Selenium in the Therapy of Neurological Diseases. Where is it Going?

Abstract

Selenium (₃₄Se), an antioxidant trace element, is an important regulator of brain function. These beneficial properties that Se possesses are attributed to its ability to be incorporated into selenoproteins as an amino acid. Several selenoproteins are expressed in the brain, in which some of them, e.g. glutathione peroxidases (GPxs), thioredoxin reductases (TrxRs) or selenoprotein P (SelP), are strongly involved in antioxidant defence and in maintaining intercellular reducing conditions. Since increased oxidative stress has been implicated in neurological disorders, including Parkinson’s disease, Alzheimer’s disease, stroke, epilepsy and others, a growing body of evidence suggests that Se depletion followed by decreased activity of Se-dependent enzymes may be important factors connected with those pathologies. Undoubtedly, the remarkable progress that has been made in understanding the biological function of Se in the brain has opened up new potential possibilities for the treatment of neurological diseases by using Se as a potential drug. However, further research in the search for optimal Se donors is necessary in order to achieve an effective and safe therapeutic income.

INTRODUCTION

Selenium (₃₄Se) is a trace element that is of fundamental importance to human health. In nature, this mineral occurs as a combination of organic and inorganic forms. Inorganic Se compounds, i.e. selenate (SeO₄^2—), selenite (SeO₃^2—) and selenide (Se^2—), are mainly present in water and soil, whereas the organic forms, selenomethionine (Se-Met) and selenocysteine (Sec), are synthesised in plants [1, 2]. Food is a considerable Se supplier, in particular brazil nuts, pork kidney and fish are known for their high Se concentration. The Se content greatly depends on regional soil, but it can be also modulated by food processing such as cooking, or roasting [3]. Humans suffer from health problems due to Se deficiency in areas that are poor in traces of this micronutrient [4]. Se is mainly consumed via food, which is a major source of organic Se at oxidation state II, which is better absorbed than the inorganic forms that are present in water. The total amount of Se in adult humans varies from between 10 to 20 mg and depends on many factors, such as place of living or health [3, 5].

The beneficial properties of Se are attributed to its ability to be incorporated in various proteins [6]. However, unlike other metals that are mainly co-factors of enzymes, Se becomes integrated into the polypeptide chain as part of the amino acid selenocysteine (Sec) [7]. In order to incorporate Se in the catalytic site of the protein chain, the organic form at oxidation level IV must be converted to an inorganic compound, hydrogen selenide. Se is co-translationally incorporated into the polypeptide chain at stop UGA codons located on mRNA. To ensure proper binding of Sec instead of termination of the translation process, mRNA encoding selenoprotein is required to contain a single, specific secondary stem-loop structure, called a selenocysteine insertion sequence (SECIS). Unlike the other 20 amino acids in the genetic code, Sec is synthesised universally on its own tRNA with the ACU anticodon (tRNA[Ser]Sec) by using serine as an intermediate (Seryl-tRNA[Ser]Sec) [8, 9] (Fig. 1). A remarkable aspect of Sec biosynthesis is that this process requires the production of the active Se donor, monoselenophosphate (H₂Se-P), by selenophosphate synthetase (SPS2), which itself is a selenoprotein [10, 11]. Moreover, Se can be non-specifically incorporated into the polypeptide chain as a selenomethionine, by replacing sulphur in methionine. Finally, Se can be non-covalently bound by Se-binding proteins [12].

Fig. (1).

Pathway of selenoprotein biosynthesis. Se-protein biosynthesis begins with the attachment of serine to Sec t-RNA, then SeryltRNA^[Ser]Sec is phosphorylatedby phosphoseryl tRNA kinase and followed by replacement of phosphate by the Se donor selenide (H2Se-P),yielding selenocysteyl-tRNA [Ser]Sec. The resulting selenocysteyl-tRNA [Ser]Sec molecule is a provider of Sec into the growingpolypeptide chain. The Sec residue can be incorporated in the N-terminal or C-terminal region [10].

Most selenoproteins that contain Sec as an integral part of their polypeptide chain exhibit enzymatic activity that is dependent on the presence of Se in their catalytic domain. Based on the location of the Sec residue, mammalian selenoproteins can be divided into two groups. The first group, where Sec is located in the C-terminal region, consists of: thioredoxin reductases (TrxRs), selenoprotein I (SelI), selenoprotein O (SelO), selenoprotein R (SelR) and selenoprotein S (SelS). The second group, characterised by Sec in the N-terminal region, includes glutathione peroxidases (GPxs), iodothyronine deiodinases (Dios), selenophosphate synthetase (SPS2) and the other selenoproteins (Fig. 1) [11, 13, 14]. Selenoproteins are involved in the regulation of many cellular functions e.g. glutathione peroxidases are critical for cellular antioxidant defence (catalysing the reduction of organic hydroperoxides and hydrogen peroxide) [15], iodothyronine deiodinases regulate thyroid hormone activity [6], thioredoxin reductases are important in the regulation of gene expression and redox potential, cell proliferation as well as activation of the immune response [16].

Se-binding proteins also play an important role in maintaining cellular homeostasis. Se-binding protein 1 (SeBP1) is implicated in detoxification processes [17], regulation of cellular growth [18], intra-Golgi protein transport [19] and lipid metabolism [20]. Other Se compounds, which are not related to proteins, have the ability to bind mercury [21], copper and iron [22], thus preventing the toxic effects evoked by these heavy metals [23]. Moreover, selenite can regulate the generation ATP and stabilise the mitochondrial membrane potential [24].

Recent data have placed new emphasis on the long debated and largely unrecognised relationship between Se and brain physiology. However, the specific roles of selenoproteins for normal brain function and neurological diseases are to be elucidated. Here, we will review current findings in Se biology and in the impairment of selenoprotein activity in neurological disorders. We will primarily focus on the most common neurodegenerative diseases, whose incidence increases with age, i.e. especially Alzheimer’s and Parkinson’s diseases, but references to other diseases will also be presented here in order to shape a new view on the role of Se in the treatment and prevention of brain disorders.

SELENIUM AND SELENOPROTEINS IN NORMAL BRAIN FUNCTION

The human brain is characterised by a low (2,3%) content of Se [5]; however, it shows high priority for Se uptake in the case of its dietary deficiency. Even markedly decreased levels of Se in blood as observed in young rats on an Se-deficient 13-week diet, did not induce the significant changes in Se concentration in the brain [25]. The same result was observed in rats that were kept on a Se-deficient diet up to 6 generations. Although a drastic decrease in the level of Se in skeletal muscles, liver and blood was observed, only a small Se decline in the brain was reported in these animals [12, 26]. It was also shown that after injection of a radiochemical labelled SeO₃^2- in rats on basal feed devoid of Se, the highest uptake of the radiotracer (⁷⁵Se) was observed in the brain, in contrast to selenium-adequate nourished rats, where a quite ubiquitous distribution of ⁷⁵Se was observed [27]. It has been suggested that this priority of the brain to use up Se is probably due to its enhanced susceptibility to oxidative stress, caused by intensified oxidative metabolism and limited antioxidant defence [28].

High metabolic activity of the brain causes excessive production of reactive oxygen and nitrogen species. Uncontrolled rise of free radicals caused by the imbalance between their overproduction and enzymatic or non-enzymatic detoxification has deleterious multi-directional effects that, in consequence, cause cell death. Moreover, the brain is abundant in iron, a substrate for the Fenton reaction, and is rich in polyunsaturated fatty acids, i.e. the potential target for peroxidation. These make the brain a privileged organ requiring efficient ROS scavenging that is mainly mediated by selenoenzymes [29, 30]. The expression of 24 selenoproteins was identified in the mouse brain, especially in the hippocampus, olfactory bulb and cerebral cortex (Table 1) [31]. Among these, the highest level was observed for GPx4 as well as SelP and SelW [2], thus suggesting their essential role in the central nervous system. Moreover, the expression and activity of selenoproteins in the brain depend on the amount of Se that is supplied, as some selenoproteins are constitutively expressed while others are induced. This creates a hierarchical ladder of selenoproteins in the brain, where those that are the most important for brain activity remain constant, irrespectively of external Se intake [32].

Table 1.

Selenoproteins in the mammalian brain.

Function	Selenoprotein	Brain Localisation	Subcellular Distribution
Antioxidant enzymes	GPx1	microglia activated astrocytes neurons	cytosolic
	GPx4	neurons astroglia	cytosol and membrane- associated
	SelW	neurons	cytosol
Redox signalling	TrxR1	astrocytes neurons	cytosol mitochondria
	TrxR2	neurons	mitochondria
Metabolism of thyroid the hormone	Dio2	glial	membrane-associated
	Dio3	neurons	membrane-associated
Transport and storage of Se	SelP	neurons	secreted protein binds to heparin
Calcium regulation	SelM	neurons astrocytes	Golgi apparatus, endoplasmic reticulum

Brain locations, subcellular distributions and functions of selenoproteins (GPx - glutathione peroxidase, TrxR - thioredoxin reductases, Sel - selenoprotein, Dio - iodothyronine deiodinases) are presented here [2, 10, 45, 67, 81].

Glutathione Peroxidases

Twenty percent of total Se in the brain is incorporated in the glutathione peroxidase (GPx) family of enzymes [33], which are responsible for catalysing the reduction of hydrogen peroxide, organic hydroperoxides and phospholipid hydroperoxides to alcohol or water by using glutathione, mercaptoethanol, cysteine or other protein thiols as reducing co-substrates [34]. Until now, several isoforms of GPx with different localisations within the cell and substrate specificity have been identified in humans [35, 36]. GPx1 is a major cytoplasmic and mitochondrial antioxidant enzyme, GPx3 is present in the extracellular space and GPx4 is expressed as a cytosolic (c-GPx4), mitochondrial (m-GPx4) or nuclear (n-GPx4) isoform [34, 37]. From the whole family of GPxs, only two isoforms, GPx1 and GPx4, have been detected in the human brain, both in the neurons and in glial cells [38].

GPx1, a widespread selenoenzyme mainly expressed in astroglial cells, is responsible for detoxifying hydrogen peroxides and alkyl hydroperoxides, thus limiting their deleterious effects [36]. Decreased activity of this enzyme resulted in higher brain vulnerability to oxidative damage induced by various toxins. GPx1 knockout mice were shown to be more prone to paraquat toxicity, manifested by a marked increase in H₂O₂ release and caspase-3 activation, thus leading to neurological abnormalities. Hence, full GPx1 expression is necessary, although more beneficial to protect against oxidative stress and neuronal injury is overexpression of the GPx1 [39, 40]. On the other hand, overexpression of GPx1 in mice caused insulin resistance and obesity via reducing the intracellular ROS required for insulin sensitizing. It was associated with a reduction in the insulin-stimulated phosphorylations of the insulin receptor (β-subunit) and impaired protein kinase B activation [41].

GPx4 is mainly expressed in the cerebral cortex, cerebellum and hippocampus, especially as a cytosolic and mitochondrial isoform. Its regional expression differs between the time of embryogenesis and after birth [36]. Firstly, GPx4 mRNA exists in the neural tube and later in the neuroepithelium of the fore-, mid-, and hind-brain in the embryo [42]. During postnatal development, GPx4 mRNA is the highest at postnatal day 15, with a gradual decline thereafter [36]. This enzyme is involved in the elimination of phospholipid hydroperoxides, however its detailed role in the central nervous system is unknown. GPx4-deficient heterozygous mice develop normally, while homozygous embryos die in utero [43]. Overexpression of GPx4 has been shown to be protective against oxidative damage in several cell lines. Besides these antioxidative capabilities, GPx4 possesses antiapoptotic properties, since its selective knockdown by small interfering RNA (siRNA) was shown to activate caspase-3 dependent apoptosis of neuronal cells [43].

Thioredoxin Reductases

Thioredoxin reductases (TrxRs) are a family of antioxidative enzymes that use NADPH to catalyse the reduction of protein disulfides in a variety of macromolecules and molecules. There are 3 types of distinct TrxR genes in mammals: cytosolic thioredoxin reductase (TrxR-1), mitochondrial thioredoxin reductase (TrxR-2) and thioredoxin-glutaredoxin reductase (TrxR-3). Among these, TrxR-1 and TrxR-2 are present in neuronal tissue, where they are responsible for maintaining a proper redox status and are involved in brain development. Expression and activity of TrxR in the brain is strongly retained and does not respond to an Se deficiency in the diet. Aside from its antioxidative role, TrxR is involved in communication between cells, transduction of cellular signalling, DNA synthesis and transcription regulation [44-46]. Trx also plays an important role in cell viability and in the regulation of apoptosis in vitro [47, 48]. Inactivation of TrxR-1 in mouse embryos disturbs cell proliferation, causing retardation of neural tube closure and consequently their death in utero [45, 49]. Likewise, a mitochondrial redox protein Trx-2 is required for proper embryonic development and it is necessary for actively respiring cells. A Trx-2 deficiency was associated with anterior neural tube dysfunction and fetal growth retardation [50]. Moreover, in vitro studies showed that TrxR inhibitor treatment led to enhancement of the ROS level and activation of N-terminal mitogen kinase [51].

Selenoprotein P

SelP, a ubiquitously expressed extracellular glycoprotein, is a useful biomarker of Se status. SelPs are an unusual family that can incorporate multiple Sec residues, while other selenoproteins can integrate only one. Therefore, SelP has been postulated as being involved in both Se intercellular transport and storage [52, 53]. The N-terminal domain of SelP contains one Sec residue, while the other nine are located in the C-terminal. Additionally, SelP has a cluster of histidine and lysine which enables the protein to associate with cellular membranes. SelP binds heparin in a pH-dependent manner, which suggests its attraction to inflammation sites [33]. The primary source of plasma SelP is the liver, from where SelP is transported to peripheral tissues, including brain [54, 55]. Its role in the CNS is unique, because brain tissue can take up tracer Se in a form of SelP and is unable to take up directly radioactive molecule⁷⁵Se [52]. Yet, recent data show that a deficiency of hepatic SelP does not affect brain Se levels, which indicates that this protein could be produced locally. Hence, as long as SelP expression in the brain is preserved, the Se level in the brain is independent of its plasma concentration [56]. Moreover, during Se deficiency, the brain unregulated Se level, using a specific uptake of SelP by the receptor for apolipoprotein E2 (ApoER2) [54, 55]. Extracellular SelP binds to ApoER2, which initiates endocytosis and enables the selenocysteine supply. Presumably, in those conditions astrocytes are the main source of extracellular SelP, which in turn is taken up by neurons using the ApoER2 [57, 33]. Aside from evidence that SelP serves to transport Se to brain tissue and to maintain its homeostasis, other suggesting function is a survival factor for neurons [58] as well as microtubule stabiliser. Fluorescence resonance energy transfer and immunoprecipitation assay have shown that SelP interacts with tubulin. Hence, SelP can play an important role in regulating the dynamic property of microtubules, which in turn determines cell polarity [59]. Genetic defects in this selenoprotein are associated with severe health issues. SelP−/−knockout mice display a neurological phenotype with subtle disruption of memory processes and spatial learning [60]. Moreover, other researchers have shown reduction in brain Se levels followed by a decrease in Se-dependent enzyme activity and impaired motor coordination in the same model [52, 61, 62]. Burk et al. confirmed that the knock-out of SelP or apoER2 significantly depleted the brain’s Se pool [63]. However, a combination of SelP KO with a restrictive Se-deficient diet resulted in severe degeneration of brainstem axons as well as neurological dysfunction within days of having started the Se deficient diet [64, 65].

Other Selenoproteins

SelM is abundant in the brain, with the highest expression in the olfactory bulb and cerebellum. In the SelM structure, there are thioredoxin-like domains and an active-site in the form of the CXXU motif, where C represents cysteine (Cys) and U represents selenocystein (Sec). This sequence is responsible for the formation of reversible selenenylsulfide bonds as well as it may also function as a redox regulator. SelM is involved in antioxidant function, [66], neuroprotection and regulation of intracellular calcium homeostasis [67]. In addition the neuroprotective function of SelM may result from its direct interaction with galectin-1. This mammalian lectinis widely expressed in the brain, where it regulates cell proliferation, differentiation as well as cell death [68].

SelW is highly expressed in the developing and adult brain [69, 70]. This smallest mammalian selenoprotein is widespread in cortical and hippocampal neurons, especially in dendrites and synapses both in the cytosol and plasma membrane [71]. Its expression in the brain is well maintained under Se deficits and is highly dependent on the SelP concentration. It was shown that SelP deficiencies resulted in greatly reduced levels of SelW expression in the brain [72, 73]. Like most selenoproteins, SelW is involved in maintaining redox homeostasis [74]. It binds to glutathione, acting as a glutathione-dependent antioxidant, and its overexpression reduces cellular sensitivity to hydrogen peroxide [75]. SelW was additionally shown to bind with 14-3-3 proteins β and χ and to regulate their oxidation state towards reduction. Since 14-3-3 proteins coordinate interaction of various kinases and phosphatases with receptors and structural proteins in neurons, SelW may indirectly regulate phosphorylation-dependent processes in the brain [76, 77]. Additionally, SelW is involved in p53- and p21-dependent regulation of the cell cycle [78, 79]. Knocking down SelW resulted in cell cycle arrest at the G1 stage via modulation of mitogen-activated protein kinase kinase 4 (MKK4) and MAPK signalling [78].

SelR is a crucial cytoplasmic and nuclear selenoprotein that contributes to neuronal antioxidant defence via scavenging reactive oxygen species. It is a member of the methionine sulfoxide reductases family type B (MsrB1) which catalyse the reduction of methionine sulfoxide (R-MetO) to methionine [80, 81]. SelR interacts with Trx and the chaperone protein clusterin, however, the role of this interaction is still unknown [82].

SELENIUM AND SELENOPROTEINS IN NEUROLOGICAL DISORDERS

Since Se and selenoproteins have an important physiological function in neurons, astrocytes and microglia, their down-regulation or damage may lead to dysfunction of the brain. It was proved that the Se level in the brain shows a tendency to decrease during aging and is related to cognitive decline [83-85]. A 15-day Se-deficient diet decreased brain protection and as a result, significantly increased the dopamine turnover in SN concomitant with a decrease in GPx activity in the SN and striatum [86]. It has been suggested that Se deficiency may cause irreversible changes in the neurons, thus leading to cognitive impairment and neuro-degenerative diseases. Moreover, the genetic inactivation of selenoproteins leads to progression of pathology in CNS. Several Se compounds, such as sodium selenite, ebselen, or other organo-selenium donors have been shown to have promising therapeutic potential against neuropathology. Many studies in vitro and in vivo have postulated that Se is important to reduce oxidative stress and other detrimental agents in Alzheimer’s and Parkinson’s diseases as well as in epilepsy. Apart from direct antioxidative properties, Se was demonstrated to be involved in regulation of various abnormal molecular processes, that are common to all brain disorders.

A growing body of evidence suggest that mitochondria dysfunction, including increased mitochondrial fragmentation and decreased mitochondrial fusion, defects in oxidative phosphorylation, impaired calcium influx as well as mitochondrial membrane potential dissipation are associated with selective cell death [87]. Recently, it was demonstrated, that Se may regulate mitochondrial biogenesis and may improve mitochondrial function. Treatment of murine hippocampal HT22 cells with sodium selenite resulted in the increased of the level of PGC-1α (peroxisome proliferator-activated receptor gamma coactivator 1-alpha) and NRF1 (nuclear respiratory factor 1), as well as respiratory chain proteins: cytochrome c and cytochrome c oxidase IV (COX IV). This effect was shown to be independent from antioxidative properties of Se and, in general, lead to improvement of the mitochondrial respiratory rate [24]. The similar effect on mitochondrial level and function was observed in SelH overexpressing HT22 cells [88]. Moreover, the SelH elevated the mitochondrial biogenesis through activation of protein kinase A-CREB-PGC-1α and Akt/protein kinase B-CREB-PGC-1α pathways [89].

Apart from its stimulating effect on mitochondrial biogenesis, Se was also shown to regulate calcium homeostasis. Xu et al. demonstrated that Se-deficiency increased the intracellular calcium concentration in the brain of chicken, resulting in mitochondrial damage, fusion of nuclear membrane and nuclear shrinkage as well as apoptotic death of neuronal cells [90]. Moreover, treatment with selenium in a form of sodium selenite prevented the cellular calcium overload, oxidative damage and death of dorsal root ganglion (DRG) sensory neurons of rats [91]. Se was also speculated to have the important role in regulation of brain inflammation, since its deficiency resulted in the elevation of cytokines: tumor necrosis factor (TNFα), cyclooxygenase 2 (COX 2), inducible nitric oxide synthase (iNOS) and prostaglandin E (PGE) synthase level in different regions of chicken brain [92].

Additionally, one of the first studies in humans has confirmed Se participation in both prevention and treatment of brain diseases [93]. However, it remains controversial whether a high Se level is safe to human health. Some reports have showed that Se could have a cytotoxic effect by inducing nitric oxide synthase (iNOS), NO-liberation and activation of caspase-3 [94]. It was indicated that high doses of Se, through depletion of GSH, induced oxidative stress, which led to a decrease in cholinergic signalling and degeneration of cholinergic neurons [95]. More research has to be conducted to study the impact of Se on neurological diseases.

Alzheimer’s Disease

Alzheimer’s disease (AD) is a neurodegenerative disorder of the elderly which is characterised by deposition of amyloid β (Aβ) in extracellular senile plaques and hyperphosphorylated tau in intracellular neurofibrillary tangles (NFT) [96-98]. A clinical picture of the disease is manifested by progressive loss of memory and speech as well as by cognition and behavioural deficits [99]. According to the classical hypothesis, the neurotoxic effect of Aβ is mediated by excessive free radical liberation and disturbances in Ca²⁺ homeostasis [96, 100]. An imbalance between free radical formation and efficiency of the antioxidant system is responsible for the widespread occurrence of protein and lipid oxidation, nucleic acid damage and mitochondrial abnormalities [99]. In addition, transient or sustained mitochondrial complex dysfunctions resulting in massive leakage of ROS lead to further neuronal oxidative injuries. Oxidative stress and neuronal disturbances result in cytokine release and in the activation of microglial inflammatory processes [101, 102].

Studies in humans have suggested the possible role of Se in the prevention and treatment of AD, either alone or in combination with other elements [93]. Some research showed a direct correlation between the reduced plasma concentration of Se and the cognitive decline in AD patients when compared to healthy individuals [103]. The decline in Se levels in the plasma, observed in the early stage of AD, was independent of nutritional status [104, 105, 106, 107]. The Se level was also significantly decreased in AD brain tissue, especially in the hippocampus and frontal, parietal, temporal and occipital lobes [108, 109]. Inversely, the latest study showed a significant increase in Se levels in the globus pallidus, superior temporal gyrus, and frontal cortex of Alzheimer’s disease patients [110]. Moreover, Gerhardsson et al. showed that the level of Se in the plasma and in cerebrospinal fluid from AD patients did not change when compared with the control group [111]. Despite these inconsistencies, which need to be further examined and interpreted, disturbances in selenoproteins, which are responsible for antioxidant defence in AD, have been better documented [112].

It was previously shown that changes in the GSH level and the increase of its oxidised form in the frontal cortex are associated with AD. This is probably connected with an increased level of peroxides that leads to activation of GPx which oxidises GSH to GSSG. Oxidised glutathione can be further reconverted to a reduced form by GR. Although the activity of GR was elevated in the hippocampus of AD mice [99, 113], it was unable to metabolise the whole pool of GSSG [112]. This leads to disturbances in the GSH/GSSG ratio shifting the balance to the oxidised form in the human AD brain [114]. Disturbances of GSH-dependent enzymes were confirmed in brain homogenates prepared from triple-transgenic mice with the PS1 _{M146 V}, APP _Swe, and tau _P301L mutation [115].

A noteworthy aspect is that in AD both the level and activity of other selenoenzymes are significantly altered. Overactivation of TrxR concomitant with the decrease of its substrate (Trx) were observed in the amygdala and cerebellum [116]. This is probably a compensation mechanism that is activated in order to prevent cells from oxidative stress. However, the decline in the Trx level was responsible for increased sensitivity to peroxynitrite [117], thus further exacerbating the process of neurodegeneration. Moreover, the level of SelM was significantly decreased in the brains of transgenic mice with human mutant presenilin-2 (N141I) [118, 119]. On the contrary, the concentration of SelP was significantly increased in the choroid plexus and the cerebrospinal fluid of AD patients, as compared to healthy controls, thus suggesting a higher brain Se requirement in order to prevent neurodegeneration [120]. A recent study of gene expression in AD determined that SelP was one of 240 genes upregulated in AD as compared to age-matched controls [121]. Additionally, SelP was detected in Aβ plaques and NFT tangles [118, 122]. Besides the important role of SelP and SelM in the global response to oxidative stress, they were shown to protect against transition metal-induced Aβ aggregation and toxicity [123, 124].

Although the Se and selenoprotein level is altered in the brain, cerebrospinal fluid and blood of patients with AD, and an association between Se status and cognitive function was observed, until now there has been no consistent clinical evidence showing the beneficial effects of Se supplementation in the treatment of AD. However, findings in molecular biology allow to make speculations on its potential preventive relevance. The treatment of rat hippocampal neurons with organo-Se-compounds- (PhSe)₂ and ebselen-had a protective effect against Aβ peptide toxicity and increased the level of SNAP-25, which is a marker of nerve terminals [125]. Supplementation with inorganic Se in a form of sodium selenite resulted in a significant reduction of lipid peroxidation and protein carbonylation in the cerebral cortex and hippocampus of rats after intracerebroventricular injection (ICV) with streptozotocin (STZ) [84]. Chronic administration of the inorganic Se compound as sodium selenate, characterized by less toxicity than selenite, was shown to reduce NFT formation and to prevent memory and motor deficits. These effects were mediated by the stabilisation of PP2A-tau complexes, thus resulting in a decrease in tau phosphorylation [126, 127]. Sodium selenate also attenuated the inhibition of PP2A and tau hyper-phosphorylation induced by traumatic brain injury [128]. Moreover, selenate treatment was shown to positively alter the phosphorylation status of key proteins involved in oxidative stress, energy metabolism and protein degradation in cellular model of AD as indicated in broad phospho-proteomics study. Apart from playing important roles in regulation of redox homeostasis, ATP generation, and misfolded proteins clearance, selenate-treatment also resulted in reduction of homocysteine, phospho-tau as well as Aβ levels [129].

In accordance with these data, SeMet treatment of triple transgenic AD mice ameliorated cognitive deficits by reducing the level of total tau and decreasing its phosphorylation via modulation of the expression and activity of PP2A and glycogen synthase kinase 3β. Moreover, the restoration of synaptic proteins, including synaptophysin and postsynaptic density protein 95 in the hippocampus and cortex, and inhibition of inflammatory response were observed in these mice treated with SeMet [130]. A recent study has shown the therapeutic effect of the novel organic Se compound p,p'-methoxyl-diphenyl diselenide (MeOPhSe)₂ against oxidative-nitrosative stress caused by ICV-STZ. Supplementation with organic Se improved spatial learning and memory in rats after STZ administration. This beneficial effect observed in the rat cerebral cortex and hippocampus was mediated by a reduction of the ROS level and nitration of tyrosine residues [131]. Some data point to the protective effects of another organic Se compound, ebselen 2-phenyl-1,2-benzisoselenazol-3(2H)-one, which possesses antioxidant activity in vitro and was shown to affect GPx and AchE activity in rat brains treated with STZ [132, 133]. Moreover, a recent study confirmed the beneficial effect of Se on neurotoxicity induced by aluminium chloride (AlCl₃). Lakshmi et al. showed that this micronutrition effectively prevented the harmful effects of this toxin in behaviour and biochemical changes, and a decrease in brain oxidative damages, neuronal inflammation and improvement of the working and attention memory were observed [134].

Furthermore, the beneficial effects were demonstrated of Se treatment when administered with other neuroprotective compounds. Combined therapy of inorganic Se in a form of sodium selenite with natural carotenoid dicarboxylic acid provided better neuroprotection in rats treated with STZ by significantly reducing the level of lipid peroxidation and elevating the content of GSH as well as GPx, glutathione S-transferase (GST) and catalase activities [135]. Shrivastava et al. presented the beneficial effect of the co-administration of Se and N(2-hydroxyethyl) ethylenediamine triacetic acid (HEDTA) against aluminium (Al)-induced toxicity in rats. These positive effects of combination therapy with Se resulted from ameliorating the increase of acetylcholinesterase (AChE) and at the same time elevated butyrylcholinesterase (BuChE) activities in the plasma after Al administration. Moreover, this combined therapy led to an increase in antioxidative enzyme activities (GR, GST, GPx, glucose-6-phosphate dehydrogenase-G6PDH) in the forebrain [136].

Parkinson’s Disease

Parkinson's disease (PD) is the second most common neurodegenerative disease after AD. PD is a progressive and complex disease with heterogeneous clinical features including motor and non-motor symptoms. This disease is characterised by progressive neuronal loss, especially in the substantia nigra pars compacta (SN), and by the development of intraneuronal α-synuclein (ASN) inclusions known as Lewy bodies (LB). Moreover, the latest data have implicated the importance of extracellular ASN oligomers in the progression of PD pathology in the brain. Our own data indicated that extracellular ASN stimulated free radical generation, NOS activation and induced NO-dependent caspase-3 activation and poly(ADP-ribose)polymerase 1 (PARP-1) cleavage, thus leading to apoptotic cell death [137, 138, 139, 102]. On the other hand, oxidative stress induced ASN oligomerisation and its extracellular liberation, thus promoting its brain propagation. [102, 137, 139-141].

Although the possible role of Se in the etiology of PD as well as its significance in the treatment of this disease have not been studied in as much detail as in AD, some new promising data have emerged. It was observed, for example, that there were no differences in the serum Se concentration between PD and control groups [142], but its midbrain level was significantly increased in PD [110]. However, these studies were conducted on a small population and need to be further confirmed using large cohorts. Nevertheless, deregulation of oxidative defence in PD has been better documented. Post-mortem studies demonstrated a significant decrease of the total GSH level and changes in the expression and activity of GSH-dependent enzymes exclusively in the SN and striatum of PD patients which did not involve other brain regions [143, 144]. Using immunohistochemistry, it was shown that the level of GPx-1 was significantly increased in the hypertrophied microglia as well as in degenerated neurons. Moreover, this enzyme was shown to be mainly localised around LB, thus suggesting its protective role against oxidation of LB-degrading enzymes [38]. The potent antioxidative role of this enzyme in PD was further confirmed by showing that GPx1 knockout mice were more sensitive to neurotoxin-induced SN injury [105]. Interestingly, GPx1 overexpression in nigral dopaminergic neurons in vivo had a protective effect against 6-OHDA toxicity [145, 146]. Not only the GPx1 but also the GPx4 level was changed in the PD brain. Post-mortem immunostaining showed that the total expression of GPx4 in SN was significantly reduced in PD brains as compared to control, but when related to the density of surviving nigral neurons it was increased [147]. Moreover, this enzyme was shown to be associated with TH-positive degenerating axons in the putamen, thus suggesting that up-regulation of GPx4, evoked by a buildup of oxidised dopamine in the axons, mitigates oxidative stress. It was also speculated that changes in GPx4 in dystrophic neurons could be related to this enzyme’s colocalisation with neuromelanin [148]. GPx4 was also shown to co-localise with ASN-positive LB in SN, which may be a response to elevated oxidation of this protein [147]. Another selenoprotein indicated to be concentrated within the cores of LB is SelP. Similar as in GPx4, expression of this protein in relation to cell density was increased in the SN of PD patients, but its level in the putamen was not changed [149]. In PD pathology not only Se-dependent antioxidative enzymes are affected; both the level and activity of other enzymes related to the GSH metabolism are also changed. Increased expression of glutathione S-transferase (GST), which is a major enzyme involved in the regulation of oxidative stress, was observed in SN as well as in cerebrospinal fluid from PD patients [150-152].

Previous in vitro studies pointed to the neuroprotective effect of Se against various toxins that cause permanent symptoms of Parkinson's disease. Se dietary supplementation reduced the generation of reactive nitrogen species (RNS) and ameliorated the reduction of GPx levels induced by methamphetamine (MA) in dopaminergic neurons [123, 153]. Moreover, sodium selenite blocked the decrease in DA and its metabolites caused by the administration of MA [154] or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) [154, 155]. It was also documented that Se prevented dopaminergic cell death in murine SN induced by the administration of MA or MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) [154, 155]. Furthermore, sodium selenite enhanced the activity of antioxidant enzymes, particularly GPx, GR, GST, and CAT, and it elevated the GSH level and GSH/GSSG ratio in rats after 6-hydroxydopamine treatment [156]. Moreover, sodium selenite administration led to an improvement in locomotion and muscular co-ordination in the rat model of PD [157]. More recent neurochemical, histopathological and behavioural data have pointed to the potent neuroprotective effect of novel organic Se compounds, such as ebselen in various PD models. Both in vitro and in vivo research has suggested that ebselen reversed the toxic effects mediated by MPP⁺ or MPTP treatment, prevented dopaminergic cell apoptosis, thus leading to improvement in motor performance, locomotor activity and neurological score. Apart from increasing GPx activity, ebselen was shown to act directly on mitochondrial complex I activity, thus leading to a decrease in free radical generation and energy deficits [158].

Other Diseases

Epilepsy

Epilepsy was one of the earliest characterised neurological diseases manifested by recurrent unprovoked seizures [159]. The endpoint of untreated intractable epilepsy is extensive hippocampal neuronal loss and reactive gliosis [103]. The mechanisms underlying this disease are still substantially unknown. However, there is experimental evidence suggesting that oxidative stress may be involved in the etiopathology of this disease. Frequent and chronic seizures lead to anoxia, which it turn evokes oxidative stress [159, 160] that is concomitant with elevated protein carbonylation and DNA oxidation [161, 162]. On the other hand, high oxidative status increased the severity and recurrence of epileptic seizures, thus creating a self-perpetuating vicious cycle.

It was previously suggested that the utilisation of Se may be increased in epilepsy since a decreased level in Se in the serum and erythrocytes from epileptic patients was noted [163, 164]. It is also believed that Se depletion in the brain during epilepsy constitutes a vital factor for the origin of seizures [163, 165, 166]. Nevertheless, information on the role and contribution of selenoproteins in brain disease is still limited. Some antioxidant abnormalities, such as increased activity of CAT concomitant with decreased activity of GPx, were observed in this disease [162]. However, experimental data from animal studies showed that Se-related anti-oxidative defence is markedly changed in epilepsy; for example, treatment with pilocarpine, which induced seizures and an epilepticus-like state, resulted in enhancement of the activity of SOD, GPx and CAT [167]. Similarly, kainic acid induced a significant increase in GSH oxidation, but it did not change the total GSH level in the rodent brain cortex [168]. Moreover, animals kept on an Se-deficient diet were more susceptible to neuropathological changes when exposed to glutamate or kainate [64, 169]. Interestingly, SelP and GPx-4 knockout mice displayed the tendency to develop neurological seizures [62, 170].

Based on these data, it seems plausible that Se administration might be beneficial in the therapy of epilepsy. A study on a pentylentetrazole (PTZ) rat model of epilepsy showed that Se treatment inhibited free radical production, regulated calcium-dependent processes, and supported the antioxidant redox system [171]. Additionally, chronic treatment of sodium selenate reduced the number of seizures induced by either PTZ injection or electrical stimulus of cornea or by amygdala kindling [172]. Sodium selenite or seleno-DL-methionine attenuated PTZ induced seizures in mice via inhibition of cyclooxygenase and reduction in prostaglandin E1 receptor activation [168]. There have also been human case reports showing that Se treatment has protective effects on epilepsy and refractory epilepsy-induced oxidative injury. It was shown that sodium selenite supplementation leads to reduction in the number of seizures and a decrease in neuronal damage [166]. Moreover, selenite inhibited free radical production and supported the antioxidant redox system via an increase in the GSH level and GPx activity in human erythrocytes [163, 173].

Cerebral Ischemia

Cerebral ischemia, characterised by high morbidity and mortality, remains the third main cause of human death and disability. Occlusion of brain arteries responsible for cerebral ischemia leads to insufficient oxygen and glucose delivery to support cellular homeostasis. This initiates a complex cascade of metabolic events, involving oxidative-nitrosative stress, inflammation and apoptosis, thus leading to neuronal damage and death [174]. The main result of ischemia is elevated glutamate release, which is responsible for excitotoxicity and an increased cytosolic calcium concentration. The NMDA-receptor - mediated calcium influx was shown to activate both constitutive (neuronal and endothelial) and inducible isoforms of nitric oxide (NO) synthase. In addition to an elevated NO level, brain ischemia leads to the generation of superoxide through various mechanisms, such as activation of NOS and xanthine oxidase, or disturbances of the mitochondrial electron transport chain [175]. Overproduction of free radicals, leads to inactivation of detoxification systems and perturbation of antioxidative defence mechanisms. It was previously demonstrated that elevated consumption of GSH with its maximum loss in the striatum and hippocampus as well as depletion of the enzymatic defence system (GPx, GR, GST) take place in ischemic brain tissue [176, 177]. In cerebral ischemia, oxidative stress is often accompanied by depletion of essential trace elements in the organism [175]. This also relates to Se, which was documented to be significantly decreased in the ischemic brain as compared with the control subject [175]. Taking into account the scavenging features of Se, treatment with Se-derived compounds was suggested as a valuable therapy in ischemia reperfusion damages [178]. An in vitro study using murine hippocampal HT22 cells indicated that Se as sodium selenite protected against glutamate toxicity and hypoxia-induced cell death by preventing changes in the mitochondrial membrane potential and ROS generation. Moreover, Se increased the expression of mitochondrial biogenesis regulators while at the same time decreasing the level of autophagy modulators [179]. In the rat ischemia model with middle cerebral artery occlusion (MCAO), sodium selenite administration increased the activity of CAT, SOD and GPx, and prevented the GSH decline, leading to decreased brain lipid peroxidation [178]. Another report indicated that sodium selenite restored the level of mitochondrial ATP, cellular Ca²⁺ and decreased the activity of caspase-3 [180]. Moreover, Se as sodium selenite inhibited monoamine oxidase activity in the mitochondria, which are also capable of producing ROS during ischemia/reperfusion. The intake of inorganic Se can also have a suppressive effect against DNA damages and PARP activation [178, 180]. More recent data based on histopathological analyses have suggested that sodium selenite treatment improved neuron density and decreased perineuronal and pericapillary edema in the prefrontal cortex and hippocampus of the rat model of ischemia/reperfusion. Moreover, the same study showed that inorganic Se treatment significantly decreased the level of inflammatory cytokines such as interleukin-1 beta (IL-1β), tumor necrosis factor alfa (TNF-α) and, at the same time, increased the levels of NGF in the prefrontal cortex and hippocampus [148]. Interestingly, the organic Se compound, ebselen, significantly decreased malondialdehyde (MDA) and NO levels as well as increased the activity of antioxidative enzymes, thus leading to morphological improvement of brain tissue in the rat ischemia/reperfusion model [181]. A promising effect was observed using a combination of sodium selenite and melatonin (Se-Mel) therapy on transient cerebral ischemia [176]. In the MCAO model, Se-Mel attenuated protein and lipid oxidation, caspase-3 activity and the NO level were reflected in improvement of animal behaviour [176].

DOES A SELENIUM-BASED DRUG DESIGN HAVE THERAPEUTIC POTENTIAL?

With accumulating evidence for oxidative injuries and disturbance of antioxidative defence as important factors in disorders of CNS, several trials have explored the efficiency of antioxidants in the prevention and treatment of those diseases. The anti-inflammatory and anti-aging properties of vitamin E, C or β-carotene have been demonstrated in various studies. Unfortunately, promising results were not reflected in clinical trials [182-185]. Therefore, the search for new compounds that might have potential application in the treatment of neurodegenerative and other brain diseases still continues. Taking into account the multiple protective effects of Se, the possibility of using this compound as a protective drug against neuronal cell death can be plausible. Thus current scientific challenge is finding the appropriate Se donor with high bioavailability and respectively low toxicity. The large number of experimental data suggests that neurotoxicity of inorganic Se species exceeds that of the organic-Se-compounds [186]. However, there are some exceptions, like methylseleninic acid, L-selenocystine or selenodiglutathione, that were shown to induce cell death already in low μM concentrations. On the other hand, despite its high bioavailability, the toxicity of inorganic sodium selenite (LD50 3.5 mg/kg body weight in rats [187] and 1.2-1.9 mg/kg body weight in sheep after oral administration [188]) is significantly higher than that of ebselen or SeMet [189]. Interestingly, it was demonstrated that the cytotoxicity of sodium selenite strongly depends on its uptake and cellular accumulation, which in turn is regulated by the extracellular environment. The extracellular reduction of selenite to selenide is mainly mediated by cysteine and leads to a high-affinity uptake of this reduced Se form, causing its accumulation and cell death. The efficacy of this process is determined by the cystine/glutamate antiporter (x_c^-)-dependent cystine uptake, its intracellular reduction to cysteine by NADPH-dependent redox systems and cysteine resecretion by multidrug resistance proteins [190, 191]. These findings highlight that therapeutic potential of reducible selenium compounds strongly depend on the type of the tissue and conditions of individual patients, hence the possibility of using certain Se species in therapy is limited and controversial [189]. Noteworthy is that organic forms of Se at IV oxidation state are characterized by higher bioavailability and lower toxicity, thus administration of these compounds allows to achieve a higher Se concentration in tissue [192-194].

One of the first synthetic organic Se compounds, ebselen, a mimetic for glutathione peroxidases, was previously considered as a potential pharmacological agent (Fig. 2A) [158,195, 196]. The mechanisms underlying the neuroprotective effect of ebselen are still not completely understood, but they are certainly related to its antioxidant and anti-inflammatory properties [197, 198]. Several pre-clinical animal studies have supported the neuroprotective effect of this chemical in stroke. It was shown that ebselen not only provided protection against oxidative damage-induced neuronal death from focal ischemia in hypertensive rats [199], but that is also had beneficial effects in transient forebrain ischemia [200, 201]. Moreover, initial clinical studies with humans indicated that ebselen may be a prophylactic neuroprotective agent in acute ischemic stroke [202, 203]. More recent ongoing trials have demonstrated that ebselen displayed efficacy for reducing the infarct volume by 27% on average in nine focal ischemia studies, with 10 of the 16 experimental contrasts showing protection [204], which seems promising for future therapies for stroke.

Fig. (2).

Molecular structure of organic selenium compounds. Ebselen (A) [133], Selenomethionine (B) [215], Selol (C) [210],Selenoneine (D) [216] described in this study.

Other clinical trials that have been conducted concerned the therapeutic use of Se in AD treatment. However, in those studies Se was tested in combination with other compounds. In one small study, geriatric patients received a mixture of inorganic and organic Se with Vitamin E. These patients significantly improved in several cognitive parameters as compared to control placebo [205]. A larger study with Se in the form of SeMet (Fig. 2B) and Vitamin E is currently undergoing the “preadvise” trial, which is being held on more than 10,000 healthy men between 60 and 90 years old. The initial goal of this trial is to study the effect of this treatment formulation on preventing AD and other brain disorders [206]. However, the latest study coming from “SELECT” clinical trial bring negative reports about possible application of SeMet in treatment of human diseases. In this experiment the l-selenomethionine intake not only had any positive impact on prostate cancer prevention, but also increased the risk to develop type 2 diabetes mellitus. The adverse effects of this compound resulted from its random incorporation into various proteins instead of methionine that, together with its long biological half-life, constitutes the reason for the failure of the therapy [207, 208].

Furthermore, research is being conducted in several research centres on a new Se compound, called Selol (Fig. 2C) [209-211]. Selol is an organic mixture of selenitetriglycerides which are obtained on the basis of sunflower oil. A compound containing Se at IV oxidation state meets the requirements of low toxicity and is characterised by fast absorption and distribution as well as high bioavailability [192, 193, 212, 213]. Until now, Selol has undergone various experimental and clinical investigations as an anticancer drug, and currently it is at the final stage of pre-clinical studies [211, 212, 214]. Additionally, due to the ability for Selol to provide a high Se concentration in the brain [193], its neuroprotective effects are currently being investigated. While testing this compound in vitro, it was found that it has the ability to suppress oxidative-nitrosative stress in neuronal cells. The mechanism of Selol’s cytoprotective effect involves stimulation of the endogenous antioxidant system via elevation of the level and activity of the Se-dependent enzyme – glutathione peroxidase (GPx) [220]. However, many more studies should be conducted so that the potential in vitro antioxidant effects of the organic Se (IV) form can be harnessed in the clinic of neurological diseases.

The latest studies introduced a novel organic Se-containing compound, selenoneine (Fig. 2D), isolated from the blood and tissues of bluefin tuna. However, the biological significance of this compound is still unclear. It was demonstrated that apart from strong antioxidant capacity, selenoneine has the ability to bind heme proteins, such as hemoglobin and myoglobin and to protect them from iron auto-oxidation. It was also shown to prevent MeHg accumulation and toxicity via acting on organic cations/carnitine transporter, OCTN1 [216, 221].

CONCLUSIONS

Recent studies have identified Se as crucial for preserving brain function and preventing age-related neurodegenerative disorders. An insufficient supply of Se may have a detrimental effect on brain cells by exacerbating neuronal dysfunction and death. Undoubtedly, the remarkable progress that has been made in understanding the biological function of Se in the brain, especially in relation to the antioxidative and anti-inflammatory functions of selenenoproteins, has also opened up new, potential possibilities for treatment of neurodegenerative diseases by using Se as a potential drug (Fig. 3). Thus, an assessment of optimal doses and forms of Se supplementation is still a matter of debate. On account of the higher toxicity of inorganic Se forms and, in consequence, their limited application in therapy, there is a need for a further search for Se donors, especially in organic form, in order to achieve an effective and safe therapeutic income.

Fig. (3).

Summary of selenium therapy effects in neurological diseases. Aβ-amyloid β, CAT-catalase,GPx-glutathione peroxidase, GSHglutathione,pTau-phosphorylated tau protein, Sel-selenoprotein, SOD-superoxide dismutase [167, 217-219].

CONFLICT OF INTEREST

The authors confirm that this article content has no conflict of interest.

LIST OF ABBREVIATIONS

Aß =

amyloid β

AChE =

acetylcholinesterase

AD =

Alzheimer’s disease

APP =

β-amyloid precursor protein

ALS =

amyotrophic lateral sclerosis

ApoER2 =

apolipoprotein receptor 2

ASN =

α-synuclein

CAT =

catalase

CSF =

cerebrospinal fluid

Dio =

iodothyronine deiodinase

GPx =

glutathione peroxidise

GSH =

glutathione

GR =

glutathione reductase

GSSG =

glutathione disulfide

GST =

glutathione S-transferase

G6PDH =

glucose-6-phosphate dehydrogenase

HEDTA =

N(2-hydroxyethyl)ethylenediamine triacetic acid

ICV-STZ =

intracerebroventricular injection of streptozotocin

iNOS =

inducible nitric oxide synthase

LD50 =

median lethal dose

LB =

Lewy bodies

MCAO =

ischemia model with middle cerebral artery occlusion

MDA =

malondialdehyde

MPP+ =

1-methyl-4-phenylpyridinium

MPTP =

1-methyl-4-phenyl-1,2,3,6-tetrahydro-pyridine

NFT =

neurofibrillary tangle

NRF1 =

nuclear respiratory factor 1

PARP-1 =

poly(ADP-ribose) polymerase-1

PD =

Parkinson’s disease

PGC-1α =

peroxisome proliferator-activated receptor-γ coactivator 1 alpha

PGE =

prostaglandin E

PP2 =

protein phosphatase 2

pro-NGF =

precursor of nerve growth factor

ROS =

reactive oxygen species

Sec =

selenocysteine

SECIS =

selenocysteine insertion sequence

Sel =

selenoprotein

SeBP1 =

selenium-binding protein 1

SeMet =

selenomethionine

SN =

substantia nigra pars compacta

SPS2 =

selenophosphate synthetase

TH =

tyrosine hydroxylase

Trx =

thioredoxin

TrxR =

thioredoxin reductase