Neuronal A2A receptor exacerbates synapse loss and memory deficits in APP/PS1 mice.

Early pathological upregulation of adenosine A2A receptors (A2ARs), one of the caffeine targets, by neurons is thought to be involved in the development of synaptic and memory deficits in Alzheimer's disease (AD) but mechanisms remain ill-defined. To tackle this question, we promoted a neuronal upregulation of A2AR in the hippocampus of APP/PS1 mice developing AD-like amyloidogenesis. Our findings revealed that the early upregulation of A2AR in the presence of an ongoing amyloid pathology exacerbates memory impairments of APP/PS1 mice. These behavioural changes were not linked to major change in the development of amyloid pathology but rather associated with increased phosphorylated tau at neuritic plaques. Moreover, proteomic and transcriptomic analyses coupled with quantitative immunofluorescence studies indicated that neuronal upregulation of the receptor promoted both neuronal and non-neuronal autonomous alterations, i.e. enhanced neuroinflammatory response but also loss of excitatory synapses and impaired neuronal mitochondrial function, presumably accounting for the detrimental effect on memory. Overall, our results provide compelling evidence that neuronal A2AR dysfunction, as seen in the brain of patients, contributes to amyloid-related pathogenesis and underscores the potential of A2AR as a relevant therapeutic target for mitigating cognitive impairments in this neurodegenerative disorder.

Exacerbation of C1q dysregulation, synaptic loss and memory deficits in tau pathology linked to neuronal adenosine A2A receptor.

Accumulating data support the role of tau pathology in cognitive decline in ageing and Alzheimer's disease, but underlying mechanisms remain ill-defined. Interestingly, ageing and Alzheimer's disease have been associated with an abnormal upregulation of adenosine A2A receptor (A2AR), a fine tuner of synaptic plasticity. However, the link between A2AR signalling and tau pathology has remained largely unexplored. In the present study, we report for the first time a significant upregulation of A2AR in patients suffering from frontotemporal lobar degeneration with the MAPT P301L mutation. To model these alterations, we induced neuronal A2AR upregulation in a tauopathy mouse model (THY-Tau22) using a new conditional strain allowing forebrain overexpression of the receptor. We found that neuronal A2AR upregulation increases tau hyperphosphorylation, potentiating the onset of tau-induced memory deficits. This detrimental effect was linked to a singular microglial signature as revealed by RNA sequencing analysis. In particular, we found that A2AR overexpression in THY-Tau22 mice led to the hippocampal upregulation of C1q complement protein-also observed in patients with frontotemporal lobar degeneration-and correlated with the loss of glutamatergic synapses, likely underlying the observed memory deficits. These data reveal a key impact of overactive neuronal A2AR in the onset of synaptic loss in tauopathies, paving the way for new therapeutic approaches.

New piperazine multi-effect drugs prevent neurofibrillary degeneration and amyloid deposition, and preserve memory in animal models of Alzheimer's disease.

Alzheimer's Disease is a devastating dementing disease involving amyloid deposits, neurofibrillary tangles, progressive and irreversible cognitive impairment. Today, only symptomatic drugs are available and therapeutic treatments, possibly acting at a multiscale level, are thus urgently needed. To that purpose, we designed multi-effects compounds by synthesizing drug candidates derived by substituting a novel N,N'-disubstituted piperazine anti-amyloid scaffold and adding acetylcholinesterase inhibition property. Two compounds were synthesized and evaluated. The most promising hybrid molecule reduces both the amyloid pathology and the Tau pathology as well as the memory impairments in a preclinical model of Alzheimer's disease. In vitro also, the compound reduces the phosphorylation of Tau and inhibits the release of Aß peptides while preserving the processing of other metabolites of the amyloid precursor protein. We synthetized and tested the first drug capable of ameliorating both the amyloid and Tau pathology in animal models of AD as well as preventing the major brain lesions and associated memory impairments. This work paves the way for future compound medicines against both Alzheimer's-related brain lesions development and the associated cognitive impairments.

Brain insulin response and peripheral metabolic changes in a Tau transgenic mouse model.

Accumulation of hyper-phosphorylated and aggregated Tau proteins is a neuropathological hallmark of Alzheimer's Disease (AD) and Tauopathies. AD patient brains also exhibit insulin resistance. Whereas, under normal physiological conditions insulin signaling in the brain mediates plasticity and memory formation, it can also regulate peripheral energy homeostasis. Thus, in AD, brain insulin resistance affects both cognitive and metabolic changes described in these patients. While a role of Aß oligomers and APOE4 towards the development of brain insulin resistance emerged, contribution of Tau pathology has been largely overlooked. Our recent data demonstrated that one of the physiological function of Tau is to sustain brain insulin signaling. We postulated that under pathological conditions, hyper-phosphorylated/aggregated Tau is likely to lose this function and to favor the development of brain insulin resistance. This hypothesis was substantiated by observations from patient brains with pure Tauopathies. To address the potential link between Tau pathology and brain insulin resistance, we have evaluated the brain response to insulin in a transgenic mouse model of AD-like Tau pathology (THY-Tau22). Using electrophysiological and biochemical evaluations, we surprisingly observed that, at a time when Tau pathology and cognitive deficits are overt and obvious, the hippocampus of THY-Tau22 mice exhibits enhanced response to insulin. In addition, we demonstrated that the ability of i.c.v. insulin to promote body weight loss is enhanced in THY-Tau22 mice. In line with this, THY-Tau22 mice exhibited a lower body weight gain, hypoleptinemia and hypoinsulinemia and finally a metabolic resistance to high-fat diet. The present data highlight that the brain of transgenic Tau mice exhibit enhanced brain response to insulin. Whether these observations are ascribed to the development of Tau pathology, and therefore relevant to human Tauopathies, or unexpectedly results from the Tau transgene overexpression is debatable and discussed.

The FK506-binding protein FKBP52 in vitro induces aggregation of truncated Tau forms with prion-like behavior.

Tauopathies, including Alzheimer's disease (AD), are neurodegenerative diseases associated with the pathologic aggregation of human brain Tau protein. Neuronal Tau is involved in microtubule (MT) formation and stabilization. We showed previously that the immunophilin FK506-binding protein of MW ∼52 kDa (FKBP52) interferes with this function of full-length Tau and provokes aggregation of a disease-related mutant of Tau. To dissect the molecular interaction between recombinant human FKBP52 and Tau, here, we study the effect of FKBP52 on a functional Tau fragment (Tau-F4, Ser(208)-Ser(324)) containing part of the proline- rich region and MT-binding repeats. Therefore, we perform MT assembly and light-scattering assays, blue native PAGE analysis, electron microscopy, and Tau seeding experiments in SH-SY5Y human neuroblastoma cells. We show that FKBP52 (6 µM) prevents MT formation generated by Tau-F4 (5 µM) and induces Tau-F4 oligomerization and aggregation. Electron microscopy analyses show granular oligomers and filaments of Tau-F4 after short-time FKBP52 incubation. We demonstrate that the terminal parts of Tau interfere with the effects of FKBP52. Finally, we find that FKBP52-induced Tau-F4 oligomers cannot only generate in vitro, direct conformational changes in full-length Tau and that their uptake into neuronal cells can equally lead to aggregation of wild-type endogenous Tau. This suggests a potential prion-like property of these particular Tau-F4 aggregates. Collectively, our results strengthen the hypothesis of FKBP52 involvement in the Tau pathogenicity process.

From epidemiology to pathophysiology: what about caffeine in Alzheimer's disease?

AD (Alzheimer's disease) is the most prevalent form of dementia in the aged population. Definitive diagnosis of AD is based on the presence of senile plaques and neurofibrillary tangles that are identified in post-mortem brain specimens. A third pathological component is inflammation. AD results from multiple genetic and environmental risk factors. Among other factors, epidemiological studies report beneficial effects of caffeine, a non-selective antagonist of adenosine receptors. In the present review, we discuss the impact of caffeine and the adenosinergic system in AD pathology as well as consequences in terms of pathology and therapeutics.

Tau mRNA Metabolism in Neurodegenerative Diseases: A Tangle Journey.

Tau proteins are known to be mainly involved in regulation of microtubule dynamics. Besides this function, which is critical for axonal transport and signal transduction, tau proteins also have other roles in neurons. Moreover, tau proteins are turned into aggregates and consequently trigger many neurodegenerative diseases termed tauopathies, of which Alzheimer's disease (AD) is the figurehead. Such pathological aggregation processes are critical for the onset of these diseases. Among the various causes of tau protein pathogenicity, abnormal tau mRNA metabolism, expression and dysregulation of tau post-translational modifications are critical steps. Moreover, the relevance of tau function to general mRNA metabolism has been highlighted recently in tauopathies. In this review, we mainly focus on how mRNA metabolism impacts the onset and development of tauopathies. Thus, we intend to portray how mRNA metabolism of, or mediated by, tau is associated with neurodegenerative diseases.

Beneficial effects of exercise in a transgenic mouse model of Alzheimer's disease-like Tau pathology.

Tau pathology is encountered in many neurodegenerative disorders known as tauopathies, including Alzheimer's disease. Physical activity is a lifestyle factor affecting processes crucial for memory and synaptic plasticity. Whether long-term voluntary exercise has an impact on Tau pathology and its pathophysiological consequences is currently unknown. To address this question, we investigated the effects of long-term voluntary exercise in the THY-Tau22 transgenic model of Alzheimer's disease-like Tau pathology, characterized by the progressive development of Tau pathology, cholinergic alterations and subsequent memory impairments. Three-month-old THY-Tau22 mice and wild-type littermates were assigned to standard housing or housing supplemented with a running wheel. After 9 months of exercise, mice were evaluated for memory performance and examined for hippocampal Tau pathology, cholinergic defects, inflammation and genes related to cholesterol metabolism. Exercise prevented memory alterations in THY-Tau22 mice. This was accompanied by a decrease in hippocampal Tau pathology and a prevention of the loss of expression of choline acetyltransferase within the medial septum. Whereas the expression of most cholesterol-related genes remained unchanged in the hippocampus of running THY-Tau22 mice, we observed a significant upregulation in mRNA levels of NPC1 and NPC2, genes involved in cholesterol trafficking from the lysosomes. Our data support the view that long-term voluntary physical exercise is an effective strategy capable of mitigating Tau pathology and its pathophysiological consequences.

NMR spectroscopy of the neuronal tau protein: normal function and implication in Alzheimer's disease.

NMR spectroscopy was used to explore the different aspects of the normal and pathological functions of tau, but proved challenging because the protein contains 441 amino acids and has poor signal dispersion. We have set out to dissect the phosphorylation patterns of tau in order to understand better its role in the aggregation process and microtubule-binding regulation. Our current knowledge on the functional consequences of specific phosphorylations is still limited, mainly because producing and assessing quantitatively phosphorylated tau samples is far from straightforward, even in vitro. We use NMR spectroscopy as a proteomics tool to characterize the phosphorylation patterns of tau, after in vitro phosphorylation by recombinant kinases. The phosphorylated tau can next be use for functional assays or interaction assays with phospho-dependent protein partners, such as the prolyl cis-trans isomerase Pin1.

From tau phosphorylation to tau aggregation: what about neuronal death?

Tau pathology is characterized by intracellular aggregates of abnormally and hyperphosphorylated tau proteins. It is encountered in many neurodegenerative disorders, but also in aging. These neurodegenerative disorders are referred to as tauopathies. Comparative biochemistry of the tau aggregates shows that they differ in both tau isoform phosphorylation and content, which enables a molecular classification of tauopathies. In conditions of dementia, NFD (neurofibrillary degeneration) severity is correlated to cognitive impairment and is often considered as neuronal death. Using tau animal models, analysis of the kinetics of tau phosphorylation, aggregation and neuronal death in parallel to electrophysiological and behavioural parameters indicates a disconnection between cognition deficits and neuronal cell death. Tau phosphorylation and aggregation are early events followed by cognitive impairment. Neuronal death is not observed before the oldest ages. A sequence of events may be the formation of toxic phosphorylated tau species, their aggregation, the formation of neurofibrillary tangles (from pre-tangles to ghost tangles) and finally neuronal cell death. This sequence will last from 15 to 25 years and one can ask whether the aggregation of toxic phosphorylated tau species is a protection against cell death. Apoptosis takes 24 h, but NFD lasts for 24 years to finally kill the neuron or rather to protect it for more than 20 years. Altogether, these data suggest that NFD is a transient state before neuronal death and that therapeutic interventions are possible at that stage.

[Molecular actors in Alzheimer's disease: which diagnostic and therapeutic consequences?]. / Comment les acteurs moléculaires de la pathologie Alzheimer permettent de comprendre la démence ? Quelles conséquences diagnostiques et thérapeutiques ?

Alzheimer's disease is a neurodegenerative disorder characterized by neuropathological lesions: amyloid deposits and neurofibrillary degeneration. However, the links between these two brain hallmarks are still poorly understood. Until now, mainly amyloid pathology has been targeted un many clinical trials without any success. Both new therapeutic strategies and diagnosis improvement are needed.

PUFA induce antidepressant-like effects in parallel to structural and molecular changes in the hippocampus.

Epidemiological data suggest that omega-3 polyunsaturated fatty acids (PUFA) consumption may be inversely correlated to the prevalence and severity of depression but little is known about the underlying mechanisms. In this study, we experimentally investigated whether a chronic supplementation with PUFA may induce antidepressant-like effects in mice in parallel to brain structural and molecular changes. Six weeks feeding with a PUFA-enriched diet induced behavioral changes in the Forced Swim Test (FST), the Tail Suspension Test and the Novelty-Suppressed Feeding Test. Moreover, more than 5 weeks supplementation with a PUFA blend containing 70% alpha-linolenic acid induced antidepressant-like effects in the FST with an increase in both swimming and climbing behaviors. The combination of a shorter duration of PUFA supplementation with a low dose of imipramine also induced an additive effect in the FST. Finally, PUFA supplementation was associated with an increase in the hippocampal volume, an over-expression of both synaptophysin and BDNF, and a raise in the number of newborn cells. Besides the possible modulation of brain plasticity, present results highlight the effectiveness of PUFA given alone or in combination with antidepressant drug as potential treatment of depressive disorders.

Biochemistry of Tau in Alzheimer's disease and related neurological disorders.

Microtubule-associated Tau proteins belong to a family of factors that polymerize tubulin dimers and stabilize microtubules. Tau is strongly expressed in neurons, localized in the axon and is essential for neuronal plasticity and network. From the very beginning of Tau discovery, proteomics methods have been essential to the knowledge of Tau biochemistry and biology. In this review, we have summarized the main contributions of several proteomic methods in the understanding of Tau, including expression, post-translational modifications and structure, in both physiological and pathophysiological aspects. Finally, recent advances in proteomics technology are essential to develop further therapeutic targets and early predictive and discriminative diagnostic assays for Alzheimer's disease and related disorders.

Tau deletion promotes brain insulin resistance.

The molecular pathways underlying tau pathology-induced synaptic/cognitive deficits and neurodegeneration are poorly understood. One prevalent hypothesis is that hyperphosphorylation, misfolding, and fibrillization of tau impair synaptic plasticity and cause degeneration. However, tau pathology may also result in the loss of specific physiological tau functions, which are largely unknown but could contribute to neuronal dysfunction. In the present study, we uncovered a novel function of tau in its ability to regulate brain insulin signaling. We found that tau deletion leads to an impaired hippocampal response to insulin, caused by altered IRS-1 and PTEN (phosphatase and tensin homologue on chromosome 10) activities. Our data also demonstrate that tau knockout mice exhibit an impaired hypothalamic anorexigenic effect of insulin that is associated with energy metabolism alterations. Consistently, we found that tau haplotypes are associated with glycemic traits in humans. The present data have far-reaching clinical implications and raise the hypothesis that pathophysiological tau loss-of-function favors brain insulin resistance, which is instrumental for cognitive and metabolic impairments in Alzheimer's disease patients.

Highly successful therapeutic vaccinations combining dendritic cells and tumor cells secreting granulocyte macrophage colony-stimulating factor.

In an attempt to induce potent immune antitumor activities, we investigated, within the rat 9L gliosarcoma model, distal therapeutic vaccinations associating three therapies: dendritic cell vaccination, intratumoral granulocyte macrophage colony-stimulating factor (GM-CSF) gene transfer, and tumor apoptosis induction. Vaccines of dendritic cells coinjected with processed GM-CSF secreting 9L cells induced systemic responses, resulting in the complete regression of distant preimplanted 9L tumor masses in, with the best strategy, 94% of male rats. All of the cured rats developed a long-term resistance to a rechallenge with parental cells. The curative responses were correlated with the detection of elevated specific cytotoxic activities and a CD4+, CD8+ T cell-, and natural killer (NK) cell-mediated IFN-gamma production. The survival rate of the rat seemed more directly linked to the amount of GM-CSF secreted by the transduced tumor cells, which in turn depended on the toxicity of the apoptosis-inducing treatment, than to the level of apoptosis induced. Unexpectedly, alive GM-CSF secreting 9L cells became apoptotic when injected in vivo. Thus we documented the positive role of apoptosis in the induction of therapeutic antitumor responses by comparing, at equal GM-CSF exogenous supply, the effects of dendritic cells coinjected with apoptotic or necrotic 9L cells. The data showed the superior therapeutic efficiency of combined vaccines containing apoptotic tumor cells. In conclusion, vaccinations with dendritic cells associated with apoptotic tumor cells secreting GM-CSF show a very high therapeutic potency that should show promise for the treatment of human cancer.

The caffeine-binding adenosine A2A receptor induces age-like HPA-axis dysfunction by targeting glucocorticoid receptor function.

Caffeine is associated with procognitive effects in humans by counteracting overactivation of the adenosine A2A receptor (A2AR), which is upregulated in the human forebrain of aged and Alzheimer's disease (AD) patients. We have previously shown that an anti-A2AR therapy reverts age-like memory deficits, by reestablishment of the hypothalamic-pituitary-adrenal (HPA) axis feedback and corticosterone circadian levels. These observations suggest that A2AR over-activation and glucocorticoid dysfunction are key events in age-related hippocampal deficits; but their direct connection has never been explored. We now show that inducing A2AR overexpression in an aging-like profile is sufficient to trigger HPA-axis dysfunction, namely loss of plasmatic corticosterone circadian oscillation, and promotes reduction of GR hippocampal levels. The synaptic plasticity and memory deficits triggered by GR in the hippocampus are amplified by A2AR over-activation and were rescued by anti-A2AR therapy; finally, we demonstrate that A2AR act on GR nuclear translocation and GR-dependent transcriptional regulation. We provide the first demonstration that A2AR is a major regulator of GR function and that this functional interconnection may be a trigger to age-related memory deficits. This supports the idea that the procognitive effects of A2AR antagonists, namely caffeine, on Alzheimer's and age-related cognitive impairments may rely on its ability to modulate GR actions.

Role of the Tau N-terminal region in microtubule stabilization revealed by new endogenous truncated forms.

Tau is a central player in Alzheimer's disease (AD) and related Tauopathies, where it is found as aggregates in degenerating neurons. Abnormal post-translational modifications, such as truncation, are likely involved in the pathological process. A major step forward in understanding the role of Tau truncation would be to identify the precise cleavage sites of the several truncated Tau fragments that are observed until now in AD brains, especially those truncated at the N-terminus, which are less characterized than those truncated at the C-terminus. Here, we optimized a proteomics approach and succeeded in identifying a number of new N-terminally truncated Tau species from the human brain. We initiated cell-based functional studies by analyzing the biochemical characteristics of two N-terminally truncated Tau species starting at residues Met11 and Gln124 respectively. Our results show, interestingly, that the Gln124-Tau fragment displays a stronger ability to bind and stabilize microtubules, suggesting that the Tau N-terminal domain could play a direct role in the regulation of microtubule stabilization. Future studies based on our new N-terminally truncated-Tau species should improve our knowledge of the role of truncation in Tau biology as well as in the AD pathological process.

Chloroquine and chloroquinoline derivatives as models for the design of modulators of amyloid Peptide precursor metabolism.

The amyloid precursor protein (APP) plays a central role in Alzheimer's disease (AD). Preventing deregulated APP processing by inhibiting amyloidogenic processing of carboxy-terminal fragments (APP-CTFs), and reducing the toxic effect of amyloid beta (Aß) peptides remain an effective therapeutic strategy. We report the design of piperazine-containing compounds derived from chloroquine structure and evaluation of their effects on APP metabolism and ability to modulate the processing of APP-CTF and the production of Aß peptide. Compounds which retained alkaline properties and high affinity for acidic cell compartments were the most effective. The present study demonstrates that (1) the amino side chain of chloroquine can be efficiently substituted by a bis(alkylamino)piperazine chain, (2) the quinoline nucleus can be replaced by a benzyl or a benzimidazole moiety, and (3) pharmacomodulation of the chemical structure allows the redirection of APP metabolism toward a decrease of Aß peptide release, and increased stability of APP-CTFs and amyloid intracellular fragment. Moreover, the benzimidazole compound 29 increases APP-CTFs in vivo and shows promising activity by the oral route. Together, this family of compounds retains a lysosomotropic activity which inhibits lysosome-related Aß production, and is likely to be beneficial for therapeutic applications in AD.

Neurofibrillary degeneration of the Alzheimer-type: an alternate pathway to neuronal apoptosis?

Neuronal death is a process which may be either physiological or pathological. Apoptosis and necrosis are two of these processes which are particularly studied. However, in neurodegenerative disorders, some neurons escape to these types of death and "agonize" in a process referred to as neurofibrillary degeneration. Neurofibrillary degeneration is characterized by the intraneuronal aggregation of abnormally phosphorylated microtubule-associated Tau proteins. A number of studies have reported a reactivation of the cell cycle in the neurofibrillary degeneration process. This reactivation of the cell cycle is reminiscent of the initiation of apoptosis in post-mitotic cells where G1/S markers including cyclin D1 and cdk4/6, are commonly found. However, in neurons exhibiting neurofibrillary degeneration, both G1/S and G2/M markers are found suggesting that they do not follow the classical apoptosis and an aberrant cell cycle occurs. This aberrant response leading to neurofibrillary degeneration may be triggered by the sequential combination of three partners: the complex Cdk5/p25 induces both apoptosis and the "abnormal mitotic Tau phosphorylation". These mitotic epitopes may allow for the nuclear depletion of Pin1. This latter may be responsible for escaping classical apoptosis in a subset of neurons. Since neurofibrillary degeneration is likely to be a third way to die, molecular mechanisms leading to changes in Tau phosphorylation including activation of kinases such as cdk5 or other regulators such as Pin1 could be important drug targets as they are possibly involved in early stages of neurodegeneration.

Pin1: a therapeutic target in Alzheimer neurodegeneration.

In Alzheimer's disease, the peptidyl prolyl cis/trans isomerase Pin1 binds to phospho-Thr231 on Tau proteins and, hence, is found within degenerating neurons, where it is associated to the large amounts of abnormally phosphorylated Tau proteins. Conversely, Pin1 may restore the tubulin polymerization function of these hyperphosphorylated Tau. In the present work, we investigated, both at the cellular and molecular levels, the role of Pin1 in Alzheimer's disease through the study of its interactions with phosphorylated Tau proteins. We also showed that in neuronal cells, Pin1 upregulates the expression of cyclin D1. This, in turn, could facilitate the transition from quiescence to the G1 phase (re-entry in cell cycle) in a neuron and, subsequently, neuronal dedifferentiation and apoptosis. The involvement of Pin1 in the G0/G1 transition in neurons points to its function as a good target for the development of new therapeutic strategies in neurodegenerative disorders.