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Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

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Background: The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular 'reactive oxygen species' (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. Review: We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation).The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible.This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, since in some circumstances (especially the presence of poorly liganded iron) molecules that are nominally antioxidants can actually act as pro-oxidants. The reduction of redox stress thus requires suitable levels of both antioxidants and effective iron chelators. Some polyphenolic antioxidants may serve both roles.Understanding the exact speciation and liganding of iron in all its states is thus crucial to separating its various pro- and anti-inflammatory activities. Redox stress, innate immunity and pro- (and some anti-)inflammatory cytokines are linked in particular via signalling pathways involving NF-kappaB and p38, with the oxidative roles of iron here seemingly involved upstream of the IkappaB kinase (IKK) reaction. In a number of cases it is possible to identify mechanisms by which ROSs and poorly liganded iron act synergistically and autocatalytically, leading to 'runaway' reactions that are hard to control unless one tackles multiple sites of action simultaneously. Some molecules such as statins and erythropoietin, not traditionally associated with anti-inflammatory activity, do indeed have 'pleiotropic' anti-inflammatory effects that may be of benefit here. Conclusion: Overall we argue, by synthesising a widely dispersed literature, that the role of poorly liganded iron has been rather underappreciated in the past, and that in combination with peroxide and superoxide its activity underpins the behaviour of a great many physiological processes that degrade over time. Understanding these requires an integrative, systems-level approach that may lead to novel therapeutic targets.
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BMC Medical Genomics
Review
Iron behaving badly: inappropriate iron chelation as a major
contributor to the aetiology of vascular and other progressive
inflammatory and degenerative diseases
Douglas B Kell*
Address: School of Chemistry and Manchester Interdisciplinary Biocentre, The University of Manchester, 131 Princess St, Manchester, M1 7DN, UK
E-mail: Douglas B Kell* - dbk@manchester.ac.uk
*Corresponding author
Published: 08 January 2009 Received: 2 September 2008
BMC Medical Genomics 2009, 2:2 doi: 10.1186/1755- 8794-2-2 Accepted: 8 January 2009
This article is available from: http://www.biomedcentral.com/1755-8794/2/2
© 2009 Kell; licensee BioMed Central Ltd.
This is an Open Access articl e distributed under t he te rms of the Creative Commons Attribution License (
http://creativecommons.org/license s/by/2.0),
which permits unre stricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Background: The production of peroxide and superox ide is an inevitable consequence of
aerobic metabolism, and while these particular 'reactive oxygen species' (ROSs) can exhibit a
number of biological effects, they are not of themselves excessively reactive and thus they are not
especially damaging at physiological concentrations. However, their reactions with poorly liganded
iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl
radical, which is exceptionally damaging, and a major cause of chronic inflammation.
Review: We review the considerable and wide-ranging evidence for t he involvement of this
combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed
pathological processes and inflammatory disorders, especially those involving the progressive
degradation o f cellular and organismal performance. These diseases share a great many similarities
and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially
hydroxyl radical generation).
The studies reviewed include those focused on a series of cardiovascular, metabolic and
neurological diseases, where iron can be found at the si tes of plaques an d lesions, as well as studies
showing the significance of iron to aging and l ongevity. The effective chelation of iron by natural or
synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems
properties, we need to recognise that physiological observables have multiple molecular causes,
and studying them in isolati on leads to inconsistentpatternsofapparentcausalitywhenitisthe
simultaneous combination of multiple factors that is responsible.
This explains, for instance, the decidedly mixed effects of antioxidants that have been observed,
since in some circumstances (especially the presence of poorly liganded iron) molecules that are
nominally antio xidants can actually act as pro-oxid ants. The reduction of redox stress thus re quires
suitable levels of both antioxidants and effective iron chelators. Some polyphenolic antioxidants
may serve both roles.
Understanding the exact speciation and ligand ing of iron in all its states is thus crucial to separating
its various pro- and anti-inflammatory activities. Redox stress, innate immunity and pro- (and some
anti-)inflammatory cytokines are linked in particular via signalling pathways involvi ng NF-kappaB
and p38, with the oxidative roles of iron here seemingly involved upstream of the IkappaB kinase
(IKK) reaction. In a number of cases it is p ossible to identify mechanisms by which ROSs and poorly
Page 1 of 79
(page number n ot f or citation purposes)
BioMed Central
Open Access
liganded iron act synergi stical ly and autocatalytically, leadi ng to 'runaway' reactions that are hard to
control unless one tackles multiple sites of action simultaneously. Some molecules such as statins
and erythropoietin, not traditionally associated with anti-inflammatory activity, do indeed have
'pleiotropic' anti-inflammatory effects that may be of benefit here.
Conclusion: Overall we argue, by synthesising a widely dispersed literature, that the role of
poorly li ganded iron has been rather underappreciated in the past, and that in combination with
peroxide and superoxide its activity underpins the behaviour of a great many physiolo gical
processes that degrade over time. Understanding these requires an integrative, systems-level
approach that may lead to novel therapeutic targets.
Background a nd preamble
The 'balkanisation' of the literature is in part due to the
amount of it (some 25,000 journals with presently 2.5
million peer-reviewed papers per year, i.e. ~5 per
minute [1]), with a number http://www.nlm.nih.gov/
bsd/medline_cit_counts_yr_pub.html increas ing by
something approaching 2 per minute at PubMed/Med-
line alone. In addition, the disconnect between the
papers in the literature (usually as pdf files) and the
metadata describing them (author, journal, year, pages,
etc) is acute and badly needs filling [2]. Without solving
this problem, and without automation of the processes
of reading, interpreting and exploiting this literature and
its metadata in a digital format, we cannot make use of
the existing tools for text mi ning and natural language
processing (e.g. [3-5]), for joining disparate concepts [6],
for literature-based discovery (e.g. [7-11], and for studies
of bibliometrics [12, 13], literature dynamics [14],
knowledge domains [15], detecting republication [16]
and so on. Until we recognise these possibilities we are
unlikely to seek to realise them.
The present article (and see [17] for a preprint) serves to
show some of the benefits than can accrue from a more
overarching view of the otherwise highly disparate
literature in a particular domain (see also [18]), but
was done 'the h ard way', i.e. with a f ew bibl iographic
and bibliometric tools but without the kind of automa-
tion implied above. For the record, the main tools used
(see a review in [2]) were Web of Knowledge a nd Scopus
for literature and citation searching, supplemented by
Google Scholar. Some use was also made of ARROW-
SMITH [6, 19, 20] and GOPubMed [21], as well as
various workflows in t he Taverna environment [22-26],
includ ing the BioAID_DiseaseDiscovery workflow http://
www.myexperiment.org/workflows/72 writt en by Marco
Roos. Citations and attendant metadata were stored in
Endnote (latterly version X).
Introduction
Even under 'normal' conditions, as well as during
ischaemia when tissue oxygenation levels are low, the
redox poise of the mitochondrial respiratory chain is
such that the normally complete four-electron reduction
of dioxygen t o water is also accompanied by the
production, at considerable rates (ca 14% of O
2
reduced), of partially reduced forms of dioxygen such
as hydrogen peroxide and superoxide (e.g. [27-45]).
These 1- and 2-electron reductions of O
2
are necessarily
exacerbated when the redox poise of t he b-type
cytochromes is low, for instance when substrate supplies
are in ex cess o r w hen the terminal electron acceptor O
2
is
abnormally low due to hypoxia or ischaemia. Various
other oxygenases, oxidases and peroxidases can also lead
direct ly to the production of such 'reduc ed' forms of
dioxygen in vivo (e.g.[46-48]),withH
2
O
2
from xanthine
oxidase being especially implicated in ischaemia/reper-
fusion injury (e.g. [47, 49-54]). These molecules (per-
oxide and superoxide) can cause or contribute to various
kinds of oxidative stress. However, this is mainly not in
fact because they can react d irectly with tissue compo-
nents th em selves, since they a re
compar atively non-
toxic, cells have well-known means of dealing with them
[55], and they are even used in cellular signalling (e.g.
[56-60]). Much more importantly, it is because they can
react with other particular species to create far more
reactive and damaging products such as hydroxyl
radicals, with all these agents nevertheless being known
collectively (and indiscriminately) as reactive oxygen
species ( ROSs). Possibly the commonest means by
which such much more damaging species, in particular
the hydroxyl radical, are created is by reaction with
unliganded or incompletely liganded iron ions [61-63].
The themes of this r eview are thus (i) that it i s this
combination of poorly ligan ded iron spec ies, coupled to
the natural production of ROSs, that is especially
damaging, (ii) that the role of iron has received far less
attention than has the general concept of ROSs, albeit
the large literature that we review, and (iii) that this
basic combination underpins a great many (and often
similar) physiological chang es leading to a variety of
disease manifestations, and in particular those where
the development of the disease is manifestly progres-
sive and degenerative.
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An overview of the structure of the review is given in Fig 1, in
the form of a 'mind map' [64]. The main literature review for
this meta-analysis was completed on June 30
th
, 2008, with
some updates being added following the refereeing process.
Some relevant chemistry of iron and
reduced forms of oxygen
While superoxide and peroxide are the proximate forms
of incomplete O
2
reduction in biology, a reaction
catalysed by the enzyme superoxide dismutase [65]
serves to equilibrate superoxide and peroxide:
2O
2
-
+2H
+
Æ H
2
O
2
+O
2
(1)
Arguably the most important reaction of hydrogen
peroxide with (free or poorly liganded) Fe(II) is the
Fenton reaction [66], leading to the very reactive and
damaging hydroxyl radical (OH
)
Fe(II) + H
2
O
2
Æ Fe(III) + OH
-
+OH
(2)
Superoxide can also react with ferric iron in the Haber-
Weiss reaction [67] to produce Fe(II) again, thereby
effecting redox cycling:
O
2
-
+ Fe(III) Æ O
2
+ Fe(II) (3)
Figure 1
An overview of this article, set out in the f orm of a 'mind map' [64].
BMC Medical Genomics 2009, 2:2 http://www.biomedcentral.com/1755-8794/2/2
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Ascorbate can replace O
2
-
within the cell for reducing
the Fe(III) to Fe(II) [68]. F urther reactions, that are not
the real focus here, follow from the ability of hydroxyl
radicals and indeed Fe(n) directly to interact with many
biological macro- and small molecules, especially
including DNA, proteins and unsaturated lipids. Thus
[69-73], Fe(II) and certain Fe(II) chelates react w ith lipid
hydroperoxides (ROOH), as they do with hydrogen
peroxide, splitting the OO bond. This gives RO
,an
alkoxyl radical, which can also abstract H
from
polyunsaturated fatty acids and from hydroperoxides.
The resulting peroxyl radicals ROO
can continue
propagation of lipid peroxidation. Oxidative stress also
leads to considerable DNA damage [74-76] and to the
polymerisation and denaturation of proteins [77-79]
and proteolipids that can together form insoluble
structures typically known as lipofucsin (see e.g. [80,
81]) or indeed plaques. Such plaques can also entrap the
catalysts of their formation, and thereby point them up.
Some of the evidence for these is described below. Many
small molecule metabolic markers for this kind of
oxidative stress induced by the hydroxyl r adical and
other 'reactive oxygen species' (ROSs) are known [ 43,
82-89], and include 8-oxo-guanine [90-94], 8-hydroxy
guanine [95], 8-hydroxy-2'-deox y-guanosine [96, 97],
8-oxo-GTP [98 ], 4-hydroxy-2-hexenal [99], 4-hydroxy-
nonenal [100], 4-hydroperoxy-2-nonenal, various iso-
prostanes [101-107], 7-keto- cholesterol [108], many
other cholesterol derivatives [109], malondialdehyde
[110], neopterin [111], nitrotyrosine [112-115] and
thymidine glycol [116, 117]. Note that the trivial
names in common use for this kind of metabolite are
not helpful and may even be ambiguous or misleading,
and it is desirable (e.g. [118]) to refer to such molecules
using terminology that relates th em either to molecules
identified in persistent curated datbases [119] such as
ChEBI [120] or KEGG [121], or better to describe t hem
via database-independent encodings such as SMILES
[122] or InChI [123-128] strings. (There are other
oxidative markers that may be less direct, such as the
ratio of 6-keto-prostaglandin F1a to thromboxane B2
[129], but these are not our focus here.)
Overall, it i s in fact well e stablis hed that the interactions
between 'iron' sensu lato and partly reduced forms of
oxygen can lead to the production of the very damaging
hydroxyl radical (e.g. [43, 1 30-139]), and that this
radical in particular probably underpins or mediates
many higher-level manifestations of tissue damage,
disease, organ failure and ultimately death [36, 137,
140-143]. While the role of ROSs generally in these
processes has been widely discussed, the general
recognition of the impor tance of inadequately liganded
iron in each of them has perhaps been less than fully
appreciated. One of our tas ks here will therefore be to
stress this role of 'iron', and to assess the various means
of chelating 'iron' such that it does not in fact do this.
(Throughout we use 'iron' to refer to forms of Fe(n,
n > 0) with unspecified ligan ds, though we absolutely
stress that it is the
exact speciation and liganding that
determines the reactivity of 'iron' in catalysing r eactions
such as that of hydroxyl radical formation, and indeed its
bioavailability generally inadequate liganding of iron
in the r equired forms can be a cause of anaemia even if
the total amount of 'iron' is plentiful.)
For completeness we note the reactions catalysed by
superoxide dismutase
2O
2
-
+2H
+
Æ O
2
+H
2
O
2
(4)
and by catalase
H
2
O
2
Æ H
2
O+1/2O
2
(5)
These to gether, were th eir activi ty in the relevant
locations sufficiently gr eat, might serve to remove (su)
peroxide from cells completely.
In addition to reactive oxygen species there are ions such
as the perferryl ion (Fe-O) [144] and reactive nitrogen
species [ 60, 145-147]. These latter are mainly formed
from the natural radical NO, an important inflammatory
mediator [148], with peroxynitrite production (from the
reaction of NO and superoxide) [46, 149-154] leading to
nitrotyrosine [112], or nitro-fatty acid [155, 156] or
protein cystein nitrosylation [157, 158 ] being a common
means of t heir detection downstream. Other toxic
products of the reactions of NO include NO
2
,N
2
O
3
,
and S-nitrosothiols [159], and the sequelae of some of
these may also involve iron [160].
Overall, we recognise that these kinds of inflammatory,
oxidative stress-related reactions are accumulative and
somewhat irre versible [161], that they are consequently
age-related, and (see [162-165] and later), and that most
diseases and causes of mortality that are prevalent in the
developed world are in this sense largely manifestations
of this kind of aging.
Ligands and siderophores
As well as the reactions described above, ferrous ions will
react with oxygen under aerobic conditions to produce
ferric ions, and in natural environments there is little to
stop this. Consequen tly, and because these reflect
fundamental physicochemical properties of such ions,
the problems of both solubility and toxicity were faced
by bacteria (and indeed fungi [166-169]) long ago in
evolution, and were solved by their creation and
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excretion of (mainly ferric-)iron chelators known as
siderophores [170-189] (and for haemophores see
[190]). These typically have extremely tight binding
constants (K
f
>10
30
[191]) and can solubilise and
sequester iron such that it can be internalised via suitable
transporter molecules within the bacterial plasma
membrane [192]. Bacterial and fungal siderophores
usually form hexadentate o ctahedral complexes with
ferric iron and typical ly employ hydroxamates, a-
hydroxycarboxylates and catechols as extremely effective
Fe
3+
ligands [182]. Since bacterial growth requires iron,
it is unsurprising that siderophores are effectively
virulence factors ( e.g. [174, 193-196]). While upwards
of 500 microbial siderophores have been identified
[182], with new