**12. Iron metabolism in healthy and Alzheimer's disease brain**

About 48% of the iron in the body is bound to hemoglobin and is involved in oxygen transport in the body. About 17% of the iron is found as the cofactor in proteins to carry out functions in several crucial biological processes such as the tricarboxylic acid cycle, oxidative phosphorylation, DNA synthesis and repair, and iron homeostasis. In the brain, iron is involved in myelination, neurotransmitter synthesis, and antioxidant enzyme function, and its entry and exit are tightly regulated by a variety of molecules. Aging, inflammation, and oxidative stress, which disturb the functions of molecules involved in iron metabolism, present as the main contributors to iron dyshomeostasis. Iron transport across blood-brain barrier in the brain, transferrin receptor 1 (TfR1), responsible for the strict control of the level of iron transported into the brain, is expressed on the luminal side of the brain microvascular endothelial cells (BMECs) and the blood-cerebrospinal fluid barrier. After circulation, a complex was formed (holo-Tf) by iron with transferrin (Tf), it binds to TfR1 on the surface of the BMECs, followed by entry into the BMECs via clathrin-mediated endocytosis. Fe3+ detaches from Tf in the acidic environment of the endosome and is reduced to Fe2+ by six-transmembrane epithelial antigen of prostrate 3(STEAP3) or duodenal cytochrome b (DCYTB), both of which are metalloreductases. It then enters the cytoplasm via divalent metal transporter 1 (DMT1). Fe2+ in BMECs can then enter the brain by the secretion of ferroportin 1 (FPN1), followed by the oxidation by extracellular ceruloplasmin (Cp) or hephaestin. Non-transferrin-bound iron (NTBI) can cross the blood-brain barrier (BBB) via

receptor-mediated transcytosis after binding to heavy-chain ferritin (H-ferritin;Lf). It was also reported that Lf increased in the brains of aged individuals and those with AD, allowing large amounts of non-Tf-bound iron to enter the brain. Iron transport and storage within the brain neuronal iron metabolism TfR1 is highly expressed on the surface of neurons, and similar to BMECs, iron enters neurons via clathrin-mediated endocytosis of holo-Tf/TfR1 and exits the endosomes in the form of reduced Fe2+ via DMT1. NTBI can also enter neurons in a DMT1-dependent manner independent of Tf. Cellular prion protein (PrPC) is abundantly expressed on the surface of neuronal membranes. It functions as a ferrireductase partner for DMT1, mediating Fe2+ uptake in the plasma membrane in the form of complex PrPc/DMT1. PrPC knockout in mice can lead to iron deficiency in brain and uptake increase of holo-Tf. By comparing the brain tissues of juvenile, adult, and aged rats that had the pathological features of AD, it was found that DMT1 abnormally increased with age. They supposed that DMT1 may be one of the main reasons why the iron concentration in the brain gradually increases with age. Some Fe2+ undergo normal metabolism in the cytoplasm of neurons, while some are stored in ferritin in the form of nontoxic Fe3+; when neurons are low in iron, ferritin can be degraded by lysosomes to release the stored iron to meet the physiological needs of the neurons. Ferritin is positively correlated with iron overload and is found deposited in senile plaques in the AD brain. It had been shown that there was an age-dependent increase in ferritin in the brain, probably a contributor to the iron overload in aged and AD brains. Autopsy studies of AD patients have revealed that mitochondrial ferritin is upregulated. Ferritin in the cerebrospinal fluid (CSF) of AD patients has been shown significantly increased, which is negatively correlated with cognitive decline and hippocampal atrophy in AD. Additionally, iron can enter mitochondria to form iron ferroptosis and Alzheimer's disease sulfur cluster and participate in the process of aerobic respiration. Regarding the transport of excess iron out of neurons, FPN1 is the only known iron exporter to date. Both Cp and hephaestin (Heph) can oxidize Fe2+ and facilitate FPN1to export iron, so the FPN1/Cp and FPN1/Heph are the main iron efflux pathways. Decrease of any of these three export proteins can induce iron retention and consequently the memory impairment. It was reported that FPN1 was downregulated in the brains of AD patients and triple-transgenic AD mouse models; thus, excessive iron could not be excreted normally, initiating intracellular iron deposition. Since Cp is a crucial partner of FPN1 to oxidize Fe2+ before it is excreted by FPN1, the dysfunction of Cp serves as an upstream event of iron retention, which has been found in AD. Noteworthily, both of amyloid precursor protein (APP) and tau, which are the substrates of the AD hallmarks in pathological condition, are crucial for neuronal iron efflux. APP is defined as a metalloprotein involved in iron homeostasis. With the assistance of soluble tau protein, APP is transported to the cell membrane where it stabilizes FPN1 and facilitates the efflux of iron. APP or tau knockdown can lead to abnormal FPN1 function and the inability of neuronal iron to flow out normally, resulting in neuronal iron overload. APP with the pathogenic Italian mutation A673V is more prone to be cleaved by β-secretase to produce Aβ1–42, impeding its support of FPN1 and thus increasing iron retention. Because of the continuous cleavage of APP and hyperphosphorylation of tau in AD brain, the iron efflux was hindered in neurons. Glial support for neuronal iron metabolism glial cells help to maintain the iron availability at a safe level in neurons. Astrocytes and microglia respond during iron overload or deficiency in order to maintain neuronal iron homeostasis. As a buffer pool, astrocytes express abundant TfR1 and DMT1, which facilitates taking up of both holo-Tf and NTBI from the abluminal side

#### *Iron Oxide Nanoparticles: A Mighty Pioneering Diagnostic Tool But Is It Really Safe… DOI: http://dx.doi.org/10.5772/intechopen.112074*

of BMECs and the brain interstitium, precisely regulating the iron concentration in neurons. Microglia also express TfR1 and reduce iron toxicity by promoting the influx of excess iron (for storage in ferritin) via the TfR1/DMT1 pathway. Microglia and astrocytes are capable of releasing ferritin carrying Fe3+ to supplement the iron deficiency or to support oligodendrocytes for myelination or remyelination. Iron is essential for myelination in oligodendrocytes, which are the most iron-rich cell type in the brain. TfR1 is absent in oligodendrocytes, while H-ferritin is the main source of iron for oligodendrocyte by interaction with T-cell immunoglobulin mucin domain 2 (TIM2). Noteworthily, when iron is overloaded, oligodendrocytes provide an antioxidant defense for neurons by secreting H-ferritin, scavenging extracellular extra iron [40–46].

### **13. Impact of iron overload on Alzheimer's disease pathology**

Currently, the involvement of iron in the early pathology of AD has been well accepted since the discovery of the link between dysregulation of brain iron homeostasis and AD pathogenesis in 1953. In the preclinical stage of AD, there is significant abnormal iron elevation in cortical, hippocampal, and cerebellar neurons while much severe in the cortex and hippocampus, the main brain areas affected by AD [47]. The iron overload in the brain is corresponding to the severity of AD lesions and the rate of cognitive decline. It is also proposed that hippocampal iron deposition could be the predictor of the rate of cognitive decline caused by Aβ. Iron overload drives a series of events, including glial activation, formation of Aβ plaque and tau tangles, and even neuronal loss, pushing the progress of the disease and accelerating cognitive decline. Iron interaction with Aβ plaques and neurofibrillary tangles iron accumulation was demonstrated to accelerate senile plaque deposition and the production of neurofibrillary tangles [48, 49]. Autopsy evidence and magnetic resonance imaging analysis provide evidence that there are a large amount of iron deposition not only in and around senile plaques but also in the sites of cortical tau accumulation, indicating the potential cross talk of iron with both of senile plaques and neurofibrillary tangles. Perturbations in iron homeostasis is one of key players in Aβ deposition. High intracellular iron concentration enhances the interaction of IRP/IRE, inducing APP upregulation. Furthermore, the enzymes that cleave APP named α- and β-secretase are tightly balanced and modulated by furin. More β-secretase is activated when α-secretase is suppressed by furin impairment in the condition of excessive iron. Upregulated APP is cleaved by more β-secretase to Aβ40/42, accelerating the Aβ deposition [50, 51]. Meanwhile, APP can no longer assist FPN1, resulting in impaired iron efflux and aggravated iron deposition. Some researchers have even proposed that Aβ is nontoxic in the absence of redox metals and that aggregation of Aβ requires the involvement of metals. Soluble Aβ binds to Fe3+ when extracellular iron increases so as to remove excess iron, but it is difficult to dissociate them after they interact; Aβ can promote the reduction of Fe3+ to Fe2+, and the reactive oxygen species (ROS) released during this process allow Aβ to be deposited more easily and rapidly, forming more senile plaques. The interactions of iron with APP and Aβ greatly increase the formation rate and degree of senile plaques. Therefore, some researchers believe that iron deposition should be included in the "Aβ cascade hypothesis" of AD. Iron can also interact with tau. Reduced soluble tau in the brain of AD patients increased brain iron deposition by suppressing FPN1 activity. On the contrary, a diet high in iron can lead to cognitive decline in mice, increased abnormal tau phosphorylation in neurons, and

abnormal expression of insulin pathway-related proteins. Insulin supplementation can reduce iron-induced phosphorylation of tau, indicating that iron deposition may lead to tau hyperphosphorylation by interfering insulin signaling. In vivo research has found that iron can be involved in tau hyperphosphorylation by activating the cyclindependent kinase 5 (CDK5)/P25 complex and glycogen synthase kinase3β (GSK-3β). Excessive intracellular Fe2 + −induced production of oxygen free radicals can also promote tau hyperphosphorylation by activating the extracellular. Ferroptosis and Alzheimer's disease signal-regulated kinase 1/2 (Erk1/2) or mitogen-activated protein kinase (MAPK) signaling pathways. Glial activation and neuroinflammation has been demonstrated to be a prominent characteristic of AD pathology. Microglial are highly reactive cells responding to increased iron levels in the brain. When iron level increases in brain, microglia become activated, with soma volume increased and process length decreased. Iron may activate microglia through proinflammatory cytokines mediated by the nuclear factor-κB (NF-κB). After activated, they express more ferritin to scavenge the extracellular iron, resulting in intracellular iron retention, increased TNFα expression, and finally infiltrated with Aβ-plaques. Activated microglia also secret Lf, which can interact with APP, promoting the Aβ formation. Conversely, formation of Aβ induces more IL-1β expression in microglia in the environment of elevated iron, exacerbating the proinflammatory effects. Astrocytes are highly resistant to metal-induced toxicity within the brain as the critical cell type in maintaining a balanced extracellular environment and supporting the normal functioning of neurons. In the environment of high iron, astrocytes respond with a significant increase in glutathione, catalase, and manganese superoxide dismutase levels to resist the oxidative stress. They show less impairment by iron than neurons and oligodendrocytes. But later, the astrocytes were found activated with increased glial fibrillary acidic protein (GFAP). Activated astrocytes release inflammatory mediators and induce oxidative stress, which facilitate the formation of Aβ and tau tangles and hinder Aβ clearance. Iron overload induces oxidative stress and neuronal loss; iron toxicity is largely based on Fenton chemistry [52–54].

### **14. Conclusion**

Along with the increasing importance of novel cancer immunotherapies in the fight against cancer and their translation from preclinical research to clinical practice, there is an increase in the demand for noninvasive imaging techniques that can measure macrophage responses. Although there are other imaging methods available, including PET, Gd-enhanced MRI, and 19F MRI, using MRI with superparamagnetic iron oxide contrast agents is probably the most promising. Theranostic properties, magnetic gradient actuation forces for transport to the target, and multimodal imaging capacity (MRI-MPI) are a few of the main advantages of these NPs. Additionally, despite the fact that clinical development of SPIOs has been stopped, a number of contrast agents, such as Resovist® and Feromuxytol, are still available.

As a redox-active transition metal, iron is a key player during the process of oxidative stress. Elevated iron promotes the production of ROS, which further depletes the cellular antioxidant GSH and promotes lipid peroxidation, finally triggering ferroptosis and neuronal loss.

As previously mentioned, oxidative stress, protein aggregation, and iron buildup all have a positive feedback loop where one factor encourages the other. By inducing iron buildup, oxidative stress, or protein aggregation, iron oxide nanoparticles

*Iron Oxide Nanoparticles: A Mighty Pioneering Diagnostic Tool But Is It Really Safe… DOI: http://dx.doi.org/10.5772/intechopen.112074*

(IONPs) can turn on this loop. Additionally, IONPs could cause the neurons to undergo apoptotic cellular death. IONPs may cause neurodegeneration given the roles that iron buildup, oxidative stress, protein aggregation, and apoptosis play in neurodegenerative disorders. However, IONPs' properties, such as size, shape, concentration, surface charge, type of coating, and functional groups, have an impact on how toxic they are. Therefore, taking into account the properties of IONPs is crucial when applying them to the CNS.
