|Hemoglobin and methemoglobin catabolism; Brain
capillary; Ferric iron; Blood brain barrier; Apoptosis; Neurovascular
unit; Neurodegenerative brain disease
|It is still not known why redox-active iron levels are abnormally
high in some regions of the brain in neurodegenerative disorders, just
as it is not clear whether iron accumulation in the brain is an initial
event that causes neuronal death or a consequence of the disease
process. We looked at the sources of redox-active iron as a key factor
in understanding the role of oxidants and oxidative stress and ironinduced
oxidative stress as a constitutent and common mechanism
involved in the development of the neurodegenerative process in
Alzheimer’s disease. In the light of previous research we consider that,
at least in some neurodegenerative disorders, brain iron misregulation
is an initial cause of neuronal death and that this misregulation might
be linked to either genetic or non-genetic factors.
|Ferric-iron brain accumulation as a cause of neurodegenerative
brain disease: A new insight to understanding the mechanism
of iron transport
|The current view has been derived from research results, with
particular attention paid to the role of environmental toxicity factors
in brain endothelial small vessels. Our research showed that people
continuously inhaling strong oxidants such as NOx (NO and NO2)
reversibly oxidize oxyhemoglobin (Fe2+) to methemoglobin (Fe3+).Irreversible methemoglobinemia can arise because of the disruption of
the oxidant/antioxidant balance, supported by synergic SO2 metabolites
leading to the degradation of antioxidant thiols . Methemoglobin by
itself, and heme, have prooxidant properties and induce structural and
functional changes in the vascular endothelium [2,3]. These changes
include cell growth arrest, senescence, morphological alterations and
cell apoptosis, and they lead to both vessel thrombosis, and endothelial
cell denudation under the influence of redox-active ferric iron (Fe3+), as
a product of heme-oxygenase, which is responsible for methemoglobinheme
degradation . In the blood, nitric oxide and superoxide
form peroxinitrites (ONOO) that convert oxyhemoglobin into
methemoglobin, and the methemoglobin-released from heme induces
endothelial cytolysis . The toxicity of H2O2 is also dependent upon its reaction with ferrous iron to form hydroxyl radicals by the Fenton
reaction. The ferrous iron needed for this reaction is formed by the
reduction of cellular ferric iron by superoxide ions . We would like to
emphasize the difference between physiological hemoglobin catabolism,
and pathological methemoglobin catabolism, because their different
final products, ferrous and ferric iron, have distinct characteristics.
Ferrous iron has the potential for catalyzing and generating highly
cytotoxic hydroxyl radicals as from the Fenton reaction (Ferrous iron
+ H2O2 → Ferric iron+OH+OH). Ferric iron is then reduced back to
ferrous iron, a peroxide radical and proton by the same hydrogen
peroxide. The substantial difference between the intracellular ferric
iron originates from the Fenton reaction. Ferric iron originates from
methemoglobin catabolism perform a level of methemoglobin cellular
uptake which increases the methemoglobinemia leading to Ferric-ironinduced
oxidative stress injury. We consider that from methemoglobin
catabolism the last product of Ferric iron is a significant added source
of ferric iron derived from the Fenton reaction, whose continuious
formation has an impact upon the brain neurovascular unit. According
to our hypothesis this could be the cause of neuronal death in humans,
and of the ageing process, leading finally to hard neurodegenerative
disorders such as AD, PD and others. Our view confirms the statement
that iron and iron-induced oxidative stress constitute a common
mechanism that is involved in the development of neurodegeneration.
According to the suggestions of previous research we consider that, at
least in some neurodegenerative disorders, brain iron misregulation is
an initial cause of neuronal death and that this misregulation might
be the result of either genetic or non-genetic factors . Our previous
research results suggest that methemoglobin plays a particularly
important role as carrier and donor of the cytotoxic and redox-active
Ferric(Fe3+) iron, and determines how iron is transported intracellularly
(Figures 1-3). Neuroscience has traditionally focused on the neurons of
the central and peripheral nervous systems, and it is now becoming
clear that neurons, glia and microvessels are organized into a well
structured neurovascular unit, and recent studies have highlighted
the importance of brain endothelial cells in this modular organization
. Iron is essential for normal cell function, but it also generates
toxic reactive oxygen species (ROS) that adversely affects the vascular
endothelium of the blood-brain barrier. Human neuroblastoma IMR-
32 cells’ exposure to ferric ammonium citrate(FAC) as a model of
neuronal iron overload and neurodegeneration was investigated. In
the consequences of the exposure of cells to ferric ammonium citrate
it was found that exposure was associated with increased oxidant cell
levels, activation of redox-sensitive signals, and apoptosis . Leung
et al. have demonstrated that ferric heme is significantly more prooxidant
than is ferrous heme. In addition they have shown that this
ferric heme has a much higher relaxivity than its ferrous counterparts.
In tandem, this evidence suggests that the MR imaging–detected T1
(longitudinal relaxation time) high signal intensity within the vessel
wall is an endogenous biomarker of an intraplaque environment that
is highly pro-oxidant and proatherogenic. MR imaging measures
showed a T1 relaxivity that was 10 times higher for ferric than for
ferrous forms of hemoglobin. Their results support the hypothesis
that ferric methemoglobin–generated T1 high signal intensity reflects
a prooxidant environment that, in the setting of vessel wall disease,
might be proatherogenic . This justifies the study of the oxidant
effect of methemoglobin catabolic products on vital organs and the
CNS, resulting in their dysfunction. The research MRI data showed
that extracellular methemoglobin generates significantly more lipid
oxidation than intracellular products; however, methemoglobin in
both these environments has similar measures of r1 (longitudinal
relaxivity of relaxation rate). Therefore, the T1 high signal intensity
due to methemoglobin is not solely restricted to an environment that causes lipid oxidation. Thus, the ability of this high signal intensity to
reflect at-risk plaque may be diminished. However, it is known that in
the absence of any chemical modifications, ferric heme substantially
degrades the integrity of the RBC membrane, and the eventual fate
of a static RBC is lysis. Thus, intracellular methemoglobin is destined
to rapidly become extracellular, thereby adding to the oxidative drive
. The cellular and intercellular iron transport mechanisms in the
central nervous system (CNS) are still poorly understood, meanwhile
accumulating evidence suggests that impaired iron metabolism is an
initial cause of neurodegeneration . Brar et al. concluded that the
development of parkinsonism during the course of AD appears to be
associated with the accumulation of iron, which in turn may contribute
to the pathogenesis of neurologic decline .
|Our results point out the consequences of brain damage caused by
toxic environmental oxidants with a view to the role of methemoglobin
catabolism in pregnancy as the source of ferric (Fe3+) iron form
concentrated in various brain regions. Methemoglobin and hemolysis
both occur as a result of oxidative stress, but the prevalent difference between them is that methemoglobin is a reversible phenomenon
(oxidant–antioxidant balance) whereas hemolysis, which occurs as a
result of oxidative stress on the erythrocyte membrane, is an irreversible
event. Methemoglobinemia can additionally exacerbate an existing
anemia, stimulating hypoxia that may be additionally dangerous. Our
prospective study of methemoglobin in pregnancy revealed a significant
rise in the level of methemoglobin >1.5 g/L (r=0.72, p<0.01) in the airpolluted
exposure period, which can be explained on the basis of an
oxidant–antioxidant imbalance, resulting in methemoglobinemia .
Methemoglobinemia and stillbirth recorded throughout the exposure
period were significantly higher than those recorded in the control
period (p=0.0205) and the frequencies of reproductive loss were
significantly lower in the control than in the exposure period (p<0.05)
. As we have found no evidence of the consequences of mother
methemoglobinemia on the fetus, the second objective was to direct
attention to methemoglobin as an early biomarker of the oxidative
stress effects caused by environmental toxicity, which put pregnancy
at risk and may later impair the health of newborns, children and
adolescents. Our research found neonatal jaundice incidence (p=0.034),
heart murmur at a later age (p=0.011), as well as child and adult mild
disorders such as dyslalia and learning/memory impairments (p=0.002)
which were significantly higher than in children and adults of control
mothers without pregnancy methemoglobinemia . Lavezzi et
al. recently presented findings, confirmed with pathohistological
techniques, that under adverse conditions, ferric iron positivity in
capillary endothelial cells of the blood-brain barrier in the fetus rise,
also resulting in preterm birth, stillbirth or early neonatal death .
The application of the Blue Prussian method highlighted accumulations
of blue granulations, indicative of nonheme Fe3+ -positive reactions, in
the brainstem and cerebellum of 12 (33%) of 36 victims and in none of
the control group. In the positive cases, iron deposits were widespread
in brain parenchyma or localized in specific areas showing a variable
extent and intensity (Figure 4).
|According to our observations, we point out specific cellular
methemoglobin and heme catabolism when the last product leading to
Ferric iron which will yield cytotoxic and paramagnetic property has
an notable role ‘in situ’. We propose that ferric iron and ferric ironinduced
oxidative stress constitute a common mechanism involved
in the development of neurodegeneration, and also sugests an initial
cause of neuronal death as a result of environmental toxicity factors.
The experiments showed that ferric and ferrous iron can enter cells via
different pathways, they do not indicate which pathway is dominant in
humans . Heme, the major functional form of iron, is synthesized in
the mitochondria. Smith et al. suggest that iron is able to participate in
‘in situ’ oxidation and readily catalyzes an H2O2-dependent oxidation,
and indicates that iron accumulation could be an important contributor
toward the oxidative damage of Alzheimer’s disease . Our work,
according to our standpoint, supports the above statement about the
importance of disturbed heme metabolism when the heme oxygenase-1,
an enzyme that catalyzes the conversion of methemoglobin and heme
to ferric iron, is increased in Alzheimer’s disease suggesting increased
heme turnover as a source of redox-active iron. Perry et al. have found
that while mitochondrial DNA and cytochrome C oxidase activity
are increased in Alzheimer’s disease, the number of mitochondria
is decreased, indicating accelerated mitochondria turnover, and
suggesting mitochondrial dysfunction as a potentially inseparable
component of the initiation and progression of Alzheimer’s disease
. It was also found that oxidative damage occurs primarily
within the cytoplasm rather than in mitochondria. According to this hypothesis that mitochondria acts as a source of hydrogen peroxide,
an intermediate, once in the cytoplasm, will be converted into highly
reactive hydroxyl radicals through the Fenton reaction in the presence
of metal ions (iron and copper) causing damage to the cytoplasm .
Cell apoptosis is initiated by extracellular and intracellular signaling
pathways, the death receptor- and the mitochondria-mediated
pathway. Various pathologies can result from oxidative stress-induced
apoptotic signaling consequently leading to ROS increases and/or
antioxidant decreases, disruption of intracellular redox homeostasis,
and irreversible oxidative modifications of lipid, protein, or DNA .
Furthermore, iron participates in diverse pathologic processes by the
Fenton reaction, which catalyzes the formation of reactive oxygen
species (ROS). To test the hypothesis that this reaction accelerates
apoptosis, Jacob et al. used human umbilical vein endothelial cells
(HUVECs) as surrogates for the microvasculature in vivo. HUVECs
were loaded with Fe3+ (ferric chloride and ferric ammonium citrate),
and apoptosis executed after a heat shock stimulus . Brain iron is
a major contributor to magnetic resonance imaging (MRI) contrast
in normal gray matter. Non-heme brain iron is present mainly in
the form of ferritin. The quantitation of non-heme brain ferric iron
indicated by MRI helps in the diagnosis and monitoring of different
neurological diseases . Most of the brain non-heme iron is believed
to be present as a storage pool consisting of ferritin or hemosiderin
and also as a product of methemoglobin catabolism . However,
the concentration of transferin–bound iron is always far too small to
affect MRI. This fact suggests considering the role of methemoglobin
catabolism as the source of ferric iron (Fe3+) form concentrated in
various brain regions. Nowdays, non-heme-bound Fe3+ is quantified
using Magnetic Resonance Imaging (MRI), thanks to its paramagnetic
properties. It is believed that most non-heme-bound iron is deposited
in the form of ferritin, haemosiderin, or methemoglobin catabolic
products, whereas transferrin-bound iron concentration is always low
and can not be detected by MRI . Recent research results indicate
a ferrihydrite-magnetite core-shell ferritin structure. It was also found
that the magnetite in the ferritin iron core is not a source of free
toxic ferrous iron, as previously believed. Therefore, the presence of
magnetite in the ferritin cores of patients with Alzheimer’s disease is
not a cause of their increased brain ferrous iron(II) concentration .
|Methemoglobin and heme have prooxidant properties. Abundant
in the source of cytotoxic and redox-active ferric (Fe3+) iron which
without ferrous-ferric inversions, ‘in situ’ as a cause of iron –induced
oxidative stress, have an direct and specific impact on the brain endothelial small vessels, and increase the rate of endothelial cell
apoptosis and so make possible the accumulation of methemoglobin,
heme and ferric iron, in brain parenchyma. Our results identify
the consequences of mother-fetal methemoglobinemia caused by
environmental oxidants. Under the gradual influence of free radicals
on physiological erythrocytes and pathological methemoglobin
degradation, we found significant incidence of neonatal bilirubinemia,
heart murmur and learning/memory impairments in childhood and
teenagers, which has not been precisely demonstrated yet. In conclusion
we point out the importance of methemoglobinemia not only as the
biomarker and precursor of the effects of environmental oxidants, but a
carrier and donor of redox-active ferric iron. We identify ferric iron as
an originator having an important role in crossing brain microvessels
to neurons (neurovascular unit), causing neuronal death and continous
ageing process, and leading finally to hard neurodegenerative disorders
such as AD, PD, MS and other diseases. Nevertheless our findings as to
the relation between environmental oxidants and the pathogenesis of
neurodegenerative diseases need further research.
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