Molecular Aspects of Neurodegeneration and Neuroprotection

Docosahexaenoic acid and arachidonic acid are major polyunsaturated fatty acids in neural membrane glycerophospholipids. Docosahexaenoic acid and arachidonic acid are metabolically and functionally distinct molecules that have opposing physiological functions. Docosahexaenoic acid is metabolized to docosanoids, whereas arachidonic acid is metabolized to eicosanoids. Like their precursors, docosanoids and eicosanoids are different types of lipid mediators, which play important and opposing roles in modulating inflammatory reactions, oxidative stress, neuroprotection, and neurodegeneration. Increase in levels of eicosanoids occurs in acute neural trauma (stroke and traumatic injury to brain and spinal cord) and neurodegenerative diseases (Alzheimer disease, Parkinson disease, and Huntington disease), whereas consumption of DHA increases levels of docosanoids, which have antioxidant, anti-inflammatory, and anti-apoptotic properties. Synthesis of docosanoids is an endogenous neuroprotective mechanism against acute neural trauma and neurodegenerative diseases.

Neuroinflammation is a host defense mechanism involved in restoration of normal neural cell function. Most major CNS diseases are characterized by neuroinflammation. Following brain injury, microglial cells are activated and initiate a rapid response, which involves cell migration, proliferation, release of cytokines/ chemokines and trophic and/or toxic effects. Cytokines/chemokines mediate the stimulation of phospholipases A2 and cyclooxygenases. Stimulation of these enzymes results in breakdown of membrane glycerophospholipids with release of arachidonic acid and docosahexaenoic acid. Oxidation of arachidonic acid produces proinflammatory eicosanoids. Lyso-glycerophospholipids, the other products of reactions catalyzed by phospholipase A2, are converted to pro-inflammatory platelet-activating factor. Eicosanoids and platelet activating factor intensify neuroinflammation. Lipoxin, an oxidized product of arachidonic acid through 5-lipoxygenase, is involved in resolution of inflammation and is anti-inflammatory. Docosahexaenoic acid is metabolized to resolvins and neuroprotectins. These lipid mediators retard the generation of eicosanoids. Levels of eicosanoids are markedly increased in many neurological disorders. Docosahexaenoic acid and its oxidized products retard neuroinflammation by inhibiting transcription factor NF-κB, preventing cytokine secretion, blocking the synthesis of eicosanoids, and modulating leukocyte trafficking. Depending on its timing and magnitude in brain tissue, inflammation serves multiple purposes. It is involved in protection of uninjured neurons and removal of degenerating neuronal debris and also in assisting repair and recovery processes. The dietary ratio of arachidonic acid to docosahexaenoic acid modulates inflammation in acute neural trauma and neurodegenerative diseases. Increase in dietary intake of docosahexaenoic acid offers the possibility of counterbalancing the harmful effects of high levels of arachidonic acid-derived pro-inflammatory eicosanoids.

Free radicals have been implicated and considered as associated risk factors for a variety of human disorders including neurodegenerative diseases, although it is not yet clear whether oxidative stress acts as a causative agent of neuronal degeneration. In human tissues, a condition of oxidative stress can be revealed through searching for specific biomarkers of oxidative damage to lipids, proteins and nucleic acids. Markers of oxidative damage to lipids include 4-hydroxynonenal, malondialdehyde, lipid hydroperoxides and isoprostanes, and thiobarbituric acid–reactive substances. Protein carbonyls and protein nitration are common markers of oxidative damage to proteins. The levels of 8-hydroxyguanine, or alternatively 8-hydroxy-2’- deoxyguanosine, can be measured in brain specimens, blood and urines, and are commonly used as a marker of oxidative DNA damage. Besides DNA a growing body of evidence indicates that also RNA undergoes oxidative damage. Studies have been performed also on markers of antioxidant defence such as oxidative modifications of plasma proteins, the activity of enzymes of the antioxidant defence (superoxide dismutase and catalase), the levels and the activity of proteins involved in the repair of oxidative DNA damage and the state of components of blood glutathione system. Significant biological changes related to a condition of oxidative stress have been found not only in brain tissues but also in biological fluids such as urine, peripheral blood or cerebrospinal fluid, and in peripheral tissues such as blood cells and fibroblasts of individuals affected by Alzheimer’s disease, Mild Cognitive Impairment, Parkinson's disease and Amyotrophic Lateral Sclerosis.

In addition to integral proteins, neural membranes are composed of glycerophospholipids, sphingolipids, and cholesterol, which provide membranes with structural integrity (suitable stability, fluidity, and permeability). Kainic acid (KA), an excitotoxin, markedly upregulates glycerophospholipids, sphingolipids, and cholesterol metabolism resulting not only in loss of essential lipids and inducing changes in neural membrane fluidity and permeability, but also in elevations in glycerophospholipid, sphingolipid, and cholesterol-derived lipid mediators. These processes result in depolarization, sustained increase in Ca2+ and stimulation of Ca2+-dependent enzymes including PLA2, PLC, nitric oxide synthase, calpains, and endonucleases. Sustained stimulation of these enzymes, generation, and interplay among glycerophospholipid-, sphingolipid-, and cholesterol-derived lipid mediators along with mitochondrial dysfunction, decrease in ATP levels, changes in redox status of neural cell may be responsible for neurodegeneration through apoptosis and necrosis in KA-mediated neurotoxicity. Other KAmediated neurochemical changes include synaptic reorganization associated with recapitulation of hippocampal development and synaptogenesis following KA-induced seizures.

Neurodegenerative diseases, particularly those associated with aging such as Alzheimer’s disease, represent a significant public health concern. The development of effective treatments is, however, hindered by the complex, multigenic nature of these diseases and by their poorly understood molecular pathophysiology. Mitochondria seem to play a primary role in neurodegeneration, due to the high energy demand of the brain. These organelles are the main producers of energy through the tricarboxylic acid cycle and host a high number of biochemical pathways including those involved in storage and maintenance of intracellular calcium levels, cellular homeostasis and survival pathways. However, mitochondria are a double edge sword. In the presence of certain oxidative stimuli, for instance, when oxygen demand exceeds supply (hypoxia), mitochondria can activate several death pathways. Indeed, hypoxia has been implicated in several neurodegenerative diseases including Alzheimer’s disease. Current knowledge supports the idea that during hypoxic events mitochondrial complex III produces high levels of reactive oxygen species (ROS), which play a key role in the regulation of the transcription factor hypoxia inducible factor 1 that triggers several death effectors. In this chapter we will discuss the involvement of mitochondria in AD putting focus on the mitochondrial pathways activated by hypoxia, which could eventually lead neurodegenerative events.

Dopamine-mediated neurotoxicity has been speculated as a potential contribution to the pathogenesis of Parkinson disease (PD), including a diffuse protein aggregation pathology but relatively selective death of dopaminergic neurons in the substantia nigra. The dopamine-mediated oxidative stress, produced by dopamine oxidation resulting in reactive oxygen species (ROS) and reactive dopamine quinones, is hypothesized to be the key event in the specific cell death of dopaminergic neurons in the pathogenesis of sporadic PD and neurotoxininduced parkinsonism. The cytotoxity in dopaminergic neurons occurs primarily due to the generation of highly reactive cyclized O-quinones, which damage mitochondia by opening the mitochondrial permeability transition pore, leading to cell death. These toxic quinones conjugate with several key PD pathogenic molecules, such as tyrosine hydroxylase, a-synuclein and parkin, forming a complex of protein-bound-quinone (quinoprotein), consequently inhibiting enzyme/protein function. Furthermore, cyclized dopamine quinones also inhibit proteasome activity, resulting in protein aggregation that may facilitate Lewy body formation in PD. Dopamine quinone formation is also closely linked to other representative hypotheses for PD. However, reductants such as glutathione (GSH), ascorbic acid (AA), and superoxide dismutase (SOD) may protect dopaminergic neurons from dopamineinduced toxicity or by various other biochemical insults associated with PD. The chaperone heat-shock protein 70 (HSP70) reduces protein misfolding and aggregation in cells, implicating its protective role against a variety of insults including oxidative stress. There are several pathogenic mechanisms possibly involved with death of dopaminergic neurons, but this overview focuses on dopamine-mediated oxidative stress that may contribute to selective neurodegeneration of dopaminergic neurons in PD.

The complement system is a powerful and vital component of the innate immune system that plays a dual role in neurodegenerative diseases. When present at an optimum level in the normal brain, the complement system plays a neuroprotective role as it is involved in a number of processes like clearance of apoptotic cells, opsonisation of pathogens,etc. However factors such as oxidative stress and age can modify this protective ability can lead to chronic inflammation resulting in neurodegeneration. In a diseased brain, aggregated polypeptides can potentially present their different charge patterns to C1q, which is a vital charge pattern recognition molecule of the complement system. Consequently activation of complement leads to microglial activation which in turn leads to defective clearance of the aggregated polypeptides by macrophages leading to chronic inflammation, especially in age-related neurodegenerative disease (e.g., Alzheimer’s disease). The current article aims at discussing the role of the complement system (especially C1q) and its consequences in initiation/progression in neurodegenerative diseases such as amyloid-associated dementias (Alzheimer’s disease, Down’s syndrome), non-amyloid associated dementia (Familial dementia, Huntington’s disease, Parkinson’s disease) and pathogen-induced dementia (prion diseases). Evidences point towards the existence of an over-activated complement system in a diseased brain can directly or indirectly lead to neuroinflammation which subsequently leads to neurodegeneration, the effects of which are manifested through the various clinical signs and symptoms. As C1q is the initiation molecule of the classical pathway, C1q-inhibitors that down regulate the complement cascade without negatively affecting the protective functions of complement can pave way for potential future immunotherapeutic approaches.

Platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero- 3-phosphocholine) displays a variety of biological activities in the nervous system. It has been suggested that PAF plays important roles in neuronal physiological function via activation of its specific membranes receptors. Under certain pathological conditions, PAF acts as a potent mediator of leukocyte functions, platelet aggregation, pro-inflammatory signaling and others. Therefore, PAF has been implicated in the pathophysiology of neuronal diseases such as ischemic stroke, hemorrhagic stroke, chronic subdural hematoma after head injury, brain tumor and associated brain edema, dementia due to neurodegenerative diseases such as Parkinson’s disease and Prion diseases, and HIV-1-associated dementia. PAF is synthesized in platelet, monocytes/macrophage, neutrophils and endothelial cells in response to physiological and pathological stimuli through the de novo and remodeling pathways from cellular membrane phospholipids. PAF is thought to be a pro-inflammatory and pro-thrombotic mediator and also causes direct damage to neuronal cells. At least three types of platelet-activating factor acetylhydrolase (PAF-AH) have been identified in mammals, i.e., intracellular type I and II, and a plasma type. The type I PAF-AH hydrolyzes the sn-2 ester bound in PAF-like phospholipids with a marked preference for very short acyl chains, typically acetyl bound. On the other hand, the type II PAF-AH has its substrate specificity similar to the plasma PAF-AH. Both PAFAHs hydrolyze phospholipids with short to medium length sn-2 acyl chains including truncated ones derived from oxidative cleavage of long chain polyunsaturated fatty acyl groups. With respect to atherosclerosis it is not fully understood whether this enzyme plays an anti-atherogenic role or pro-atherogenic role. In this review, the roles of PAF, its signaling and related metabolism including PAH-AHs in a variety of pathological conditions in the central nervous system are discussed.

Role of oxidative stress and oxidative cellular deficits have been considered for a long time in the etiopathogenesis as well as in course and treatment outcome of schizophrenia. A large number of reviews and monograms have been published primarily on altered levels of indices of oxidative stress and oxidative cell injury in mostly chronic medicated patients and few in drug naïve early psychotic patients. However, since schizophrenia is now generally considered to have neurodevelopmental deficits that most likely start in utero it is important to have a specific mechanism that can trigger the oxidative stress at the critical developmental time. This lack of information has limited the success in developing effective treatment strategies in amelioration of oxidative stress-mediated cellular damage and improved neurodevelopment and consequent clinical outcome. Furthermore, chronic use of the major antipsychotics worsens the oxidative cellular damage and contributes to the poor clinical outcome by increased negative symptoms and the cognitive and motor deficits. We have here presented a hypothesis that states as, the altered metabolism of key maternal nutrients such as folic acid and B12 and omega-3 fatty acids synergistically will trigger the oxidative stress which will alter the early foetal neurodevelopment and alter later in life the cognitive deficits and psychosis key behavioural symptomatology of schizophrenia. The altered metabolism of these maternal nutrients will contribute to the altered one carbon metabolism leading to increased homocysteine and reduced expression of antioxidant enzyme genes and neurotrophic factors by altered chromatin methylation (epigenesis) that contribute to increased oxidative stress. This novel mechanism will also explain the role of a large number of genetic (e.g., altered expression of antioxidant enzymes and neurotrophins) and environmental (e.g., maternal use of alcohol and drug abuse, smoking, under- and mal-nutrition, and socioeconomic stressors) risk factors reported for etiopathogenesis and course of schizophrenia since all of these risk factors trigger the oxidative stress and cellular oxidative damage such as reported in schizophrenia with and without treatment with antipsychotics. Finally, the mechanism based on altered micronutrients may provide effective specific nutritional supplementation strategies for prevention of oxidative stress-mediated pathologies associated with onset of schizophrenia as well as its improved clinical outcome by conventional treatments.

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are of benefit in Alzheimer’s disease by virtue of their anti-inflammatory actions, ability to modulate neural function, including neurotransmission, membrane fluidity, ion channel, enzyme regulation and gene expression. EPA, DHA, and ω-6 arachidonic acid (AA) form precursors to anti-inflammatory compounds: lipoxins, resolvins, protectins and maresins that suppress leukocyte migration and activation, inhibit NF-κB activation, production of pro-inflammatory cytokines tumor necrosis factor-α and interleukin-6, free radical generation, and enhance endothelial nitric oxide generation and augment the healing process. In animal models, the protective action of EPA and DHA against Alzheimer’s disease correlated with increased formation of lipoxins, resolvins, protectins and maresins. EPA, DHA and AA stimulate neurite outgrowth by activating syntaxin 3 that is specifically involved in fast calcium-triggered exocytosis of neurotransmitters. SNAP25 (synaptosomal-associated protein of 25 kDa), a syntaxin partner implicated in neurite outgrowth, interacted with syntaxin 3 only in the presence of AA that allowed the formation of the binary syntaxin 3-SNAP 25 complex. AA stimulated syntaxin 3 to form the ternary SNARE complex (soluble N-ethylmaleimide-sensitive factor attachment protein receptor), which is needed for the fusion of plasmalemmal precursor vesicles into the cell surface membrane that leads to membrane fusion that facilitates neurite outgrowth. These results imply that EPA, DHA, and AA when given in optimal amounts are of benefit in the prevention and treatment of Alzheimer’s disease. PUFAs enhance the concentrations of neurotrophic factors in the brain that may provide additional protection to neurons. Thus, PUFAs by themselves or their stable synthetic analogues could be of benefit in Alzheimer’s disease and other neurodegenerative diseases.

Neuronal cell death in neurodegenerative and neurotraumatic diseases is accompanied by excitotoxicity, oxidative stress, and inflammation. These processes lead to the generation of lipid mediators. The intensity of cross talk among lipid mediators dictates the rate of neurodegeneration in above neurological diseases. In neurodegenerative diseases cell death occurs slowly (months to years), where as in neurotraumatic diseases, neurodegeneration occurs rapidly (minutes to hours) through necrosis at the core of injury, whereas in penumbral region neurons undergo delayed neurodegeneration through apoptosis. Presence of lipid mediators in biological fluids can be used for the diagnosis of above neurological disorders.

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