Recently, extracellular vesicles (EVs), such as exosomes, have been proposed to play an influential role in the cell-to-cell spread of neurodegenerative diseases, including the intercellular transmission of α-synuclein (α-syn). However, the regulation of EV biogenesis and its relation to Parkinson’s disease (PD) is only partially understood. The generation of EVs through the ESCRT-independent pathway depends on the hydrolysis of sphingomyelin by neutral sphingomyelinase 2 (nSMase2) to produce ceramide, which causes the membrane of endosomal multivesicular bodies to bud inward. nSMase2 is sensitive to oxidative stress, a common process in PD brains; however, little is known about the role of sphingomyelin metabolism in the pathogenesis of PD. This is the first study to show that inhibiting nSMase2 decreases the transfer of oligomeric aggregates of α-syn between neuron-like cells. Furthermore, it reduced the accumulation and aggregation of high-molecular-weight α-syn. Hypoxia, as a model of oxidative stress, reduced the levels of nSMase2, but not its enzymatic activity, and significantly altered the lipid composition of cells without affecting EV abundance or the transfer of α-syn. These data show that altering sphingolipids can mitigate the spread of α-syn, even under hypoxic conditions, potentially suppressing PD progression.

Parkinson's disease (PD) and other synucleinopathies are characterized by accumulation of misfolded aggregates of α-synuclein (α-syn). The normal function of α-syn is still under investigation, but it has been generally linked to synaptic plasticity, neurotransmitter release and the maintenance of the synaptic pool. α-Syn localizes at synaptic terminals where it can bind to synaptic vesicles as well as to other cellular membranes. It has become clear that these interactions have an impact on both α-syn functional role and its propensity to aggregate. In this study, we investigated the aggregation process of α-syn covalently modified with 4-hydroxy-2-nonenal (HNE). HNE is a product of lipid peroxidation and has been implicated in the pathogenesis of different neurodegenerative diseases by modifying the kinetics of soluble toxic oligomers. Although HNE-modified α-syn has been reported to assemble into stable oligomers, we found that slightly acidic conditions promoted further protein aggregation. Lipid vesicles delayed the aggregation process in a concentration-dependent manner, an effect that was observed only when they were added at the beginning of the aggregation process. Co-aggregation of lipid vesicles with HNE-modified α-syn also induced cytotoxic effects on differentiated SHSY-5Y cells. Under conditions in which the aggregation process was delayed cell viability was reduced. By exploring the behavior and potential cytotoxic effects of HNE-α-syn under acidic conditions in relation to protein-lipid interactions our study gives a framework to examine a possible pathway leading from a physiological setting to the pathological outcome of PD.

The gradual deterioration of cognitive functions in Alzheimer’s disease is paralleled by a hierarchical progression of amyloid-beta and tau brain pathology. Recent findings indicate that toxic oligomers of amyloid-beta may cause propagation of pathology in a prion-like manner, although the underlying mechanisms are incompletely understood. Here we show that small extracellular vesicles, exosomes, from Alzheimer patients’ brains contain increased levels of amyloid-beta oligomers and can act as vehicles for the neuron-to-neuron transfer of such toxic species in recipient neurons in culture. Moreover, blocking the formation, secretion or uptake of exosomes was found to reduce both the spread of oligomers and the related toxicity. Taken together, our results imply that exosomes are centrally involved in Alzheimer’s disease and that they could serve as targets for development of new diagnostic and therapeutic principles.

The aged brain may be used as a tool to investigate altered metabolism of amyloid beta protein (Aβ42) that may have implications in the pathogenesis of Alzheimer's disease (AD). In the present study, we have observed a striking increase in the amyloid precursor protein (APP) level in the brain cortex of aged rats (22-24 months) along with a mild but statistically significant increase in the level of APP mRNA. Moreover, the activity of β secretase is elevated (nearly 55%) and that of neprilysin diminished (48%) in brain cortex of aged rats compared to that in young rats (4-6 months). All these changes lead to a markedly increased accumulation of Aβ42 in brain cortical tissue of aged rats. Long-term dietary supplementation of rats with a combination of N-acetylcysteine, α-lipoic and α-tocopherol from 18 months onwards daily till the sacrifice of the animals by 22-24 months, attenuates the age-related alterations in amyloid beta metabolism. In separate experiments, a significant impairment of spatial learning and memory has been observed in aged rats, and the phenomenon is remarkably prevented by the dietary supplementation of the aged animals by the same combination of N-acetylcysteine, α-lipoic acid and α-tocopherol. The results call for further explorations of this combination in suitable animal models in ameliorating AD related brain deficits.

The metabolic turnover of sphingolipids produces several signaling molecules that profoundly affect the proliferation, differentiation and death of cells. In particular, an enormous body of information is available that defines the varied role of ceramide and sphingosine-1-phosphate in cell death and survival. This review specifically examines the role of ceramide and sphingosine-1- phosphate in triggering neuronal death in Alzheimer's disease by analyzing the data from post-mortem studies and experimental research. There is compelling evidence that ceramide plays a key role in the neurodegeneration and amyloidogenesis occurring in the brain in Alzheimer's disease. Further, it appears that ceramide and amyloid beta protein orchestrate an attack on mitochondria to set in the pathways of cell death. However, the complexity of metabolic and signaling pathways of sphingolipid derivatives precludes an immediate identification of effective drug targets for the therapy of Alzheimer's disease.

Oxidative stress and inflammatory response are important elements of Alzheimer's disease (AD) pathogenesis, but the role of redox signaling cascade and its cross-talk with inflammatory mediators have not been elucidated in details in this disorder. The review summarizes the facts about redox-signaling cascade in the cells operating through an array of kinases, phosphatases and transcription factors and their downstream components. The biology of NF-κB and its activation by reactive oxygen species (ROS) and proinflammatory cytokines in the pathogenesis of AD have been specially highlighted citing evidence both from post-mortem studies in AD brain and experimental research in animal or cell-based models of AD. The possibility of identifying new disease-modifying drugs for AD targeting NF-κBsignaling cascade has been discussed in the end.

The elucidation of the intriguing relationship between oxidative stress and Alzheimer's disease is crucial to understand the pathogenesis of the disease as also to design a suitable drug trial with antioxidants against this condition. We begin by reviewing the basic facts about Alzheimer's disease and the chemistry and biology of oxygen free radicals with particular reference to the cellular adaptive response through redox-signalling pathways. The post-mortem evidence of oxidative damage in the brain of Alzheimer's disease patients is overwhelming which is also supported by the similar changes in transgenic mice models of this disease. However, the causal relationship of oxidative stress with amyloid beta pathology or the genesis of Alzheimer's disease is not clear. Considering the available evidence the review suggests that the oxidative stress could be an early event in the disease process and may trigger various adaptive responses such as the alterations of amyloid beta metabolism and the activation of stress responsive kinases which can subsequently lead to neuronal degeneration and AD pathology. Further, we have presented a large body of evidence from various studies to highlight the beneficial effects of antioxidants against amyloid beta toxicity or AD pathology in animal or cell based models of AD. The failure of clinical trials with antioxidants against AD has been mentioned and the possible causes of such failures have been analysed.

In contrast to many studies showing the pro-oxidative nature of amyloid peptide, this work shows that aggregated Aβ42 peptide in varying concentrations (2–20μM) in cell-free systems inhibits the formation of hydroxyl radicals and H2O2 from a mixture of iron (20μM FeSO4) and ascorbate (2mM) as measured by benzoate hydroxylation assay and coumarin carboxylic acid assay. Aggregated Aβ42 in similar concentrations further prevents protein and lipid oxidation in isolated rat brain mitochondria incubated alone or with FeSO4 and ascorbate. Moreover, mitochondria exposed to FeSO4 and ascorbate show enhanced formation of reactive oxygen species and this phenomenon is also abolished by aggregated Aβ42. It is suggested that the antioxidant property of Aβ42 in various systems is mediated by metal chelation and it is nearly as potent as a typical metal chelator, such as diethylenetriaminepentaacetic acid, in preventing oxidative damage. However, aggregated Aβ42 causes mitochondrial functional impairment in the form of membrane depolarization and a loss of phosphorylation capacity without involving reactive oxygen species in the process. Thus, the present results suggest that the amyloid peptide exhibits a protective antioxidant role in biological systems, but also has toxic actions independent of oxidative stress.

Various serum or CSF biomarkers of Alzheimer's disease (AD) have been identified in a large number of case-control studies, but neither their utility in the differential diagnosis of AD, nor their role in the pathogenesis of sporadic AD has been clearly understood. This study attempts to identify a panel of serum biomarkers for AD to examine their usefulness in distinguishing AD from vascular dementia (VAD) and to test some of these biomarkers in a cell based model for their plausible role in affecting amyloid beta peptide metabolism. Serum levels of Hcy, DHEA-S and Lp(a) were measured by ELISA using commercial kits in patients of AD (n = 40), VAD (n = 40) and age-matched controls (n = 50). In another set of case-control observations, fasting serum levels of adiponectin, leptin and insulin were measured in 35 AD and 35 age-matched control subjects.In the experimental part SHSY5Y cells are exposed to exogenously added insulin, homocysteine and adiponectin in varying concentrations for a period of 48 h and the effects on intracellular APP and amyloid beta peptide content are monitored by immunoblotting and ELISA. Our results show that serum levels of Hcy, adiponectin and insulin are significantly elevated while those of DHEA-S and leptin are markedly decreased in AD subjects, but no significant difference in Lp(a) concentration in serum is found between control and AD groups. Further, the serum levels of Hcy, DHEA-S and Lp(a) can distinguish clearly the AD group from the VAD subjects.At the experimental level, insulin but not Hcy causes a marked increase in both the APP and Aβ42 content along with alterations in β secretase and neprilysin activities in SHSY5Y cells. The effects of adiponectin on SHSY5Y cells are under study. Serum parameters like Hcy, DHEA-S, adiponectin, leptin , insulin are biomarkers useful in the differential diagnosis of AD and insulin in particular also promotes AD pathogenesis in a cell based model.

The study has demonstrated that dopamine induces membrane depolarization and a loss of phosphorylation capacity in dose-dependent manner in isolated rat brain mitochondria during extended in vitro incubation and the phenomena are not prevented by oxyradical scavengers or metal chelators. Dopamine effects on brain mitochondria are, however, markedly prevented by reduced glutathione and N-acetyl cysteine and promoted by tyrosinase present in the incubation medium. The results imply that quinone oxidation products of dopamine are involved in mitochondrial damage under this condition. When PC12 cells are exposed to dopamine in varying concentrations (100-400μM) for up to 24h, a pronounced impairment of mitochondrial bio-energetic functions at several levels is observed along with a significant (nearly 40%) loss of cell viability with features of apoptotic nuclear changes and increased activities of caspase 3 and caspase 9 and all these effects of dopamine are remarkably prevented by N-acetyl cysteine. N-acetyl cysteine also blocks nearly completely the dopamine induced increase in reactive oxygen species production and the formation of quinoprotein adducts in mitochondrial fraction within PC12 cells and also the accumulation of quinone products in the culture medium. Clorgyline, an inhibitor of MAO-A, markedly decreases the formation of reactive oxygen species in PC12 cells upon dopamine exposure but has only mild protective actions against quinoprotein adduct formation, mitochondrial dysfunctions, cell death and caspase activation induced by dopamine. The results have indicated that quinone oxidation products and not reactive oxygen species are primarily involved in cytotoxic effects of dopamine and the mitochondrial impairment plays a central role in the latter process. The data have clear implications in the pathogenesis of Parkinson's disease.

Sporadic Alzheimer's disease (AD) is multifactorial in origin, but the key elements of the pathogenic process are mitochondrial dysfunction, oxidative stress, inflammatory response, protein aggregation. The molecular links among these potential pathogenic mechanisms are not fully elucidated, but oxidative stress because of its ability to alter cell signaling, gene expression, membrane properties, enzyme and receptor functions is most likely to influence the cross-talk among the probable pathogenic processes of AD. In an ongoing study in the lab we have observed that the age-related oxidative stress alters amyloid peptide metabolism in rat brain which is prevented by long-term dietary supplementation. This study has attempted to explore the role of oxidative stress in triggering and propagating amyloid peptide pathology in a cell based model. PC 12 cells have been subjected to an oxidative insult by exposure to H202 (100-400 μM) or t-butylhydroperoxide (100-400 μM) in the presence or absence of radical scavengers and antioxidants. The altered metabolism of amyloid peptide has been monitored by measuring the expression levels of APP, the intracellular and extra cellular level of Aß-42, the activities/ levels of ß secretase, δ secretase as well as the degrading enzymes of Aß-42 namely neprilysin and insulin degrading enzyme (IDE). Oxidative damage markers, protein aggregation and cell death have also been measured in this experimental model. Significant alterations in APP expression and intracellular content of Aß-42 have been observed along with accumulation of oxidative damage markers and conspicuous cell death in PC 12 cells exposed to oxidative stress. Conclusions: When extrapolated to in vivo situation, our results imply that oxidative stress present in aged brain can be a major factor in triggering and promoting altered APP metabolism leading to the genesis of sporadic AD. The study shows oxidative stress mediated Amyloid precursor protein metabolism alteration as also accumulation of Aß-42 in cell. This study has clear implications in understanding the common mechanisms of brain aging and AD.

The entangled relationship of brain aging, mitochondrial dysfunction, and amyloid-β peptide (Aβ₄₂) toxicity occupies the center stage in the pathogenesis of Alzheimer's disease (AD). The present study examines some of the toxic effects of Aβ₄₂ on brain mitochondria and provides evidence that aged brain mitochondria are significantly more vulnerable to Aβ₄₂ toxicity. In particular, the study has shown that the aggregated, but not the monomeric, form of Aβ₄₂ in varying concentrations (10-40 μM) during in vitro incubation causes a loss of mitochondrial membrane potential, a decrease in phosphorylation capacity and ATP synthesis, and the release of cytochrome c from the mitochondria but without any noticeable change in the activities of respiratory chain complexes. Such effects of Aβ₄₂ are strikingly more conspicuous on aged rat (22-24 months) brain mitochondria compared to that on brain mitochondria of young rats (4-6 months). More interestingly is the observation that in contrast to young rat brain mitochondria, a significantly higher level of Aβ₄₂ remains associated with aged brain mitochondria under basal incubation condition as well as after exposure to exogenously added peptide. Extrapolated to an in vivo scenario, the results have clear implications in AD pathogenesis and also partly explain why brain aging is a dominant risk factor for this disease condition.

Mitochondrial dysfunction and oxidative stress are two interdependent and reinforcing damage mechanisms that play a central role in brain aging. Oxidative stress initiated and propagated by active oxyradicals and various other free radicals in the presence of catalytic metal ions not only can damage the phospholipid, protein and DNA molecules within the cell but can also modulate cell signalling pathways and gene expression pattern and all these processes may be of critical importance in the aging of brain. The present article describes the mechanism of formation of reactive oxyradicals within mitochondria and then explains how these can initiate mitochondrial biogenesis program and activate various transcriptional factors in the cytosol to boost up the antioxidative capacity of the mitochondria and the cell. However, a high level of oxidative stress finally inflicts critical damage to the oxidative phosphorylation machinery and mitochondrial DNA (mtDNA). The latter part of the article is a catalogue showing the accumulating evidence in favour of oxidative inactivation of mitochondrial functions in aged brain and the detailed reports of various studies with antioxidant supplementation claiming variable success in preventing the age-related brain mitochondrial decay and cognitive decline. The antioxidant supplementation approach may be of potential help in the management of neurodegenerative diseases like Alzheimer's disease. The newly developed mitochondria-targeted antioxidants have brought a new direction to experimental studies related to oxidative damage and they may provide potential drugs in near future for a variety of diseases or degenerative conditions including brain aging and neurodegenerative disorders.

The study has shown that in aged (22-24 months) rat brains an elevation of homocysteine level (42%) and a decrease in dehydroepiandrosterone sulphate (DHEA-S) content (32%) occur compared to those in the brains of young rats (4-6 months). Such changes in the brain levels of homocysteine and DHEA-S in aged rats are prevented, when the diet daily of the rats is supplemented with a combination of antioxidants (N-acetyl cysteine 50 mg, alpha-lipoic acid 3 mg and alpha-tocopherol 1.5 mg - each per 100 g of body weight) starting from 18 months until these are sacrificed between 22 and 24 months. The brain content of reduced glutathione is also decreased in aged rats as compared to that in young ones and the phenomenon can again be prevented completely by the same regimen of antioxidant supplementation. The changes in the levels of homocysteine and DHEA-S in aged rat brain have been related to associated glutathione depletion and oxidative stress and the implications of the results highlighted in the pathogenesis of Alzheimer's disease.

The etiopathogenesis of sporadic Alzheimer's Disease (AD) is uncertain, but altered APP metabolism with associated toxicity of amyloid beta protein Aβ (42/40) probably plays a central role in the onset and progression of this neurodegenerative disorder which causes devastating failure of memory and cognition in elderly people. Aging is considered as the most dominant risk factor for AD, although the molecular and cellular links are not clear. The present study addresses several questions related to AD and brain aging in the context of APP metabolism, Aβ 42 toxicity and brain content of homocysteine which is considered as an important risk factor for AD from epidemiological and experimental studies. The levels of Aβ 42 and APP were measured by ELISA and semiquantitative immunoblotting techniques in brain of young (4-6 months) and aged (22-24 months) rats. Homocysteine level was measured in young and aged rat brain using an ELISA based commercial kit. Brain mitochondria from young and aged rats were isolated and incubated with purified Aβ 42 (aggregated and monomeric forms) at 370 for 2h followed by the measurement of mitochondrial functional parameters such as transmembrane potential, phosphate utilization capacity, ATP synthesis and activities of respiratory chain complexes. Significant alterations in the levels of homocysteine, APP and Aβ 42 were noted in young and aged rat brain. During in vitro incubation Aβ 42 caused a dose dependent decrease of brain mitochondrial transmembrane potential, phosphate utilization and ATP synthesis without affecting the activity of respiratory chain complexes. The differential effect of Aβ 42 on brain mitochondria isolated from young and aged rats are being investigated now. The study shows age related accumulation of Aβ 42 and homocysteine in rat brain as also the toxic effects of Aβ 42 on brain mitochondria during in vitro incubation. The results have clear implications in linking the process of brain aging with AD pathogenesis.

This study demonstrates that in vitro incubation of isolated rat brain mitochondria with recombinant human α‐synuclein leads to dose‐dependent loss of mitochondrial transmembrane potential and phosphorylation capacity. However, α‐synuclein does not seem to have any significant effect on the activities of respiratory chain complexes under similar conditions of incubation suggesting that the former may impair mitochondrial bioenergetics by direct effect on mitochondrial membranes. Moreover, the recombinant wild type α‐synuclein and different mutant forms (A30P, A53T and E46K) have essentially similar effects on rat brain isolated mitochondria. The results are significant in view of the fact that α‐synucleinopathy is involved in the pathogenesis of Parkinson's disease.