunity, can compromise host homeostasis 10. This trade-off is countered by an additional host defense strategy termed as tissue damage control 111, which confers disease tolerance to infection 11213. The term disease tolerance, as used herein, refers to the same phenomenon identified originally in host–microbe interactions in plants 14. It limits the impact of infection on host integrity and fitness without interfering with the host's pathogen burden 14. This is distinct from immunological tolerance 15, and while the two may be functionally linked, they act via distinct mechanisms. Here, we explore how adaptive responses regulating host Fe metabolism impact on resistance and disease tolerance to infection. We will put forward the notion that pathogen class-specific mechanisms regulating Fe metabolism evolved to confer protection against intracellular versus extracellular pathogens. Other recent reviews focusing on related aspects of Fe metabolism, in the context of infection, are cited throughout the manuscript 67161718. Regulation of Fe metabolism during infection Host–microbe interactions evolved in a manner that is often linked to the emergence of host adaptive responses that regulate Fe metabolism and impact on Fe availability for microbes 5. These adaptive responses carry a trade-off such that modulation of Fe metabolism can only occur within narrow limits, compatible with host homeostasis 10. The general principle being that adaptive responses supplying Fe to microbes increase, in most cases, their pathogenicity while those withholding Fe from microbes limit their virulence. In support of this notion, host Fe overload is associated with poor clinical outcomes in a number of infectious diseases such as AIDS (i.e., co-infections resulting from HIV-driven immunodeficiency), malaria, and tuberculosis 19202122, while dietary Fe supplementation exacerbates the overall rate of mortality in areas endemic for such infectious diseases 2324252627. Of note, systemic Fe chelation yields inconsistent clinical outcomes, as illustrated for malaria or invasive fungal infections 2028, reflecting the complex interplay between host Fe metabolism and the pathogenesis of infectious diseases. Microbes evolved strategies to acquire Fe from their hosts 56717. These share as a common denominator the expression of siderophores that elute Fe from host Fe-binding proteins 2930 as well as other Fe uptake systems 631. Of note, deletion of genes regulating the expression of siderophores or other Fe uptake systems is often associated with reduced pathogen virulence, as illustrated for bacteria, for example, Staphylococci and Salmonella, or fungi, for example, Aspergillus, infections 673233. Along this line, polymorphisms in host genes encoding Fe-binding proteins that counter microbial siderophores or other microbial Fe uptake systems enhance resistance to bacterial infections 34. The majority of bioavailable Fe in mammals exists in the form of heme (Fig 1). It is therefore reasonable to assume that microbes evolved strategies to extract Fe from this tetrapyrrole ring structure 730. Examples are provided by Gram-positive bacteria such as Staphylococcus aureus, which expresses a specific hemoglobin receptor (IsdB), promoting heme extraction from hemoglobin 35. This is also the case for Gram-negative bacteria, such as Escherichia (E.) coli that express evolutionary conserved heme receptors, which bind and promote heme degradation 36. The same may hold true for protozoan parasites such as Plasmodium, which can incorporate host heme into their own metabolic pathways 37. Figure 1. Interrelationship of Fe and Heme metabolismMore than 80% of the bioavailable Fe in mammals exists in the form of heme contained in hemoproteins 241. The most abundant pool of Fe in mammals are the prosthetic heme groups of hemoglobin in red blood cells (RBC), followed by the heme groups of myoglobin in muscle cells and those of cytochromes and other ubiquitously expressed hemoproteins in all cells 3854. The Fe required to sustain hemoglobin synthesis is made available by hemophagocytic Mø as these clear senescent RBC by erythrophagocytosis (top right) 49. The heme contained in hemoglobin is transported by heme-responsive gene-1 (HRG1) into the cytoplasm where Fe is extracted by heme-oxygnease-1 (HO-1), exported via ferroportin (FPN), and delivered to traferrin (Tf) in plasma. Surplus cytoplasmic Fe in Mø is incorporated and stored within ferritin. Tf transports and provides Fe to the erythropoietic compartment via TfR, where it is used for heme synthesis (bottom right). Heme synthesis occurs via eight successive enzymatic reactions that take place back and forward in the mitochondria (brown) and the cytosol (blue). For details, see Box 1. Download figure Download PowerPoint Box 1. Heme synthesis Heme synthesis occurs via eight successive enzymatic reactions that take place back and forward in the mitochondria and the cytosol. The first consists in the condensation of glycine and succinyl-CoA into δ-aminolevulinic acid (ALA), catalyzed by the mitochondrial δ-aminolevulinate synthase (ALAS1/2). ALA translocates to the cytosol where it reacts with pyridoxal phosphate to form a porphobilinogen, via a reaction catalyzed by porphobilinogen synthase (PBGS). The resulting four porphobilinogen molecules are condensed by hydroxylmethylbilane synthase (HMBS) to generate hydroxymethylbilane, which is converted to uroporphyrinogen I and then to coproporphyrinogen I by a non-enzymatic and an enzymatic reaction catalyzed by uroporphyrinogen III co-synthase or isomerase (UROS). Decarboxylation of uroporphyrinogen III forms coproporphyrinogen III, via a reaction catalyzed by uroporphyrinogen III decarboxylase (UROD). Coproporphyrinogen III is then transported to the mitochondria where coproporphyrinogen oxidase (CPOX) catalyzes its oxidative decarboxylation into protoporphyrinogen IX, subsequently oxidized to protoporphyrin IX (PPIX) by protoporphyrin oxidase (PPOX). The final step consists in the insertion of the Fe, originating from hemophagocytic Mø and provided via transferrin, into newly synthesized protoporphyrinogen IX, via an enzymatic reaction catalyzed by ferrochelatase (FECH) 242. See also Fig 1. Host heme metabolism is controlled by several genes 38, including some that might restrict Fe access to microbes. This is probably the case for haptoglobin (HP) and hemopexin (HPX), two acute-phase proteins that scavenge extracellular hemoglobin and labile heme in plasma, respectively 39. Moreover, heme catabolism by heme oxygenase-1 (HO-1), a stress-responsive enzyme that converts labile heme into equimolar amounts of Fe, carbon monoxide (CO), and biliverdin 40, should also contribute to deprive microbes from accessing Fe contained in heme. That these genes modulate the pathogenesis of infectious diseases in humans is supported by the association of polymorphisms in the human HP genes with an unfavorable course of co-infections resulting from HIV-driven immunodeficiency 41, malaria 42, or tuberculosis 43. This also holds true for polymorphisms in the human HMOX1 gene coding for HO-1, as illustrated for HIV infection 44 and malaria 4546. More recently, an association between plasma levels of HO-1 and active versus latent or treated pulmonary tuberculosis has been established 47. This suggests that host adaptive responses regulating heme metabolism and depriving microbes from accessing this source of Fe can exert a major impact on the outcome of infectious diseases. Regulation of Fe metabolism by macrophages during infection Monocytes/macrophages (Mø) play a central role in the maintenance of Fe homeostasis, delivering approximately 95% of the Fe required to support de novo heme/hemoglobin synthesis during erythropoiesis 4849 (Fig 1). Hemophagocytic Mø engulf senescent and damaged erythrocytes, digest their hemoglobin content in phagolysosomes, and extract the prosthetic heme groups from hemoglobin (Fig 1). Labile heme is transferred to the cytoplasm via a mechanism assisted by the heme transporter heme-responsive gene-1 (HRG1) 5051 (Fig 1, top right). Once heme reaches the cytoplasm, it can be secreted from Mø via different cellular heme exporters 49 or degraded by HO-1, which extracts Fe from the tetrapyrrole ring of heme. Labile Fe is then excreted by the transmembrane protein ferroportin-1 (FPN) 5253, the only known cellular Fe exporter (Fig 1, top right). Alternatively, labile Fe is stored in Mø by ferritin, a multimeric protein composed of 24 heavy/heart (FTH) and light/liver (FTL) subunits 5455. Ferritin can store and, through the ferroxidase activity of FTH, convert about 4,500 atoms of Fe2+ into inert Fe3+ 5455 (Fig 1, top right). Heme catabolism by HO-1 induces the expression of ferritin, via a mechanism involving the production of labile Fe, which inhibits the binding of Fe regulatory proteins (IRP) to the 5′UTR of FTL and FTH mRNA, hence promoting their translation and ferritin expression 56. Fe export from Mø is controlled systemically by hepcidin, a 2.8-KDa acute-phase peptide secreted by hepatocytes, which binds FPN and triggers its proteolytic degradation 17. This results in sustained inhibition of Fe cellular export, leading to intracellular Fe accumulation and hypoferremia 5758. Hepcidin expression is regulated mainly at the level of transcription of the hepcidin (HAMP) gene, being induced in response to: (i) hyperferremia, that is, higher than normal levels of circulating Fe, (ii) cytokines, for example, interleukin (IL)-1, IL-6, and IL-22, or (iii) recognition of bacterial lipopolysaccharide by Toll-like receptor 4 (TLR4) 596061 and repressed in response to: (i) hypoferremia, that is, lower than normal levels of circulating Fe, (ii) anemia, (iii) tissue hypoxia, and (iv) hormones, for example, growth and differentiation factors 15, erythroferrone, platelet-derived growth factor-BB (PDGF-BB) and testosterone 62636465. Regulation of Mø intracellular Fe content by the hepcidin/FPN axis impacts on Mø polarization and presumably therefore resistance to infection 66667. Specifically, the intracellular Fe content of Mø modulates their response to interferon γ (IFNγ) 6869, a cytokine produced by activated natural killer (NK) cells and T helper type 1 (TH1) cells, which plays a central role in driving Mø microbicidal activity 70. Moreover, accumulation of intracellular Fe in T cells mitigates the activation, differentiation, and proliferation of TH1 cells while fostering TH type 2 (TH2) responses, as illustrated in the context of Candida albicans infection in mice 6971. Reduced cellular Fe content, resulting from blockage of Fe cellular import by the transferrin receptor in T cell, also inhibits TH1 cell responses, whereas TH2 cells are less sensitive to this effect, presumably due to their higher intracellular Fe content 72. While perhaps unfavorable in the context of infections by intracellular pathogens, which are cleared by TH1 responses 6973, Fe-driven TH2 responses might promote host resistance against large extracellular parasites, for example, helminthes, but this remains to be tested experimentally. The microbicidal activity of activated Mø depends critically on the phagocytic nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX2/gp91phox) and the inducible form of nitric oxide (NO) synthase (iNOS/NOS2) 7475. Both NOX2/gp91phox and iNOS/NOS2 are prototypical hemoproteins that use Fe-heme to catalyze the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS), respectively 76. While NOX2/gp91phox 77 expression and iNOS/NOS2 78 expression are regulated essentially at the transcriptional level, via a mechanism involving the activation of the nuclear factor kappa B (NF-κB) family of transcription factors, their activity requires the insertion of a heme prosthetic group into these apoproteins. This argues that expression of these enzymes must be functionally integrated with mechanisms regulating Fe-heme metabolism as to support their activity and hence Mø microbicidal activity. The underlying mechanisms remain elusive. In addition to their intrinsic cytotoxicity, ROS and RNS regulate Fe metabolism in a manner that also contributes to the overall microbicidal activity of activated Mø. Both ROS and RNS have high affinity toward Fe2+ in the prosthetic heme groups or Fe-sulfur clusters of a variety of proteins 7980. These include IRPs, to which NO can bind and modulate their stability as well as activity 8182, resulting in IRE-mediated regulation of Fe metabolism 5866818283848586. ROS also regulate IRP activity indirectly, via kinase/phosphatase signaling pathways 5866. The physiologic relevance of this mechanism was recently underpinned by the description of hepcidin-independent hypoferremia in mouse models of inflammation 87, and the fact that tumor necrosis factor (TNF) causes sustained hypoferremia in mice, irrespective of hepcidin 88. Whether ROS or RNS contribute to hepcidin-independent hypoferremia has not been established. ROS and RNS are also reactive toward thiol groups in the cysteines of redox-regulated proteins, such as the Kelch-like ECH-associated protein 1 (Keap1) 899091. This adaptor for the Cullin (Cul)3–RING (really interesting new gene)-box protein (Rbx)1 ubiquitin ligase complex targets constitutively the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) for proteolytic degradation by the 26S proteasome 91. When Keap1 Cys151 is targeted by RNS, the tertiary structure of Keap1 is altered, impairing its ubiquitin ligase activity 899091. This allows newly transcribed Nrf2 to undergo nuclear translocation and drive the transcription of target genes containing DNA-antioxidant-responsive elements in their promoter 91. These genes include HO-1, which generates CO via heme catabolism. This gasotransmitter compels bacteria to produce and release ATP, which is then sensed by the P2 × 7 purinergic receptor expressed by Mø 92. This modulates a K+ efflux pump which activates the NACHT, LRR, and PYD domains-containing protein 3 (NALP3), leading to caspase 1 activation, pro-IL-1β cleavage, and IL-1β secretion by Mø 92. This process of "microbe metabolic sensing" appears to be required to promote bacterial clearance by activated Mø 92. Some intracellular pathogens, such as Mycobacteria, can subvert this sensing system, going into a state of dormancy in response to CO 93. Taken together, these observations argue that regulation of Mø intracellular Fe content impacts on resistance to infection. This effect goes beyond secluding Fe from invading pathogens, encompassing the modulation of innate and adaptive immune responses against those pathogens. Pathogen class-specific regulation of Fe metabolism Pathogen class-specific mechanisms evolved most probably to limit Fe availability to extracellular versus intracellular pathogens, while enhancing resistance mechanisms 179495. Induction of hypoferremia, via systemic inhibition of extracellular Fe export coupled to increased Mø Fe/heme uptake and intracellular retention 56, reduces the growth and virulence of extracellular pathogens, as illustrated for Vibrio vulnificus infection in mice 9697. However, this strategy can be detrimental against intracellular pathogens, enhancing their growth and proliferation, as illustrated for bacteria from Chlamydia, Legionella, Salmonella, and Mycobacteria spp. 9899100101102. The impact of such pathogen class-specific mechanisms is particularly relevant in the context of co-infections, where restricting Fe access to one pathogen can be detrimental against co-infection by another pathogen. For example, heme catabolism by HO-1 103104105106 and subsequent intracellular Fe storage by ferritin 54107 confers host protection against the blood stage of Plasmodium infection while increasing susceptibility to the intracellular bacteria Salmonella 108. Whether this principle can be extrapolated to co-infections by other pathogens is likely, but remains to be established. Regulation of Fe metabolism as a host resistance mechanism against extracellular pathogens The general strategy to restrict Fe access to extracellular pathogens consists in the retention of intracellular Fe by Mø leading to hypoferremia (Fig 2). This is achieved essentially by the induction of Fe import and retention mechanisms, coupled to suppression of cellular Fe export (Fig 2). Activated Mø can produce minute amounts of hepcidin in response to bacterial lipopolysaccharide (LPS) or cytokines, targeting FPN for proteolytic degradation 109110. However, while this may provide a fast acting strategy to limit Mø Fe export, sustained Fe retention requires additional inputs provided by cytokines such as IFNγ, which represses FPN transcription 6 (Fig 2). IFNγ also induces the expression of iNOS/NOS2 78, via a mechanism regulated at a transcriptional level by the activation of the signal transducers and activators of transcription (STAT) family of transcription factors (Fig 2). As described above, the NO produced by iNOS/NOS2 regulates the binding affinities of IRP toward target IRE in genes such as transferrin receptor (TfR) and ferritin, inhibiting cellular Fe import and intracellular retention, respectively 588182111. However, once intracellular Fe levels increase in Mø, iNOS/NOS2 transcription is repressed creating a negative feedback loop, which promotes intracellular Fe retention 85111. IFNγ also induces the expression of the divalent metal transporter-1 (DMT1) in Mø, promoting ferrous Fe uptake and intracellular incorporation by ferritin 112 (Fig 2). Cytokine-driven NF-κB activation induces FTH transcription, supporting further retention of Fe in Mø 5455 (Fig 2). Figure 2. Regulation of Fe metabolism in response to extracellular pathogens(A) Immune responses to extracellular pathogens encompass the production of cytokines, for example, IL-1, IL-6, and IL-22, which induce the transcription of the hepcidin (HAMP) gene in hepatocytes. Cellular Fe retention by Mø can lead to the development of anemia of inflammation, which triggers the production of several hormones, such as Epo, Erfe, GDF15, PDGF-BB, or the activation of the HIF family of transcription factors, which reduce hepcidin expression. (B) Circulating hepcidin targets systemically the Fe export protein FPN for degradation, which inhibits Fe cellular retention in many cell types, including Mø, which are pivotal to the maintenance of Fe homeostasis. This is supported by the autocrine production of hepcidin by Mø. In addition, cytokines, such as IFNγ and TNF, also inhibit FPN transcription in Mø (not shown) promoting Fe retention and limiting further Fe availability to extracellular microbes. (C) Upon infection by extracellular pathogens, Mø can uptake extracellular Fe via different mechanisms involving Tf/TfR interaction, (D) the lactoferrin/lactoferrin receptor, (E) the Lcn2/Lcn2R, or (F) the divalent metal transporter DMT1. All of these contribute to scavenge extracellular Fe and prevent extracellular pathogens from accessing Fe. (G) In addition, Mø can engulf damaged RBC and prevent hemoglobin release or (H) scavenge extracellular hemoglobin/haptoglobin via the scavenger receptor CD163 as well as (I) heme/HPX complexes via the scavenger receptor CD91. (J) Uptake of heme/HPX, damaged RBC, or hemoglobin/haptoglobin by Mø is coupled to heme transport from phagolysomes to the cytoplasm, by the heme transporter HRG1, and to heme catabolism by HO-1. These pathways are induced by several cytokines in the course of an infection. Download figure Download PowerPoint Other cytokines such as IL-4, IL-10, and IL-13 promote TfR-mediated Fe uptake and intracellular retention by ferritin 113. Moreover, IL-10 and IL-6 also increase heme uptake from circulating hemoglobin–haptoglobin or heme–HPX complexes, via a mechanism involving the induction of the hemoglobin scavenger receptor CD163 and the HPX scavenger receptor CD91 114115, respectively (Fig 2). This is coupled to the induction of HO-1 in Mø 116, which extracts Fe from heme and promotes intracellular Fe storage by ferritin (Fig 2). Another resistance mechanism against extracellular pathogens, based on regulation of Fe metabolism, involves the expression of lactoferrin, a member of the transferrin family of Fe-binding proteins (Fig 2). Lactoferrin is expressed in many tissues and secretions where its primary function is to bind and restrict Fe delivery to extracellular microbes, a mechanism which accounts for its antimicrobial activity in breast milk 117. Activated Mø express and release lactoferrin, which scavenges and restricts Fe availability to extracellular pathogens 6117118 (Fig 2). Lactoferrin also protects Mø from oxidative stress 117119 and modulates Mø microbicidal activity, as shown in the context of Staphylococcus aureus infection 120121. The Fe-binding capacity of lactoferrin can also interfere with microbial virulence pathways such as biofilm formation 122. Moreover, lactoferrin modulates the activation and proliferation of lymphocytes, NK cells, and Mø, via a mechanism mediated in part by lactoferrin receptors expressed by these cells 117123. Cellular internalization of Fe-loaded lactoferrin by Mø can lead to cellular Fe overload and cytotoxicity 124. Of note, some bacteria evolved strategies to subvert the protective effects of lactoferrin extracting Fe from lactoferrin 125126. Although the exact mechanism by which bacteria can extract iron from lactoferrin remains to be determined, some Gram-negative bacteria can import lactoferrin following its binding to microbial transmembrane lactoferrin binding proteins A and B 127128. Lipocalin (Lcn2/NGAL) is a peptide produced by activated polymorphonuclear (PMN) cells and Mø, which captures Fe-laden bacterial catechole type siderophores, such as enterobactin, derived from Gram-negative bacteria (Fig 2). Lcn2 inhibits the acquisition of siderophore-bound Fe by bacteria, as illustrated for E. coli or Klebsiella spp. 129130. Lcn2 delivers bacterial as well as mammalian siderophore-bound Fe 131132 to the Lcn2 receptor expressed by host cells, suggesting that it modulates Fe trafficking and hence Fe metabolism during infection (Fig 2). Lcn2 also promotes resistance to non-bacterial pathogens, as demonstrated for Plasmodium infection 133. It should be noted, however, that Lcn2 exerts contrasting effects depending on the pathogen and tissue localization, improving or mitigating resistance to infection. For example, Lcn2 aggravates the outcome of Streptococcus pneumoniae infection 134 through a immunomodulatory effect possibly acting via regulation of Fe metabolism or another not yet fully elucidated pathway 135136. Of note, E. coli can use the mammalian siderophore, 2,5-dihydroxy-benzoic acid (2,5-DHBA) as Fe source 137. Accordingly, Mø respond to bacterial challenge by downregulating 2,5-DHBA synthesis, leading to 2,5-DHBA depletion and enhanced resistance to E. coli infection. Both ROS and RNS produced by activated Mø or PMN cells can elicit different forms of stress and damage to bystander red blood cells (RBC), eventually leading to the release of their hemoglobin content. Given the sheer number of RBCs (~2–3 × 1013 in adult humans) and their extremely high hemoglobin (~3 × 108 molecules/RBC) and heme (~1.2 × 109 molecules/RBC) content, disruption of a relatively small numbers of RBCs is sufficient to release significant amounts of hemoglobin 49. Extracellular hemoglobin exerts antimicrobial effects, driven by the peroxidase activity of the Fe contained in
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