Age-Dependent Changes in Nrf2/Keap1 and Target Antioxidant Protein Expression Correlate to Lipoxidative Adducts, and Are Modulated by Dietary N-3 LCPUFA in the Hippocampus of Mice.

The brain has a high metabolism rate that may generate reactive oxygen and nitrogen species. Consequently, nerve cells require highly efficient antioxidant defenses in order to prevent a condition of deleterious oxidative stress. This is particularly relevant in the hippocampus, a highly complex cerebral area involved in processing superior cognitive functions. Most current evidence points to hippocampal oxidative damage as a causal effect for neurodegenerative disorders, especially Alzheimer's disease. Nuclear factor erythroid-2-related factor 2/Kelch-like ECH-associated protein 1 (Nrf2/Keap1) is a master key for the transcriptional regulation of antioxidant and detoxifying systems. It is ubiquitously expressed in brain areas, mainly supporting glial cells. In the present study, we have analyzed the relationships between Nrf2 and Keap1 isoforms in hippocampal tissue in response to aging and dietary long-chain polyunsaturated fatty acids (LCPUFA) supplementation. The possible involvement of lipoxidative and nitrosative by-products in the dynamics of the Nrf2/Keap1 complex was examined though determination of protein adducts, namely malondialdehyde (MDA), 4-hydroxynonenal (HNE), and 3-nitro-tyrosine (NTyr) under basal conditions. The results were correlated to the expression of target proteins heme-oxygenase-1 (HO-1) and glutathione peroxidase 4 (GPx4), whose expressions are known to be regulated by Nrf2/Keap1 signaling activation. All variables in this study were obtained simultaneously from the same preparations, allowing multivariate approaches. The results demonstrate a complex modification of the protein expression patterns together with the formation of adducts in response to aging and diet supplementation. Both parameters exhibited a strong interaction. Noticeably, LCPUFA supplementation to aged animals restored the Nrf2/Keap1/target protein patterns to the status observed in young animals, therefore driving a "rejuvenation" of hippocampal antioxidant defense.

Changes in Oxidative Stress and Intestinal Permeability during Pregnancy in Women with Gestational Diabetes Mellitus Treated with Metformin or Insulin and Healthy Controls: A Randomized Controlled Trial.

Both oxidative stress and intestinal permeability are increased in hyperglycemic situations and have been shown to be reduced by metformin in type 2 diabetes mellitus (T2DM) patients. The aim of this study was to elucidate the effect of metformin on oxidative stress and intestinal permeability in women with gestational diabetes mellitus (GDM) treated with metformin compared to those treated with insulin and healthy controls. A total of 120 women were included from August 2016 to February 2022: 41 received metformin (MET group), 38 received insulin (INS group), and 41 were healthy controls. Baseline and antenatal visits were carried out at 25.4 ± 4.8 and 36.1 ± 0.8 weeks of pregnancy, respectively. Advanced oxidation protein products (AOPPs), total antioxidant capacity (TAC), and zonulin levels were measured at every visit. Zonulin levels from baseline to prepartum visit increased significantly in both healthy controls (0.6 ± 0.9 to 1.2 ± 1.7 ng/mL, p = 0.004) and the INS group (0.4 ± 0.3 to 0.6 ± 0.5 ng/mL, p = 0.034) but did not significantly change in the MET group (0.4 ± 0.4 to 0.5 ± 0.4 ng/mL, p = 0.202). However, TAC and AOPP levels significantly increased in women with GDM, both in the INS and MET groups but not in the healthy controls. In conclusion, in our population, metformin has been shown to avoid an increase in intestinal permeability but failed to avoid an increase in oxidative stress related to hyperglycemia.

The Relationship between Depressive Symptoms, Quality of Life and miRNAs 8 Years after Bariatric Surgery.

(1) Background: There are conflicting results on whether weight loss after bariatric surgery (BS) might be associated with quality of life (QoL)/depressive symptomatology. We aim to determine whether BS outcomes are associated with QoL/depressive symptomatology in studied patients at the 8-year follow-up after BS, as well as their relationship with different serum proteins and miRNAs. (2) Methods: A total of 53 patients with class III obesity who underwent BS, and then classified into "good responders" and "non-responders" depending on the percentage of excess weight lost (%EWL) 8 years after BS (%EWL ≥ 50% and %EWL < 50%, respectively), were included. Basal serum miRNAs and different proteins were analysed, and patients completed tests to evaluate QoL/depressive symptomatology at 8 years after BS. (3) Results: The good responders group showed higher scores on SF-36 scales of physical functioning, role functioning-physical, role functioning-emotional, body pain and global general health compared with the non-responders. The expression of hsa-miR-101-3p, hsa-miR-15a-5p, hsa-miR-29c-3p, hsa-miR-144-3p and hsa-miR-19b-3p were lower in non-responders. Hsa-miR-19b-3p was the variable associated with the response to BS in a logistic regression model. (4) Conclusions: The mental health of patients after BS is limited by the success of the intervention. In addition, the expression of basal serum miRNAs related to depression/anxiety could predict the success of BS.

Molecular and biophysical features of hippocampal "lipid rafts aging" are modified by dietary n-3 long-chain polyunsaturated fatty acids.

"Lipid raft aging" in nerve cells represents an early event in the development of aging-related neurodegenerative diseases, such as Alzheimer's disease. Lipid rafts are key elements in synaptic plasticity, and their modification with aging alters interactions and distribution of signaling molecules, such as glutamate receptors and ion channels involved in memory formation, eventually leading to cognitive decline. In the present study, we have analyzed, in vivo, the effects of dietary supplementation of n-3 LCPUFA on the lipid structure, membrane microviscosity, domain organization, and partitioning of ionotropic and metabotropic glutamate receptors in hippocampal lipid raffs in female mice. The results revealed several lipid signatures of "lipid rafts aging" in old mice fed control diets, consisting in depletion of n-3 LCPUFA, membrane unsaturation, along with increased levels of saturates, plasmalogens, and sterol esters, as well as altered lipid relevant indexes. These changes were paralleled by increased microviscosity and changes in the raft/non-raft (R/NR) distribution of AMPA-R and mGluR5. Administration of the n-3 LCPUFA diet caused the partial reversion of fatty acid alterations found in aged mice and returned membrane microviscosity to values found in young animals. Paralleling these findings, lipid rafts accumulated mGluR5, NMDA-R, and ASIC2, and increased their R/NR proportions, which collectively indicate changes in synaptic plasticity. Unexpectedly, this diet also modified the lipidome and dimension of lipid rafts, as well as the domain redistribution of glutamate receptors and acid-sensing ion channels involved in hippocampal synaptic plasticity, likely modulating functionality of lipid rafts in memory formation and reluctance to age-associated cognitive decline.

Effects of Dietary n-3 LCPUFA Supplementation on the Hippocampus of Aging Female Mice: Impact on Memory, Lipid Raft-Associated Glutamatergic Receptors and Neuroinflammation.

Long-chain polyunsaturated fatty acids (LCPUFA), essential molecules whose precursors must be dietary supplied, are highly represented in the brain contributing to numerous neuronal processes. Recent findings have demonstrated that LCPUFA are represented in lipid raft microstructures, where they favor molecular interactions of signaling complexes underlying neuronal functionality. During aging, the brain lipid composition changes affecting the lipid rafts' integrity and protein signaling, which may induce memory detriment. We investigated the effect of a n-3 LCPUFA-enriched diet on the cognitive function of 6- and 15-months-old female mice. Likewise, we explored the impact of dietary n-3 LCPUFAs on hippocampal lipid rafts, and their potential correlation with aging-induced neuroinflammation. Our results demonstrate that n-3 LCPUFA supplementation improves spatial and recognition memory and restores the expression of glutamate and estrogen receptors in the hippocampal lipid rafts of aged mice to similar profiles than young ones. Additionally, the n-3 LCPUFA-enriched diet stabilized the lipid composition of the old mice's hippocampal lipid rafts to the levels of young ones and reduced the aged-induced neuroinflammatory markers. Hence, we propose that n-3 LCPUFA supplementation leads to beneficial cognitive performance by "rejuvenating" the lipid raft microenvironment that stabilizes the integrity and interactions of memory protein players embedded in these microdomains.

Neuronal ER-Signalosome Proteins as Early Biomarkers in Prodromal Alzheimer's Disease Independent of Amyloid-ß Production and Tau Phosphorylation.

There exists considerable interest to unveil preclinical period and prodromal stages of Alzheimer's disease (AD). The mild cognitive impairment (MCI) is characterized by significant memory and/or other cognitive domains impairments, and is often considered the prodromal phase of AD. The cerebrospinal fluid (CSF) levels of ß-amyloid (ßA), total tau (t-tau), and phosphorylated tau (p-tau) have been used as biomarkers of AD albeit their significance as indicators during early stages of AD remains far from accurate. The new biomarkers are being intensively sought as to allow identification of pathological processes underlying early stages of AD. Fifty-three participants (75.4 ± 8.3 years) were classified in three groups as cognitively normal healthy controls (HC), MCI, and subjective memory complaints (SMC). The subjects were subjected to a battery of neurocognitive tests and underwent lumbar puncture for CSF extraction. The CSF levels of estrogen-receptor (ER)-signalosome proteins, ßA, t-tau and p-tau, were submitted to univariate, bivariate, and multivariate statistical analyses. We have found that the components of the ER-signalosome, namely, caveolin-1, flotilin-1, and estrogen receptor alpha (ERα), insulin growth factor-1 receptor ß (IGF1Rß), prion protein (PrP), and plasmalemmal voltage dependent anion channel 1 (VDAC) could be detected in the CSF from all subjects of the HC, MCI, and SMC groups. The six proteins appeared elevated in MCI and slightly increased in SMC subjects compared to HC, suggesting that signalosome proteins undergo very early modifications in nerve cells. Using a multivariate approach, we have found that the combination of ERα, IGF-1Rß, and VDAC are the main determinants of group segregation with resolution enough to predict the MCI stage. The analyses of bivariate relationships indicated that collinearity of ER-signalosome proteins vary depending on the stage, with some pairs displaying opposed relationships between HC and MCI groups, and the SMC stage showing either no relationships or behaviors similar to either HC or MCI stages. The multinomial logistic regression models of changes in ER-signalosome proteins provide reliable predictive criteria, particularly for the MCI. Notably, most of the statistical analyses revealed no significant relationships or interactions with classical AD biomarkers at either disease stage. Finally, the multivariate functions were highly correlated with outcomes from neurocognitive tests for episodic memory. These results demonstrate that alterations in ER-signalosome might provide useful diagnostic information on preclinical stages of AD, independently from classical biomarkers.

Multivariate Assessment of Lipoxidative Metabolites, Trace Biometals, and Antioxidant and Detoxifying Activities in the Cerebrospinal Fluid Define a Fingerprint of Preclinical Stages of Alzheimer's Disease.

BACKGROUND: There exists considerable interest in the identification of molecular traits during early stages of Alzheimer's disease (AD). Mild cognitive impairment (MCI) is considered the closest prodromal stage of AD, and to develop gradually from earlier stages although not always progresses to AD. Classical cerebrospinal fluid (CSF) AD biomarkers, amyloid-ß peptides and tau/p-tau proteins, have been measured in prodromal stages yet results are heterogeneous and far from conclusive. Therefore, there exists a pressing need to identify a neurochemical signature for prodromal stages and to predict which cases might progress to AD. OBJECTIVE: Exploring potential CSF biomarkers related to brain oxidative and inorganic biochemistry during prodromal stages of the disease. METHODS: We have analyzed CSF levels of lipoxidative markers (MDA and 8-isoF2α), biometals (Cu, Zn, Se, Mn, and Fe), iron-transport protein transferrin (TFER), antioxidant enzymes (SOD and GPx4), detoxifying enzymes (GST and BuChE), as well as classical amyloid-ß and total and phosphorylated tau, in cognitively healthy controls, patients with MCI, and subjects exhibiting subjective memory complaints (SMC). RESULTS: Inter-group differences for several variables exhibit differentiable trends along the HC ⟶ SMC ⟶ MCI sequence. More interestingly, the combination of Se, Cu, Zn, SOD, TFER, and GST variables allow differentiable fingerprints for control subjects and each prodromal stage. Further, multivariate scores correlate positively with neurocognitive In-Out test, hence with both episodic memory decline and prediction to dementia. CONCLUSION: We conclude that changes in the CSF biochemistry related to brain oxidative defense and neurometallomics might provide more powerful and accurate diagnostic tools in preclinical stages of AD.

Disarranged neuroplastin environment upon aging and chronic stress recovery in female Sprague Dawley rats.

Chronic stress produces long-term metabolic changes throughout the superfamily of nuclear receptors, potentially causing various pathologies. Sex hormones modulate the stress response and generate a sex-specific age-dependent metabolic imprint, especially distinct in the reproductive senescence of females. We monitored chronic stress recovery in two age groups of female Sprague Dawley rats to determine whether stress and/or aging structurally changed the glycolipid microenvironment, a milieu playing an important role in cognitive functions. Old females experienced memory impairment even at basal conditions, which was additionally amplified by stress. On the other hand, the memory of young females was not disrupted. Stress recovery was followed by a microglial decrease and an increase in astrocyte count in the hippocampal immune system. Since dysfunction of the brain immune system could contribute to disturbed synaptogenesis, we analyzed neuroplastin expression and the lipid environment. Neuroplastin microenvironments were explored by analyzing immunofluorescent stainings using a newly developed Python script method. Stress reorganized glycolipid microenvironment in the Cornu Ammonis 1 (CA1) and dentate gyrus (DG) hippocampal regions of old females but in a very different fashion, thus affecting neuroplasticity. The postulation of four possible neuroplastin environments pointed to the GD1a ganglioside enrichment during reproductive senescence of stressed females, as well as its high dispersion in both regions and to GD1a and GM1 loss in the CA1 region. A specific lipid environment might influence neuroplastin functionality and underlie synaptic dysfunction triggered by a combination of aging and chronic stress.

Synthesis and Biological Assessment of 4,1-Benzothiazepines with Neuroprotective Activity on the Ca2+ Overload for the Treatment of Neurodegenerative Diseases and Stroke.

In excitable cells, mitochondria play a key role in the regulation of the cytosolic Ca2+ levels. A dysregulation of the mitochondrial Ca2+ buffering machinery derives in serious pathologies, where neurodegenerative diseases highlight. Since the mitochondrial Na+/Ca2+ exchanger (NCLX) is the principal efflux pathway of Ca2+ to the cytosol, drugs capable of blocking NCLX have been proposed to act as neuroprotectants in neuronal damage scenarios exacerbated by Ca2+ overload. In our search of optimized NCLX blockers with augmented drug-likeness, we herein describe the synthesis and pharmacological characterization of new benzothiazepines analogues to the first-in-class NCLX blocker CGP37157 and its further derivative ITH12575, synthesized by our research group. As a result, we found two new compounds with an increased neuroprotective activity, neuronal Ca2+ regulatory activity and improved drug-likeness and pharmacokinetic properties, such as clog p or brain permeability, measured by PAMPA experiments.

DHA and Its Elaborated Modulation of Antioxidant Defenses of the Brain: Implications in Aging and AD Neurodegeneration.

DHA (docosahexaenoic acid) is perhaps the most pleiotropic molecule in nerve cell biology. This long-chain highly unsaturated fatty acid has evolved to accomplish essential functions ranging from structural components allowing fast events in nerve cell membrane physiology to regulation of neurogenesis and synaptic function. Strikingly, the plethora of DHA effects has to take place within the hostile pro-oxidant environment of the brain parenchyma, which might suggest a molecular suicide. In order to circumvent this paradox, different molecular strategies have evolved during the evolution of brain cells to preserve DHA and to minimize the deleterious effects of its oxidation. In this context, DHA has emerged as a member of the "indirect antioxidants" family, the redox effects of which are not due to direct redox interactions with reactive species, but to modulation of gene expression within thioredoxin and glutathione antioxidant systems and related pathways. Weakening or deregulation of these self-protecting defenses orchestrated by DHA is associated with normal aging but also, more worryingly, with the development of neurodegenerative diseases. In the present review, we elaborate on the essential functions of DHA in the brain, including its role as indirect antioxidant, the selenium connection for proper antioxidant function and their changes during normal aging and in Alzheimer's disease.

FLTX2: A Novel Tamoxifen Derivative Endowed with Antiestrogenic, Fluorescent, and Photosensitizer Properties.

Tamoxifen is the most widely used selective modulator of estrogen receptors (SERM) and the first strategy as coadjuvant therapy for the treatment of estrogen-receptor (ER) positive breast cancer worldwide. In spite of such success, tamoxifen is not devoid of undesirable effects, the most life-threatening reported so far affecting uterine tissues. Indeed, tamoxifen treatment is discouraged in women under risk of uterine cancers. Recent molecular design efforts have endeavoured the development of tamoxifen derivatives with antiestrogen properties but lacking agonistic uterine tropism. One of this is FLTX2, formed by the covalent binding of tamoxifen as ER binding core, 7-nitrobenzofurazan (NBD) as the florescent dye, and Rose Bengal (RB) as source for reactive oxygen species. Our analyses demonstrate (1) FLTX2 is endowed with similar antiestrogen potency as tamoxifen and its predecessor FLTX1, (2) shows a strong absorption in the blue spectral range, associated to the NBD moiety, which efficiently transfers the excitation energy to RB through intramolecular FRET mechanism, (3) generates superoxide anions in a concentration- and irradiation time-dependent process, and (4) Induces concentration- and time-dependent MCF7 apoptotic cell death. These properties make FLTX2 a very promising candidate to lead a novel generation of SERMs with the endogenous capacity to promote breast tumour cell death in situ by photosensitization.

The Impact of Microbiota on the Pathogenesis of Amyotrophic Lateral Sclerosis and the Possible Benefits of Polyphenols. An Overview.

The relationship between gut microbiota and neurodegenerative diseases is becoming clearer. Among said diseases amyotrophic lateral sclerosis (ALS) stands out due to its severity and, as with other chronic pathologies that cause neurodegeneration, gut microbiota could play a fundamental role in its pathogenesis. Therefore, polyphenols could be a therapeutic alternative due to their anti-inflammatory action and probiotic effect. Thus, the objective of our narrative review was to identify those bacteria that could have connection with the mentioned disease (ALS) and to analyze the benefits produced by administering polyphenols. Therefore, an extensive search was carried out selecting the most relevant articles published between 2005 and 2020 on the PubMed and EBSCO database on research carried out on cell, animal and human models of the disease. Thereby, after selecting, analyzing and debating the main articles on this topic, the bacteria related to the pathogenesis of ALS have been identified, among which we can positively highlight the presence mainly of Akkermansia muciniphila, but also Lactobacillus spp., Bifidobacterium spp. or Butyrivibrio fibrisolvens. Nevertheless, the presence of Escherichia coli or Ruminococcus torques stand out negatively for the disease. In addition, most of these bacteria are associated with molecular changes also linked to the pathogenesis of ALS. However, once the main polyphenols related to improvements in any of these three ALS models were assessed, many of them show positive results that could improve the prognosis of the disease. Nonetheless, epigallocatechin gallate (EGCG), curcumin and resveratrol are the polyphenols considered to show the most promising results as a therapeutic alternative for ALS through changes in microbiota.

Lipid and Lipid Raft Alteration in Aging and Neurodegenerative Diseases: A Window for the Development of New Biomarkers.

Lipids in the brain are major components playing structural functions as well as physiological roles in nerve cells, such as neural communication, neurogenesis, synaptic transmission, signal transduction, membrane compartmentalization, and regulation of gene expression. Determination of brain lipid composition may provide not only essential information about normal brain functioning, but also about changes with aging and diseases. Indeed, deregulations of specific lipid classes and lipid homeostasis have been demonstrated in neurodegenerative disorders such as Alzheimer's disease (AD) and Parkinson's disease (PD). Furthermore, recent studies have shown that membrane microdomains, named lipid rafts, may change their composition in correlation with neuronal impairment. Lipid rafts are key factors for signaling processes for cellular responses. Lipid alteration in these signaling platforms may correlate with abnormal protein distribution and aggregation, toxic cell signaling, and other neuropathological events related with these diseases. This review highlights the manner lipid changes in lipid rafts may participate in the modulation of neuropathological events related to AD and PD. Understanding and characterizing these changes may contribute to the development of novel and specific diagnostic and prognostic biomarkers in routinely clinical practice.

Differential Aggregation and Phosphorylation of Alpha Synuclein in Membrane Compartments Associated With Parkinson Disease.

The aggregation of α-synuclein (α-syn) is a major factor behind the onset of Parkinson's disease (PD). Sublocalization of this protein may be relevant for the formation of multimeric α-syn oligomeric configurations, insoluble aggregates that form Lewy bodies in PD brains. Processing of this protein aggregation is regulated by associations with distinct lipid classes. For instance, instability of lipid raft (LR) microdomains, membrane regions with a particular lipid composition, is an early event in the development of PD. However, the relevance of membrane microdomains in the regulation and trafficking of the distinct α-syn configurations associated with PD remains unexplored. In this study, using 6- and 14-month-old healthy and MPTP-treated animals as a model of PD, we have investigated the putative molecular alterations of raft membrane microstructures, and their impact on α-syn dynamics and conformation. A comparison of lipid analyses of LR microstructures and non-raft (NR) fractions showed alterations in gangliosides, cholesterol, polyunsaturated fatty acids (PUFA) and phospholipids in the midbrain and cortex of aged and MPTP-treated mice. In particular, the increase of PUFA and phosphatidylserine (PS) during aging correlated with α-syn multimeric formation in NR. In these aggregates, α-syn was phosphorylated in pSer129, the most abundant post-transductional modification of α-syn promoting toxic aggregation. Interestingly, similar variations in PUFA and PS content correlating with α-syn insoluble accumulation were also detected in membrane microstructures from the human cortex of incidental Parkinson Disease (iPD) and PD, as compared to healthy controls. Furthermore, structural changes in membrane lipid microenvironments may induce rearrangements in raft-interacting proteins involved in other neuropathologies. Therefore, we also investigated the dynamic of other protein markers involved in cognition and memory impairment such as metabotropic glutamate receptor 5 (mGluR5), ionotropic NMDA receptor (NMDAR2B), prion protein (PrPc) and amyloid precursor protein (APP), whose activity depends on membrane lipid organization. We observed a decline of these protein markers in LR fractions with the progression of aging and pathology. Overall, our findings demonstrate that lipid alterations in membranous compartments promoted by brain aging and PD-like injury may have an effect on α-syn aggregation and segregation in abnormal multimeric structures.

Lipostatic Mechanisms Preserving Cerebellar Lipids in MPTP-Treated Mice: Focus on Membrane Microdomains and Lipid-Related Gene Expression.

The cerebellum is an essential component in the control of motor patterns. Despite dramatic alteration of basal ganglia morpho-functionality in Parkinson's disease (PD), cerebellar function appears to be unaffected by the disease. Only recently this brain structure has been proposed to play compensatory roles in PD-induced motor dysfunction, particularly during the initial asymptomatic stages of PD. In PD subjects and animal models of PD, such as MPTP-treated mice, brain structures other than basal ganglia are also affected by the disease, including cortical areas not involved in motor control. Thus, it is noteworthy that the cerebellum remains unaffected. In the present study, we have analyzed the lipid composition of membrane microdomains [lipid rafts (LR) and non-raft domains] and assessed the expression levels of genes encoding enzymes synthesizing membrane-related lipids. The outcomes revealed that membrane domain lipids in cerebellum are highly preserved both in control and MPTP-treated mice as compared to control animals. Likewise, only small, mostly not significant, changes were observed in the expression of lipid-related genes in the cerebellum. Indeed, most changes were related to aging rather than to the exposure to the neurotoxin. Conversely, in the same animals, lipid composition, and gene expression were dramatically altered in the occipital cortex (OC), a brain area unrelated to the control of motor function. PCR and immunohistochemical analyses of both brain areas revealed that dopamine transporter (DAT) mRNA and protein were expressed in OC but not in the cerebellum. As MPTP neurotoxicity requires the expression of DAT to access intracellular compartments, we hypothesized that the absence of DAT in cerebellum hampers MPTP-induced toxicity. We conclude that cerebellum is endowed with efficient mechanisms to preserve nerve cell lipid homeostasis, which greatly maintain the stability of membrane microdomains involved in synaptic transmission, signal transduction, and intercellular communication, which together may participate in the compensatory role of the cerebellum in PD symptomatology.

In-Out-Test: A New Paradigm for Sorting the Wheat from the Chaff in Prodromal Alzheimer's Disease.

BACKGROUND: Assessment of hippocampal amnesia is helpful to distinguish between normal cognition and mild cognitive impairment (MCI), but not for identifying converters to dementia. Here biomarkers are useful but novel neuropsychological approaches are needed in their absence. The In-out-test assesses episodic memory using a new paradigm hypothesized to avoid reliance on executive function, which may compensate for damaged memory networks. OBJECTIVE: To assess the validity of the In-out-test in identifying prodromal Alzheimer's disease (PAD) in a clinical setting, by comparing this to the Free and Cued Selective Reminding Test (FCSRT) and cerebrospinal fluid biomarkers. METHODS: A cross-sectional study of 32 cognitively healthy, 32 MCI, and 30 progressive dementia subjects. All participants were given both the In-out-test and the FCSRT; 40 of them also received a lumbar puncture. RESULTS: Internal consistency was demonstrated using Cronbach Alpha (r = 0.81) and Inter-rater reliability with Kappa (k = 0.94). Intraclass correlation (ICC) for test-retest reliability: r = 0.57 (p = 0.57). ICC between the In-out-test and FCSRT r = 0.87 (p = 0.001). ICC between the In-out-test and Aß42 and P-tau/Aß42 for controls: 0.73 and 0.75, respectively; P-tau for MCI: 0.77 and total sample: 0.70; Aß42 for dementia: 0.71. All ICC measures between FCSRT and biomarkers were ≤0.264. AD diagnosis: In-out-test k = 0.71; FCSRT k = 0.49. PAD diagnosis (N = 35): In-out-test k = 0.69; FCSRT k = 0.44. CONCLUSIONS: The In-out-test detected prodromal AD with a higher degree of accuracy than a conventional hippocampal-based memory test. These results suggest that this new paradigm could be of value in clinical settings, predicting which patients with MCI will go on to develop AD.

"Lipid raft aging" in the human frontal cortex during nonpathological aging: gender influences and potential implications in Alzheimer's disease.

Lipid rafts are highly dynamic membrane domains featured by distinctive biochemical composition and physicochemical properties compared with the surrounding plasma membrane. These microstructures are associated not only with cellular signaling and communication in normal nerve cells but also with pathological processing of amyloid precursor protein in Alzheimer's disease. Using lipid rafts isolated from human frontal cortex in nondemented subjects aging 24 to 85 years, we demonstrate here that lipid structure of lipid rafts undergo significant alterations of specific lipid classes and phospholipid-bound fatty acids as brain cortex correlating with aging. Main changes affect levels of plasmalogens, polyunsaturated fatty acids (especially docosahexaenoic acid and arachidonic acid), total polar lipids (mainly phosphatidylinositol, sphingomyelin, sulfatides, and cerebrosides), and total neutral lipids (particularly cholesterol and sterol esters). Besides, relevant relationships between main fatty acids and/or lipid classes were altered in an age-related manner. This "lipid raft aging" exhibits clear gender differences and appear to be more pronounced in women than in men, especially in older (postmenopausal) women. The outcomes led us to conclude that human cortical lipid rafts are modified by aging in a gender-dependent fashion. Given the central role of bilayer lipid matrix in lipid rafts functionality and neuronal signaling, we hypothesize that these findings might underlie the higher prevalence of cognitive decline evolving toward Alzheimer's disease in postmenopausal women.

Estrogen Interactions With Lipid Rafts Related to Neuroprotection. Impact of Brain Ageing and Menopause.

Estrogens (E2) exert a plethora of neuroprotective actions against aged-associated brain diseases, including Alzheimer's disease (AD). Part of these actions takes place through binding to estrogen receptors (ER) embedded in signalosomes, where numerous signaling proteins are clustered. Signalosomes are preferentially located in lipid rafts which are dynamic membrane microstructures characterized by a peculiar lipid composition enriched in gangliosides, saturated fatty acids, cholesterol, and sphingolipids. Rapid E2 interactions with ER-related signalosomes appear to trigger intracellular signaling ultimately leading to the activation of molecular mechanisms against AD. We have previously observed that the reduction of E2 blood levels occurring during menopause induced disruption of ER-signalosomes at frontal cortical brain areas. These molecular changes may reduce neuronal protection activities, as similar ER signalosome derangements were observed in AD brains. The molecular impairments may be associated with changes in the lipid composition of lipid rafts observed in neurons during menopause and AD. These evidences indicate that the changes in lipid raft structure during aging may be at the basis of alterations in the activity of ER and other neuroprotective proteins integrated in these membrane microstructures. Moreover, E2 is a homeostatic modulator of lipid rafts. Recent work has pointed to this relevant aspect of E2 activity to preserve brain integrity, through mechanisms affecting lipid uptake and local biosynthesis in the brain. Some evidences have demonstrated that estrogens and the docosahexaenoic acid (DHA) exert synergistic effects to stabilize brain lipid matrix. DHA is essential to enhance molecular fluidity at the plasma membrane, promoting functional macromolecular interactions in signaling platforms. In support of this, DHA detriment in neuronal lipid rafts has been associated with the most common age-associated neuropathologies, namely AD and Parkinson disease. Altogether, these findings indicate that E2 may participate in brain preservation through a dual membrane-related mechanism. On the one hand, E2 interacting with ER related signalosomes may protect against neurotoxic insults. On the other hand, E2 may exert lipostatic actions to preserve lipid balance in neuronal membrane microdomains. The different aspects of the emerging multifunctional role of estrogens in membrane-related signalosomes will be discussed in this review.