A strategic tool to improve the study of molecular determinants of Alzheimer's disease: The role of glyceraldehyde.

Alzheimer's disease (AD) is the most prevalent form of dementia and is characterized by progressive neurodegeneration leading to severe cognitive, memory, and behavioral impairments. The onset of AD involves a complex interplay among various factors, including age, genetics, chronic inflammation, and impaired energy metabolism. Despite significant efforts, there are currently no effective therapies capable of modifying the course of AD, likely owing to an excessive focus on the amyloid hypothesis and a limited consideration of other intracellular pathways. In the present review, we emphasize the emerging concept of AD as a metabolic disease, where alterations in energy metabolism play a critical role in its development and progression. Notably, glucose metabolism impairment is associated with mitochondrial dysfunction, oxidative stress, Ca2+ dyshomeostasis, and protein misfolding, forming interconnected processes that perpetuate a detrimental self-feeding loop sustaining AD progression. Advanced glycation end products (AGEs), neurotoxic compounds that accumulate in AD, are considered an important consequence of glucose metabolism disruption, and glyceraldehyde (GA), a glycolytic intermediate, is a key contributor to AGEs formation in both neurons and astrocytes. Exploring the impact of GA-induced glucose metabolism impairment opens up exciting possibilities for creating an easy-to-handle in vitro model that recapitulates the early stage of the disease. This model holds great potential for advancing the development of novel therapeutics targeting various intracellular pathways implicated in AD pathogenesis. In conclusion, looking beyond the conventional amyloid hypothesis could lead researchers to discover promising targets for intervention, offering the possibility of addressing the existing medical gaps in AD treatment.

Exploring the Role of NCX1 and NCX3 in an In Vitro Model of Metabolism Impairment: Potential Neuroprotective Targets for Alzheimer's Disease.

Alzheimer's disease (AD) is a widespread neurodegenerative disorder, affecting a large number of elderly individuals worldwide. Mitochondrial dysfunction, metabolic alterations, and oxidative stress are regarded as cooperating drivers of the progression of AD. In particular, metabolic impairment amplifies the production of reactive oxygen species (ROS), resulting in detrimental alterations to intracellular Ca2+ regulatory processes. The Na+/Ca2+ exchanger (NCX) proteins are key pathophysiological determinants of Ca2+ and Na+ homeostasis, operating at both the plasma membrane and mitochondria levels. Our study aimed to explore the role of NCX1 and NCX3 in retinoic acid (RA) differentiated SH-SY5Y cells treated with glyceraldehyde (GA), to induce impairment of the default glucose metabolism that typically precedes Aß deposition or Tau protein phosphorylation in AD. By using an RNA interference-mediated approach to silence either NCX1 or NCX3 expression, we found that, in GA-treated cells, the knocking-down of NCX3 ameliorated cell viability, increased the intracellular ATP production, and reduced the oxidative damage. Remarkably, NCX3 silencing also prevented the enhancement of Aß and pTau levels and normalized the GA-induced decrease in NCX reverse-mode activity. By contrast, the knocking-down of NCX1 was totally ineffective in preventing GA-induced cytotoxicity except for the increase in ATP synthesis. These findings indicate that NCX3 and NCX1 may differently influence the evolution of AD pathology fostered by glucose metabolic dysfunction, thus providing a potential target for preventing AD.

miRNAs, Mesenchymal Stromal Cells and Major Neoplastic and Inflammatory Skin Diseases: A Page Being Written: A Systematic Review.

Micro RNAs (miRNAs) are a type of non-coding RNA (ncRNA) and typically interact with specific target mRNAs through complementary base pairing, affecting their translation and/or stability. MiRNAs regulate nearly all cellular functions, including the cell fate of mesenchymal stromal cells (MSCs). It is now accepted that various pathologies arise at the stem level, and, in this scenario, the role played by miRNAs in the fate of MSCs becomes of primary concern. Here we have considered the existing literature in the field of miRNAs, MSCs and skin diseases, classified as inflammatory (such as psoriasis and atopic dermatitis-AD) and neoplastic (melanoma and non-melanoma-skin-cancer including squamous cell and basal cell carcinoma) diseases. In this scoping review article, the evidence recovered indicates that this topic has attracted attention, but it is still a matter of opinion. A protocol for this review was registered in PROSPERO with the registration number "CRD42023420245". According to the different skin disorders and to the specific cellular mechanisms considered (cancer stem cells, extracellular vesicles, inflammation), miRNAs may play a pro- or anti-inflammatory, as well as a tumor suppressive, or supporting, role, indicating a complex regulation of their function. It is evident that the mode of action of miRNAs is more than a switch on-off, and all the observed effects of their dysregulated expression must be checked in a detailed analysis of the targeted proteins. The involvement of miRNAs has been studied mainly for squamous cell carcinoma and melanoma, and much less in psoriasis and AD; different mechanisms have been considered, such as miRNAs included in extracellular vesicles derived both from MSCs or tumor cells, miRNAs involved in cancer stem cells formation, up to miRNAs as candidates to be new therapeutic tools.

A new K+channel-independent mechanism is involved in the antioxidant effect of XE-991 in an in vitro model of glucose metabolism impairment: implications for Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder that represents the first cause of dementia. Although there has been significant progress in AD research, the actual mechanisms underlying this pathology remain largely unknown. There is increasing evidence that oxidative stress, metabolic alterations, and mitochondrial dysfunction are key players in the development and worsening of AD. As a result, in the past few years, remarkable attempts have been made to develop neuroprotective strategies against the impairment of mitochondrial dynamics and cell redox status. In the present study, we reveal a novel antioxidant K+ channel-independent effect of the M-current inhibitor XE-991 in SH-SY5Y cells differentiated with retinoic acid (RA) and primary rat cortical neurons exposed to the glycolysis inhibitor glyceraldehyde (GA). This experimental approach aimed to create a condition of hypometabolism accompanied by mitochondrial dysfunction and redox imbalance, as frequently observed in the beginning stage of the disease. We found that XE-991 exerted a neuroprotective action most likely through the resumption of superoxide dismutase (SOD) activity, which was significantly compromised during GA challenge. We also observed that the enhancement of SOD activity was accompanied by a sequence of positive effects; these included the reduction in basal Ca2+ levels within cytoplasmic and mitochondrial compartments, the decrease in mitochondrial reactive oxygen species (ROS) production, the modulation of AMPK/mTOR pathway, the recovery of ΔΨm collapse, the increase in the intracellular ATP content and the decrease in amyloid-ß (Aß) and hyperphosphorylated form of tau protein (pTau) levels. Collectively, our study reveals an off-target antioxidant effect of XE-991 and paves the way toward the further evaluation of new therapeutic uses of already existing molecules to accelerate the process of developing an effective therapy to counteract AD.

The Hidden Notes of Redox Balance in Neurodegenerative Diseases.

Reactive oxygen species (ROS) are versatile molecules that, even if produced in the background of many biological processes and responses, possess pleiotropic roles categorized in two interactive yet opposite domains. In particular, ROS can either function as signaling molecules that shape physiological cell functions, or act as deleterious end products of unbalanced redox reactions. Indeed, cellular redox status needs to be tightly regulated to ensure proper cellular functioning, and either excessive ROS accumulation or the dysfunction of antioxidant systems can perturb the redox homeostasis, leading to supraphysiological concentrations of ROS and potentially harmful outcomes. Therefore, whether ROS would act as signaling molecules or as detrimental factors strictly relies on a dynamic equilibrium between free radical production and scavenging resources. Of notice, the mammalian brain is particularly vulnerable to ROS-mediated toxicity, because it possesses relatively poor antioxidant defenses to cope with the redox burden imposed by the elevated oxygen consumption rate and metabolic activity. Many features of neurodegenerative diseases can in fact be traced back to causes of oxidative stress, which may influence both the onset and progression of brain demise. This review focuses on the description of the dual roles of ROS as double-edge sword in both physiological and pathological settings, with reference to Alzheimer's and Parkinson's diseases.

Control of Ca2+ and metabolic homeostasis by the Na+/Ca2+ exchangers (NCXs) in health and disease.

Spatial and temporal control of calcium (Ca2+) levels is essential for the background rhythms and responses of living cells to environmental stimuli. Whatever other regulators a given cellular activity may have, localized and wider scale Ca2+ events (sparks, transients, and waves) are hierarchical determinants of fundamental processes such as cell contraction, excitability, growth, metabolism and survival. Different cell types express specific channels, pumps and exchangers to efficiently generate and adapt Ca2+ patterns to cell requirements. The Na+/Ca2+ exchangers (NCXs) in particular contribute to Ca2+ homeostasis by buffering intracellular Ca2+ loads according to the electrochemical gradients of substrate ions - i.e., Ca2+ and sodium (Na+) - and under a dynamic control of redundant regulatory processes. An interesting feature of NCX emerges from the strict relationship that connects transporter activity with cell metabolism: on the one hand NCX operates under constant control of ATP-dependent regulatory processes, on the other hand the ion fluxes generated through NCX provide mechanistic support for the Na+-driven uptake of glutamate and Ca2+ influx to fuel mitochondrial respiration. Proof of concept evidence highlights therapeutic potential of preserving a timed and balanced NCX activity in a growing rate of diseases (including excitability, neurodegenerative, and proliferative disorders) because of an improved ability of stressed cells to safely maintain ion gradients and mitochondrial bioenergetics. Here, we will summarize and review recent works that have focused on the pathophysiological roles of NCXs in balancing the two-way relationship between Ca2+ signals and metabolism.

Taurine Stabilizing Effect on Lysozyme.

Taurine is an important organic osmolyte in mammalian cells, and it weakens inflammation and oxidative stress mediated injuries in some diseases. Recently, taurine has been demonstrated to play a therapeutic role against neurodegenerative disorders, although its parallel involvement in several biochemical mechanisms makes not clear taurine specific role in these diseases. Furthermore, the stabilizing effect of this molecule in terms of protein stability is known, but not deeply investigated. In this work we explore by Circular Dichroism the stabilizing impact of taurine in lysozyme thermal denaturation and its influence in lysozyme aggregation into amyloid fibrils. Taurine even at low concentration modifies protein-protein interactions in lysozyme native state, as revealed by Small Angle X-ray Scattering experiments, and alters the amyloid aggregation pattern without completely inhibiting it, as confirmed by UV/Vis spectroscopy with Congo Red and by Atomic Force Microscopy. Evaluation of the cytotoxicities of the amyloid fibrils grown in presence or in absence of taurine is investigated on SH-SY5Y neuroblastoma cells.

The Neuroprotective Effect of L-Carnitine against Glyceraldehyde-Induced Metabolic Impairment: Possible Implications in Alzheimer's Disease.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive cognitive regression and memory loss. Dysfunctions of both glucose metabolism and mitochondrial dynamics have been recognized as the main upstream events of the degenerative processes leading to AD. It has been recently found that correcting cell metabolism by providing alternative substrates can prevent neuronal injury by retaining mitochondrial function and reducing AD marker levels. Here, we induced an AD-like phenotype by using the glycolysis inhibitor glyceraldehyde (GA) and explored whether L-carnitine (4-N-trimethylamino-3-hydroxybutyric acid, LC) could mitigate neuronal damage, both in SH-SY5Y neuroblastoma cells and in rat primary cortical neurons. We have already reported that GA significantly modified AD marker levels; here we demonstrated that GA dramatically compromised cellular bioenergetic status, as revealed by glycolysis and oxygen consumption rate (OCR) evaluation. We found that LC ameliorated cell survival, improved OCR and ATP synthesis, prevented the loss of the mitochondrial membrane potential (Δψm) and reduced the formation of reactive oxygen species (ROS). Of note, the beneficial effect of LC did not rely on the glycolytic pathway rescue. Finally, we noticed that LC significantly reduced the increase in pTau levels induced by GA. Overall, these findings suggest that the use of LC can promote cell survival in the setting of the metabolic impairments commonly observed in AD. Our data suggest that LC may act by maintaining mitochondrial function and by reducing the pTau level.

Gateways for Glutamate Neuroprotection in Parkinson's Disease (PD): Essential Role of EAAT3 and NCX1 Revealed in an In Vitro Model of PD.

Increasing evidence suggests that metabolic alterations may be etiologically linked to neurodegenerative disorders such as Parkinson's disease (PD) and in particular empathizes the possibility of targeting mitochondrial dysfunctions to improve PD progression. Under different pathological conditions (i.e., cardiac and neuronal ischemia/reperfusion injury), we showed that supplementation of energetic substrates like glutamate exerts a protective role by preserving mitochondrial functions and enhancing ATP synthesis through a mechanism involving the Na+-dependent excitatory amino acid transporters (EAATs) and the Na+/Ca2+ exchanger (NCX). In this study, we investigated whether a similar approach aimed at promoting glutamate metabolism would be also beneficial against cell damage in an in vitro PD-like model. In retinoic acid (RA)-differentiated SH-SY5Y cells challenged with α-synuclein (α-syn) plus rotenone (Rot), glutamate significantly improved cell viability by increasing ATP levels, reducing oxidative damage and cytosolic and mitochondrial Ca2+ overload. Glutamate benefits were strikingly lost when either EAAT3 or NCX1 expression was knocked down by RNA silencing. Overall, our results open the possibility of targeting EAAT3/NCX1 functions to limit PD pathology by simultaneously favoring glutamate uptake and metabolic use in dopaminergic neurons.

NCX1 and EAAC1 transporters are involved in the protective action of glutamate in an in vitro Alzheimer's disease-like model.

Increasing evidence suggests that metabolic dysfunctions are at the roots of neurodegenerative disorders such as Alzheimer's disease (AD). In particular, defects in cerebral glucose metabolism, which have been often noted even before the occurrence of clinical symptoms and histopathological lesions, are now regarded as critical contributors to the pathogenesis of AD. Hence, the stimulation of energy metabolism, by enhancing the availability of specific metabolites, might be an alternative way to improve ATP synthesis and to positively affect AD progression. For instance, glutamate may serve as an intermediary metabolite for ATP synthesis through the tricarboxylic acid (TCA) cycle and the oxidative phosphorylation. We have recently shown that two transporters are critical for the anaplerotic use of glutamate: the Na+-dependent Excitatory Amino Acids Carrier 1 (EAAC1) and the Na+-Ca2+ exchanger 1 (NCX1). Therefore, in the present study, we established an AD-like phenotype by perturbing glucose metabolism in both primary rat cortical neurons and retinoic acid (RA)-differentiated SH-SY5Y cells, and we explored the potential of glutamate to halt cell damage by monitoring neurotoxicity, AD markers, ATP synthesis, cytosolic Ca2+ levels and EAAC1/NCX1 functional activities. We found that glutamate significantly increased ATP production and cell survival, reduced the increase of AD biomarkers (amyloid ß protein and the hyperphosphorylated form of tau protein), and recovered the increase of NCX reverse-mode activity. The RNA silencing of either EAAC1 or NCX1 caused the loss of the beneficial effects of glutamate, suggesting the requirement of a functional interplay between these transporters for glutamate-induced protection. Remarkably, our results indicate, as proof-of-principle, that facilitating the use of alternative fuels, like glutamate, may be an effective approach to overcome deficits in glucose utilization and significantly slow down neuronal degenerative process in AD.

Cracking the code of sodium/calcium exchanger (NCX) gating: Old and new complexities surfacing from the deep web of secondary regulations.

Cell membranes spatially define gradients that drive the complexity of biological signals. To guarantee movements and exchanges of solutes between compartments, membrane transporters negotiate the passages of ions and other important molecules through lipid bilayers. The Na+/Ca2+ exchangers (NCXs) in particular play central roles in balancing Na+ and Ca2+ fluxes across diverse proteolipid borders in all eukaryotic cells, influencing cellular functions and fate by multiple means. To prevent progression from balance to disease, redundant regulatory mechanisms cooperate at multiple levels (transcriptional, translational, and post-translational) and guarantee that the activities of NCXs are finely-tuned to cell homeostatic requirements. When this regulatory network is disturbed by pathological forces, cells may approach the end of life. In this review, we will discuss the main findings, controversies and open questions about regulatory mechanisms that control NCX functions in health and disease.

Mitochondrial localization of NCXs: Balancing calcium and energy homeostasis.

It is well established that mitochondria are the main source of ATP production within cells. However, mitochondria have other remarkable functions, serving as important modulators of cellular Ca2+ signaling, and it is now generally recognized that control over Ca2+ homeostasis is intrinsically interwoven with mitochondrial abilities to adjust and tune ATP production. In this review, we describe the mechanisms that mitochondria use to balance Ca2+ homeostasis maintenance and cell energy metabolism. In recent years, the knowledge on the molecular machinery mediating Ca2+ influx/efflux has been improved and, albeit still open to further investigations, several lines of evidence converge on the hypothesis that plasma membrane Na+/Ca2+ exchanger (NCX) isoforms are also expressed at the mitochondrial level, where they contribute to the Ca2+ and Na+ homeostasis maintenance. In particular, the connection between mitochondrial NCX activity and metabolic substrates utilization is further discussed here. We also briefly focus on the alterations of both mitochondrial Ca2+ handling and cellular bioenergetics in neurodegenerative diseases, such as Parkinson's and Alzheimer's disease.

NCX and EAAT transporters in ischemia: At the crossroad between glutamate metabolism and cell survival.

Energy metabolism impairment is a central event in the pathophysiology of ischemia. The limited availability of glucose and oxygen strongly affects mitochondrial activity, thus leading to ATP depletion. In this setting, the switch to alternative energy sources could ameliorate cells survival by enhancing ATP production, thus representing an attractive strategy for ischemic treatment. In this regard, some studies have recently re-evaluated the metabolic role of glutamate and its potential to promote cell survival under pathological conditions. In the present review, we discuss the ability of glutamate to exert an "energizing role" in cardiac and neuronal models of hypoxia/reoxygenation (H/R) injury, focusing on the Na+/Ca2+ exchanger (NCX) and the Na+-dependent excitatory amino acid transporters (EAATs) as key players in this metabolic pathway.

Excitatory Amino Acid Transporters (EAATs): Glutamate Transport and Beyond.

Na+-dependent excitatory amino acid transporters (EAATs) are the major transport mechanisms for extracellular glutamate removal in the central nervous system (CNS). The primary function assigned to EAATs is the maintenance of low extracellular glutamate levels, thus allowing glutamate to be used as a signaling molecule in the brain and to avoid excitotoxicity. However, glutamate has other recognized functions. For instance, it is a key anaplerotic substrate for the tricarboxylic acid (TCA) cycle, as it can be converted to α-ketoglutarate by transaminases or glutamate dehydrogenase. Furthermore, glutamate is a precursor of the main antioxidant glutathione, which plays a pivotal role in preventing oxidative cell death. Therefore, glutamate signaling/use is at the crossroad of multiple metabolic pathways and accordingly, it can influence a plethora of cell functions, both in health and disease. Here, we provide an overview of the main functions of glutamate and its transport systems, analyzing its role as a neurotransmitter and at the same time, the possible metabolic fates it can undergo in the intracellular milieu. Specifically, the metabolic role of glutamate and the molecular machinery proposed to metabolically support its transport will be further analyzed.

Selective inhibition of mitochondrial sodium-calcium exchanger protects striatal neurons from α-synuclein plus rotenone induced toxicity.

Progressive accumulation of α-synuclein (α-syn) and exposure to environmental toxins are risk factors that may both concur to Parkinson's disease (PD) pathogenesis. Electrophysiological recordings of field postsynaptic potentials (fEPSPs) and Ca2+ measures in striatal brain slices and differentiated SH-SY5Y cells showed that co-application of α-syn and the neurotoxic pesticide rotenone (Rot) induced Ca2+ dysregulation and alteration of both synaptic transmission and cell function. Interestingly, the presence of the mitochondrial NCX inhibitor CGP-37157 prevented these alterations. The specific involvement of the mitochondrial NCX was confirmed by the inability of the plasma membrane inhibitor SN-6 to counteract such phenomenon. Of note, using a siRNA approach, we found that NCX1 was the isoform specifically involved. These findings suggested that NCX1, operating on the mitochondrial membrane, may have a critical role in the maintenance of ionic Ca2+ homeostasis in PD and that its inhibition most likely exerts a protective effect in the toxicity induced by α-syn and Rot.

The dual face of glutamate: from a neurotoxin to a potential survival factor-metabolic implications in health and disease.

Glutamate is the major excitatory neurotransmitter in the central nervous system. Beyond this function, glutamate also plays a key role in intermediary metabolism in all organs and tissues, linking carbohydrate and amino acid metabolism via the tricarboxylic acid cycle. Under both physiological and pathological conditions, we have recently found that the ability of glutamate to fuel cell metabolism selectively relies on the activity of two main transporters: the sodium-calcium exchanger (NCX) and the sodium-dependent excitatory amino-acid transporters (EAATs). In ischemic settings, when glutamate is administered at the onset of the reoxygenation phase, the coordinate activity of EAAT and NCX allows glutamate to improve cell viability by stimulating ATP production. So far, this phenomenon has been observed in both cardiac and neuronal models. In this review, we focus on the most recent findings exploring the unusual activity of glutamate as a potential survival factor in different settings.

Glutamate as a potential "survival factor" in an in vitro model of neuronal hypoxia/reoxygenation injury: leading role of the Na+/Ca2+ exchanger.

In brain ischemia, reduction in oxygen and substrates affects mitochondrial respiratory chain and aerobic metabolism, culminating in ATP production impairment, ionic imbalance, and cell death. The restoration of blood flow and reoxygenation are frequently associated with exacerbation of tissue injury, giving rise to ischemia/reperfusion (I/R) injury. In this setting, the imbalance of brain bioenergetics induces important metabolic adaptations, including utilization of alternative energy sources, such as glutamate. Although glutamate has long been considered as a neurotoxin, it can also be used as intermediary metabolite for ATP synthesis, and both the Na+/Ca2+ exchanger (NCX) and the Na+-dependent excitatory amino-acid transporters (EAATs) are essential in this pathway. Here we analyzed the role of NCX in the potential of glutamate to improve metabolism and survival of neuronal cells subjected to hypoxia/reoxygenation (H/R). In SH-SY5Y neuroblastoma cells differentiated into a neuron-like state, H/R produced a significant cell damage, a decrease in ATP cellular content, and intracellular Ca2+ alterations. Exposure to glutamate at the onset of the reoxygenation phase attenuated H/R-induced cell damage and evoked a significant raise in intracellular ATP levels. Furthermore, we found that in H/R cells NCX reverse-mode activity was reduced, and that glutamate limited such reduction. All the effects induced by glutamate supplementation were lost when cells were transfected with small interfering RNA against NCX1 and EAAT3, suggesting the need of a specific functional interplay between these proteins for glutamate-induced protection. Collectively, our results revealed the potential beneficial effect of glutamate in an in vitro model of H/R injury and focused on the essential role exerted by NCX1. Although preliminary, these findings could be a starting point to further investigate in in vivo systems such protective effect in ischemic settings, shedding a new light on the classical view of glutamate as detrimental factor.

Essential role of the Na+-Ca2+ exchanger (NCX) in glutamate-enhanced cell survival in cardiac cells exposed to hypoxia/reoxygenation.

Myocardial ischemia culminates in ATP production impairment, ionic derangement and cell death. The provision of metabolic substrates during reperfusion significantly increases heart tolerance to ischemia by improving mitochondrial performance. Under normoxia, glutamate contributes to myocardial energy balance as substrate for anaplerotic reactions, and we demonstrated that the Na+/Ca2+ exchanger1 (NCX1) provides functional support for both glutamate uptake and use for ATP synthesis. Here we investigated the role of NCX1 in the potential of glutamate to improve energy metabolism and survival of cardiac cells subjected to hypoxia/reoxygenation (H/R). Specifically, in H9c2-NCX1 myoblasts, ATP levels, mitochondrial activities and cell survival were significantly compromised after H/R challenge. Glutamate supplementation at the onset of the reoxygenation phase significantly promoted viability, improved mitochondrial functions and normalized the H/R-induced increase of NCX1 reverse-mode activity. The benefits of glutamate were strikingly lost in H9c2-WT (lacking NCX1 expression), or in H9c2-NCX1 and rat cardiomyocytes treated with either NCX or Excitatory Amino Acid Transporters (EAATs) blockers, suggesting that a functional interplay between these transporters is critically required for glutamate-induced protection. Collectively, these results revealed for the first time the key role of NCX1 for the beneficial effects of glutamate against H/R-induced cell injury.