Attenuation of Autism-like Behaviors by an Anthocyanin-Rich Extract from Portuguese Blueberries via Microbiota-Gut-Brain Axis Modulation in a Valproic Acid Mouse Model.

Autism Spectrum Disorders (ASDs) are a group of neurodevelopmental pathologies whose current treatment is neither curative nor effective. Anthocyanins are naturally occurring compounds abundant in blueberries and in other red fruits which have been shown to be successful in the treatment of several neurological diseases, at least in in vitro and in vivo disease models. The aim of the present work was to study the ability of an anthocyanin-rich extract (ARE) obtained from Portuguese blueberries to alleviate autism-like symptoms in a valproic acid (VPA) mouse model of ASD and to get insights into the underlying molecular mechanisms of such benefits. Therefore, pregnant BALB/c females were treated subcutaneously with a single dose of VPA (500 mg/kg) or saline on gestational day 12.5. Male offspring mice were orally treated with the ARE from Portuguese blueberries (30 mg/kg/day) or the vehicle for three weeks, and further subjected to behavioral tests and biochemical analysis. Our data suggested that the ARE treatment alleviated autism-like behaviors in in utero VPA-exposed mice and, at the same time, decreased both neuroinflammation and gut inflammation, modulated the gut microbiota composition, increased serotonin levels in cerebral prefrontal cortex and gut, and reduced the synaptic dysfunction verified in autistic mice. Overall, our work suggests that anthocyanins extracted from Portuguese blueberries could constitute an effective strategy to ameliorate typical autistic behaviors through modulation of the microbiota-gut-brain axis.

Neuropeptide Y stimulates autophagy in hypothalamic neurons.

Aging is characterized by autophagy impairment that contributes to age-related disease aggravation. Moreover, it was described that the hypothalamus is a critical brain area for whole-body aging development and has impact on lifespan. Neuropeptide Y (NPY) is one of the major neuropeptides present in the hypothalamus, and it has been shown that, in aged animals, the hypothalamic NPY levels decrease. Because caloric restriction (CR) delays aging, at least in part, by stimulating autophagy, and also increases hypothalamic NPY levels, we hypothesized that NPY could have a relevant role on autophagy modulation in the hypothalamus. Therefore, the aim of this study was to investigate the role of NPY on autophagy in the hypothalamus. Using both hypothalamic neuronal in vitro models and mice overexpressing NPY in the hypothalamus, we observed that NPY stimulates autophagy in the hypothalamus. Mechanistically, in rodent hypothalamic neurons, NPY increases autophagy through the activation of NPY Y1 and Y5 receptors, and this effect is tightly associated with the concerted activation of PI3K, MEK/ERK, and PKA signaling pathways. Modulation of hypothalamic NPY levels may be considered a potential strategy to produce protective effects against hypothalamic impairments associated with age and to delay aging.

Lipodystrophy as a target to delay premature aging.

Lipodystrophy syndromes are rare diseases characterized by low levels and an abnormal distribution of adipose tissue, caused by diverse genetic or acquired causes. These conditions commonly exhibit metabolic complications, including insulin resistance, diabetes, hypertriglyceridemia, nonalcoholic fatty liver disease, and adipose tissue dysfunction. Moreover, genetic lipodystrophic laminopathies exhibit a premature aging phenotype, emphasizing the importance of restoring adipose tissue distribution and function. In this opinion, we discuss the relevance of adipose tissue reestablishment as a potential approach to alleviate premature aging and age-related complications in genetic lipodystrophy syndromes.

Ataxin-2 in the hypothalamus at the crossroads between metabolism and clock genes.

ATXN2 gene, encoding for ataxin-2, is located in a trait locus for obesity. Atxn2 knockout (KO) mice are obese and insulin resistant; however, the cause for this phenotype is still unknown. Moreover, several findings suggest ataxin-2 as a metabolic regulator, but the role of this protein in the hypothalamus was never studied before. The aim of this work was to understand if ataxin-2 modulation in the hypothalamus could play a role in metabolic regulation. Ataxin-2 was overexpressed/re-established in the hypothalamus of C57Bl6/Atxn2 KO mice fed either a chow or a high-fat diet (HFD). This delivery was achieved through stereotaxic injection of lentiviral vectors encoding for ataxin-2. We show, for the first time, that HFD decreases ataxin-2 levels in mouse hypothalamus and liver. Specific hypothalamic ataxin-2 overexpression prevents HFD-induced obesity and insulin resistance. Ataxin-2 re-establishment in Atxn2 KO mice improved metabolic dysfunction without changing body weight. Furthermore, we observed altered clock gene expression in Atxn2 KO that might be causative of metabolic dysfunction. Interestingly, ataxin-2 hypothalamic re-establishment rescued these circadian alterations. Thus, ataxin-2 in the hypothalamus is a determinant for weight, insulin sensitivity and clock gene expression. Ataxin-2's potential role in the circadian clock, through the regulation of clock genes, might be a relevant mechanism to regulate metabolism. Overall, this work shows hypothalamic ataxin-2 as a new player in metabolism regulation, which might contribute to the development of new strategies for metabolic disorders.

Ghrelin delays premature aging in Hutchinson-Gilford progeria syndrome.

Hutchinson-Gilford progeria syndrome (HGPS) is a rare and fatal genetic condition that arises from a single nucleotide alteration in the LMNA gene, leading to the production of a defective lamin A protein known as progerin. The accumulation of progerin accelerates the onset of a dramatic premature aging phenotype in children with HGPS, characterized by low body weight, lipodystrophy, metabolic dysfunction, skin, and musculoskeletal age-related dysfunctions. In most cases, these children die of age-related cardiovascular dysfunction by their early teenage years. The absence of effective treatments for HGPS underscores the critical need to explore novel safe therapeutic strategies. In this study, we show that treatment with the hormone ghrelin increases autophagy, decreases progerin levels, and alleviates other cellular hallmarks of premature aging in human HGPS fibroblasts. Additionally, using a HGPS mouse model (LmnaG609G/G609G mice), we demonstrate that ghrelin administration effectively rescues molecular and histopathological progeroid features, prevents progressive weight loss in later stages, reverses the lipodystrophic phenotype, and extends lifespan of these short-lived mice. Therefore, our findings uncover the potential of modulating ghrelin signaling offers new treatment targets and translational approaches that may improve outcomes and enhance the quality of life for patients with HGPS and other age-related pathologies.

PI3K/AKT/MTOR and ERK1/2-MAPK signaling pathways are involved in autophagy stimulation induced by caloric restriction or caloric restriction mimetics in cortical neurons.

Caloric restriction has been shown to robustly ameliorate age-related diseases and to prolong lifespan in several model organisms, and these beneficial effects are dependent on the stimulation of autophagy. Autophagy dysfunction contributes to the accumulation of altered macromolecules, and is a key mechanism of promoting aging and age-related disorders, as neurodegenerative ones. We have previously shown that caloric restriction (CR), and CR mimetics Neuropeptide Y (NPY) and ghrelin, stimulate autophagy in rat cortical neurons, however by unknown molecular mechanisms. Overall, we show that CR, NPY, and ghrelin stimulate autophagy through PI3K/AKT/MTOR inhibition and ERK1/2-MAPK activation. The knowledge of these kinases in autophagy regulation and the contribution to the understanding of molecular mechanism facilitates the discovery of more targeted therapeutic strategies to stimulate autophagy, which is relevant in the context of age-related disorders.

Neuropeptide Y Enhances Progerin Clearance and Ameliorates the Senescent Phenotype of Human Hutchinson-Gilford Progeria Syndrome Cells.

Hutchinson-Gilford progeria syndrome (HGPS, or classical progeria) is a rare genetic disorder, characterized by premature aging, and caused by a de novo point mutation (C608G) within the lamin A/C gene (LMNA), producing an abnormal lamin A protein, termed progerin. Accumulation of progerin causes nuclear abnormalities and cell cycle arrest ultimately leading to cellular senescence. Autophagy impairment is a hallmark of cellular aging, and the rescue of this proteostasis mechanism delays aging progression in HGPS cells. We have previously shown that the endogenous Neuropeptide Y (NPY) increases autophagy in hypothalamus, a brain area already identified as a central regulator of whole-body aging. We also showed that NPY mediates caloric restriction-induced autophagy. These results are in accordance with other studies suggesting that NPY may act as a caloric restriction mimetic and plays a role as a lifespan and aging regulator. The aim of the present study was, therefore, to investigate if NPY could delay HGPS premature aging phenotype. Herein, we report that NPY increases autophagic flux and progerin clearance in primary cultures of human dermal fibroblasts from HGPS patients. NPY also rescues nuclear morphology and decreases the number of dysmorphic nuclei, a hallmark of HGPS cells. In addition, NPY decreases other hallmarks of aging as DNA damage and cellular senescence. Altogether, these results show that NPY rescues several hallmarks of cellular aging in HGPS cells, suggesting that NPY can be considered a promising strategy to delay or block the premature aging of HGPS.

New BACE1 Chimeric Peptide Inhibitors Selectively Prevent AßPP-ß Cleavage Decreasing Amyloid-ß Production and Accumulation in Alzheimer's Disease Models.

BACKGROUND: A disease-modifying therapy for Alzheimer's disease (AD) is still an unmet clinical need. The formation of amyloid-ß (Aß) requires the initial cleavage of the amyloid-ß protein precursor (AßPP) by BACE1 (beta-site AßPP cleaving enzyme 1), which is a prime therapeutic target for AD. OBJECTIVE: We aimed to design and develop a selective BACE1 inhibitor suitable to AD treatment. METHODS: The new BACE1 inhibitors consist on a chimeric peptide including a sequence related to the human Swedish mutant form of AßPP (AßPPswe) conjugated with the TAT carrier that facilitates cell membrane permeation and the crossing of the blood-brain barrier. Additionally to the chimeric peptide in the L-form, we developed a D-retroinverso chimeric peptide. The latter strategy, never used with BACE1 inhibitors, is considered to favor a significantly higher half-life and lower immunogenicity. RESULTS: We found that both chimeric peptides inhibit recombinant BACE1 activity and decrease Aß40/42 production in Neuro-2a (N2A) cells expressing AßPPswe without inducing cytotoxicity. The intraperitoneal administration of these peptides to 3xTg-AD mice decreased plasma and brain Aß40/42 levels, as well as brain soluble AßPPß production. Also, a reduction of insoluble Aß was observed in the brain after chronic treatment. Noteworthy, the chimeric peptides selectively inhibited the AßPP-ß cleavage relatively to the proteolysis of other BACE1 substrates such as close homologue of L1 (CHL1) and seizure-related gene 6 (SEZ6). CONCLUSIONS: Overall these new BACE1 chimeric peptideshold promising potential as a selective disease-modifying therapy for AD.

Caloric restriction stimulates autophagy in rat cortical neurons through neuropeptide Y and ghrelin receptors activation.

Caloric restriction is an anti-aging intervention known to extend lifespan in several experimental models, at least in part, by stimulating autophagy. Caloric restriction increases neuropeptide Y (NPY) in the hypothalamus and plasma ghrelin, a peripheral gut hormone that acts in hypothalamus to modulate energy homeostasis. NPY and ghrelin have been shown to be neuroprotective in different brain areas and to induce several physiological modifications similar to those induced by caloric restriction. However, the effect of NPY and ghrelin in autophagy in cortical neurons is currently not known. Using a cell culture of rat cortical neurons we investigate the involvement of NPY and ghrelin in caloric restriction-induced autophagy. We observed that a caloric restriction mimetic cell culture medium stimulates autophagy in rat cortical neurons and NPY or ghrelin receptor antagonists blocked this effect. On the other hand, exogenous NPY or ghrelin stimulate autophagy in rat cortical neurons. Moreover, NPY mediates the stimulatory effect of ghrelin on autophagy in rat cortical neurons. Since autophagy impairment occurs in aging and age-related neurodegenerative diseases, NPY and ghrelin synergistic effect on autophagy stimulation may suggest a new strategy to delay aging process.