18 alpha-Glycyrrhetinic Acid Proteasome Activator Decelerates Aging and Alzheimer's Disease Progression in Caenorhabditis elegans and Neuronal Cultures

Aims: Proteasomes are constituents of the cellular proteolytic networks that maintain protein homeostasis through regulated proteolysis of normal and abnormal (in any way) proteins. Genetically mediated proteasome activation in multicellular organisms has been shown to promote longevity and to exert protein antiaggregation activity. In this study, we investigate whether compound-mediated proteasome activation is feasible in a multicellular organism and we dissect the effects of such approach in aging and Alzheimer's disease (AD) progression. Results: Feeding of wild-type Caenorhabditis elegans with 18 alpha-glycyrrhetinic acid (18 alpha-GA; a previously shown proteasome activator in cell culture) results in enhanced levels of proteasome activities that lead to a skinhead-1- and proteasomeactivation-dependent life span extension. The elevated proteasome function confers lower paralysis rates in various AD nematode models accompanied by decreased A beta deposits, thus ultimately decelerating the progression of AD phenotype. More importantly, similar positive results are also delivered when human and murine cells of nervous origin are subjected to 18 alpha-GA treatment. Innovation: This is the first report of the use of 18 alpha-GA, a diet-derived compound as prolongevity and antiaggregation factor in the context of a multicellular organism. Conclusion: Our results suggest that proteasome activation with downstream positive outcomes on aging and AD, an aggregation-related disease, is feasible in a nongenetic manipulation manner in a multicellular organism. Moreover, they unveil the need for identification of antiaging and antiamyloidogenic compounds among the nutrients found in our normal diet.Peer reviewe

Chemical analysis of Greek pollen - Antioxidant, antimicrobial and proteasome activation properties

<p>Abstract</p> <p>Background</p> <p>Pollen is a bee-product known for its medical properties from ancient times. In our days is increasingly used as health food supplement and especially as a tonic primarily with appeal to the elderly to ameliorate the effects of ageing. In order to evaluate the chemical composition and the biological activity of Greek pollen which has never been studied before, one sample with identified botanical origin from sixteen different common plant taxa of Greece has been evaluated.</p> <p>Results</p> <p>Three different extracts of the studied sample of Greek pollen, have been tested, in whether could induce proteasome activities in human fibroblasts. The water extract was found to induce a highly proteasome activity, showing interesting antioxidant properties. Due to this activity the aqueous extract was further subjected to chemical analysis and seven flavonoids have been isolated and identified by modern spectral means. From the methanolic extract, sugars, lipid acids, phenolic acids and their esters have been also identified, which mainly participate to the biosynthetic pathway of pollen phenolics. The total phenolics were estimated with the Folin-Ciocalteau reagent and the total antioxidant activity was determined by the DPPH method while the extracts and the isolated compounds were also tested for their antimicrobial activity by the dilution technique.</p> <p>Conclusions</p> <p>The Greek pollen is rich in flavonoids and phenolic acids which indicate the observed free radical scavenging activity, the effects of pollen on human fibroblasts and the interesting antimicrobial profile.</p

Disparate habitual physical activity and dietary intake profiles of elderly men with low and elevated systemic inflammation

The development of chronic, low-grade systemic inflammation in the elderly (inflammaging) has been associated with increased incidence of chronic diseases, geriatric syndromes, and functional impairments. The aim of this study was to examine differences in habitual physical activity (PA), dietary intake patterns, and musculoskeletal performance among community-dwelling elderly men with low and elevated systemic inflammation. Nonsarcopenic older men free of chronic diseases were grouped as ‘low’ (LSI: n = 17; 68.2 ± 2.6 years; hs-CRP: 1 mg/L) systemic inflammation according to their serum levels of high-sensitivity CRP (hs-CRP). All participants were assessed for body composition via Dual Emission X-ray Absorptiometry (DEXA), physical performance using the Short Physical Performance Battery (SPPB) and handgrip strength, daily PA using accelerometry, and daily macro- and micronutrient intake. ESI was characterized by a 2-fold greater hs-CRP value than LSI (p < 0.01). The two groups were comparable in terms of body composition, but LSI displayed higher physical performance (p < 0.05), daily PA (step count/day and time at moderate-to-vigorous PA (MVPA) were greater by 30% and 42%, respectively, p < 0.05), and daily intake of the antioxidant vitamins A (6590.7 vs. 4701.8 IU/day, p < 0.05), C (120.0 vs. 77.3 mg/day, p < 0.05), and E (10.0 vs. 7.5 mg/day, p < 0.05) compared to ESI. Moreover, daily intake of vitamin A was inversely correlated with levels of hs-CRP (r = −0.39, p = 0.035). These results provide evidence that elderly men characterized by low levels of systemic inflammation are more physically active, spend more time in MVPA, and receive higher amounts of antioxidant vitamins compared to those with increased systemic inflammation

Bacterial production and direct functional screening of expanded molecular libraries for discovering inhibitors of protein aggregation.

Protein misfolding and aggregation are associated with a many human disorders, including Alzheimer's and Parkinson's diseases. Toward increasing the effectiveness of early-stage drug discovery for these conditions, we report a bacterial platform that enables the biosynthesis of molecular libraries with expanded diversities and their direct functional screening for discovering protein aggregation inhibitors. We illustrate this approach by performing, what is to our knowledge, the largest functional screen of small-size molecular entities described to date. We generated a combinatorial library of ~200 million drug-like, cyclic peptides and rapidly screened it for aggregation inhibitors against the amyloid-β peptide (Aβ42), linked to Alzheimer's disease. Through this procedure, we identified more than 400 macrocyclic compounds that efficiently reduce Aβ42 aggregation and toxicity in vitro and in vivo. Finally, we applied a combination of deep sequencing and mutagenesis analyses to demonstrate how this system can rapidly determine structure-activity relationships and define consensus motifs required for bioactivity

Reduced proteasome activity in the aging brain results in ribosome stoichiometry loss and aggregation.

A progressive loss of protein homeostasis is characteristic of aging and a driver of neurodegeneration. To investigate this process quantitatively, we characterized proteome dynamics during brain aging in the short-lived vertebrate Nothobranchius furzeri combining transcriptomics and proteomics. We detected a progressive reduction in the correlation between protein and mRNA, mainly due to post-transcriptional mechanisms that account for over 40% of the age-regulated proteins. These changes cause a progressive loss of stoichiometry in several protein complexes, including ribosomes, which show impaired assembly/disassembly and are enriched in protein aggregates in old brains. Mechanistically, we show that reduction of proteasome activity is an early event during brain aging and is sufficient to induce proteomic signatures of aging and loss of stoichiometry in vivo. Using longitudinal transcriptomic data, we show that the magnitude of early life decline in proteasome levels is a major risk factor for mortality. Our work defines causative events in the aging process that can be targeted to prevent loss of protein homeostasis and delay the onset of age-related neurodegeneration

European contribution to the study of ROS: A summary of the findings and prospects for the future from the COST action BM1203 (EU-ROS).

The European Cooperation in Science and Technology (COST) provides an ideal framework to establish multi-disciplinary research networks. COST Action BM1203 (EU-ROS) represents a consortium of researchers from different disciplines who are dedicated to providing new insights and tools for better understanding redox biology and medicine and, in the long run, to finding new therapeutic strategies to target dysregulated redox processes in various diseases. This report highlights the major achievements of EU-ROS as well as research updates and new perspectives arising from its members. The EU-ROS consortium comprised more than 140 active members who worked together for four years on the topics briefly described below. The formation of reactive oxygen and nitrogen species (RONS) is an established hallmark of our aerobic environment and metabolism but RONS also act as messengers via redox regulation of essential cellular processes. The fact that many diseases have been found to be associated with oxidative stress established the theory of oxidative stress as a trigger of diseases that can be corrected by antioxidant therapy. However, while experimental studies support this thesis, clinical studies still generate controversial results, due to complex pathophysiology of oxidative stress in humans. For future improvement of antioxidant therapy and better understanding of redox-associated disease progression detailed knowledge on the sources and targets of RONS formation and discrimination of their detrimental or beneficial roles is required. In order to advance this important area of biology and medicine, highly synergistic approaches combining a variety of diverse and contrasting disciplines are needed.The EU-ROS consortium (COST Action BM1203) was supported by the European Cooperation in Science and Technology (COST). The present overview represents the final Action dissemination summarizing the major achievements of COST Action BM1203 (EU-ROS) as well as research news and personal views of its members. Some authors were also supported by COST Actions BM1005 (ENOG) and BM1307 (PROTEOSTASIS), as well as funding from the European Commission FP7 and H2020 programmes, and several national funding agencies

Study of molecular mechanisms of regulation of ageing and longevity

The development of multicellular organisms is characterized by irreversible changes of their anatomy, their physiology and their ability of adaptation to the environment. These changes, that can either be gradual or not, increase the possibility of death. The cascade of these developmental stages, with death being the last stage, is called ageing. It is a ubiquitous phenomenon that appears in all organisms. During the progression of ageing, changes in the morphology, the physiology and the molecular characteristics of the cells occur. Degradation and more precisely, the proteolytic mechanisms of the cell are playing a fundamental role in the maintenance of cellular homeostasis. These mechanisms are subjected to changes during ageing. The proteasome is the main cellular proteolytic machinery. In humans, the proteasome is composed by 14 α-regulatory subunits and 14 β-catalytic subunits (α7β7β7α7). It is responsible for the degradation of damaged proteins (unfolded, oxidized proteins etc) as well as, for the degradation of normal proteins that need to be proteolysed. However, it is not clear what is happening in the mechanism of the proteasome during ageing and its progression. The aim of this thesis is the study of the basic cellular proteolytic system, the proteasome, along with its expression profile and its role regarding its function, during the progression of ageing and longevity, in vivo and in vitro. We have shown that the protein expression levels of the proteasome subunits along with the proteasome function (studied mainly through the specific proteasome activities) are altered. These alterations may be related to the in vivo ageing and its progression. By using the model of cells from healthy donors of different ages (human fibroblasts derived from healthy donors of different ages, including centenarians), it became obvious that the RNA levels and the protein expression levels of several proteasome subunits (mainly α-regulatory and β-catalytic subunits of the 20S complex), along with the proteasome activities, were found decreased with donors age (18, 28 and 80 years old donors). The decreased proteasome activities of the cells derived from older donors were directly linked to the high levels of oxidised proteins exhibited by those cells. Given the fact that in the ageing research, healthy centenarians represent the best characterised living examples of successful ageing, fibroblasts derived from healthy centenarians were included in this study. The RNA and protein expression levels of the examined proteasome subunits in centenarians cultures were found similar to the levels expressed by the young donors cultures (28 years old) rather than the old donors cultures (80 years old). This profile is also maintained when the levels of proteasome activities of these cells are examined. Given the fact that the best model of studying ageing in vitro is represented by human normal cells that senesce in vitro, fibroblasts derived from healthy adult donors of different ages were cultured until the point of irreversible cease of proliferation (irreversible growth arrest) and their proteasome profile was studied. The RNA and protein expression levels and simultaneously, their levels of proteasome activities exhibited a decrease along with the in vitro ageing of the cells. .............................................................Η αναπτυξιακή πορεία όλων των πολυκύτταρων οργανισμών περιλαμβάνει διάφορα στάδια, τα οποία χαρακτηρίζονται από μη αντιστρεπτές μεταβολές, στην ανατομία, στη φυσιολογία αλλά και στην προσαρμοστικότητα αυτών, οι οποίες αυξάνουν την πιθανότητα θανάτου. Οι μεταβολές που υφίσταται κάθε οργανισμός μπορεί να είναι απότομες ή και βαθμιαίες. Το φαινόμενο αυτό της αλληλοδιαδοχής διαφόρων αναπτυξιακών σταδίων, με τελικό στάδιο το θάνατο, καλείται γήρανση. Πρόκειται για καθολικό φαινόμενο που εμφανίζεται σε όλους τους οργανισμούς, κατά την εξέλιξη του οποίου, εμφανίζονται διάφορες αλλαγές στη μορφολογία, τη φυσιολογία και τα μοριακά χαρακτηριστικά των δομικών λίθων των οργανισμών, τα κύτταρα. Η αποικοδόμηση και οι πρωτεολυτικοί μηχανισμοί του κυττάρου αποτελούν συστήματα που παίζουν βασικό ρόλο στη διατήρηση της κυτταρικής ομοιοστασίας και που υφίστανται τροποποιήσεις κατά την εξέλιξη του φαινομένου της γήρανσης. Το πρωτεόσωμα αποτελεί τον βασικό κυτταρικό πρωτεολυτικό μηχανισμό. Στα ανθρώπινα κύτταρα, το πρωτεόσωμα αποτελείται από 14 α-ρυθμιστικές υπομονάδες και 14 β-καταλυτικές υπομονάδες σε διάταξη α7β7β7α7. Είναι υπέυθυνο για την αποικοδόμηση τόσο κατεστραμμένων πρωτεϊνών (αποδιαταγμένων, οξειδωμένων κλπ), όσο και φυσιολογικών πρωτεϊνών που πρέπει να πρωτεολυθούν. Παρόλο ότι υπάρχουν πολλά δεδομένα τόσο για τη γήρανση όσο και για το πρωτεόσωμα, δεν είναι πλήρως γνωστό το τι συμβαίνει στον μηχανισμό αυτού κατά τη γήρανση και την εξέλιξή της. Σκοπός της διδακτορικής αυτής διατριβής είναι η μελέτη του βασικού πρωτεολυτικού συστήματος του κυττάρου, του πρωτεοσώματος και η συσχέτιση της έκφρασης και δράσης αυτού με το φαινόμενο της γήρανσης και της μακροβιότητας στον άνθρωπο, in vivo και in vitro. Δείχθηκε πως κατά την in vivo γήρανση, το πρωτεόσωμα υφίσταται αλλαγές τόσο σε επίπεδο πρωτεϊνικής έκφρασης των υπομονάδων του όσο και σε επίπεδο λειτουργικότητας αυτού (μελετώντας τις πρωτεολυτικές του ενεργότητες). Οι μεταβολές αυτές μπορούν να σχετισθούν με την in vivo γήρανση και την εξέλιξη αυτής. Χρησιμοποιώντας το μοντέλο των δοτών διαφορετικών ηλικιών (ανθρώπινοι ινοβλάστες από υγιείς δότες διαφορετικών ηλικιών), έγινε φανερό πως μεταβαίνοντας από κύτταρα δοτών μικρής ηλικίας (18, 28 ετών) σε κύτταρα δοτών μεγάλης ηλικίας (80 ετών), παρατηρείται μείωση των επιπέδων, τόσο RNA όσο και πρωτεϊνικής έκφρασης, των διαφόρων πρωτεοσωμικών υπομονάδων (η μελέτη έγινε με έμφαση στις υπομονάδες του 20S συμπλόκου, τόσο α-ρυθμιστικές όσο και β-καταλυτικές), όσο και των πρωτεοσωμικών ενεργοτήτων αυτού. Οι μειωμένες ενεργότητες του πρωτεοσώματος στους ινοβλάστες ατόμων μεγάλης ηλικίας έχουν άμεσο αντίκτυπο στα παρατηρούμενα επίπεδα οξειδωμένων κυτταρικών πρωτεϊνών, τα οποία παρουσιάζονται αυξημένα στα κύτταρα αυτά. Δεδομένου ότι στο πεδίο της γήρανσης, οι υγιείς αιωνόβιοι αποτελούν χαρακτηριστικά παραδείγματα «επιτυχούς» γήρανσης, συμπεριλήφθησαν στην παραπάνω μελέτη και ινοβλάστες από ανάλογους δότες. Τα επίπεδα RNA και πρωτεϊνικής έκφρασης των διαφόρων πρωτεοσωμικών υπομονάδων αυτών των κυττάρων, βρέθηκαν παρόμοια με αυτά που παρατηρούνται στα κύτταρα δοτών νεαρής ηλικίας (28 ετών) και όχι δοτών μεγαλύτερης ηλικίας (80 ετών). ...............................................................

Redox regulation of proteasome function

Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS) were initially regarded mainly as metabolic by-products with damaging properties. Over the last decade, our understanding of their role in metabolism was drastically changed and they were recognized as essential mediators in cellular signaling cascades, as well as modulators of biochemical pathways. Proteostasis is highly affected by the various levels of intracellular and extracellular free radicals with either mild or severe outcomes. As part of the proteostatic network, the proteasome system is equally affected by redox alterations. This short review summarizes the effects of oxidative stress on proteasome status while it also recapitulates conditions and processes where redox alterations signal changes to proteasome expression, assembly and function

Quest for Bioactive Compounds in Our Diet with Anti-Ageing and Anti-Aggregation Properties

Ageing is a complex process affected by both genetic and environmental factors, characterized by a gradual failure of functionality, reduced stress response and resistance, leading to enhanced probability for age-related diseases and mortality. During the last decades, natural compounds have attracted the attention of researchers in the quest of bioactive phytochemicals with anti-ageing properties. For a few of these compounds an extra advantage appears; many of them have been shown to decelerate the progression of age-related diseases with emphasis on aggregation-related diseases. Using the nematode Caenorhabditis elegans along with the replicative senescence model of human primary fibroblasts, we have identified compounds that are part of our diet with anti-oxidation, anti-ageing and anti-aggregation activities. Some of the identified compounds promote their anti-ageing activity through activation of the proteasome, others through the activation of Nrf2 transcription factor, while others through inhibition of glucose transporters (GLUTs). Our work identifies new bioactive compounds with anti-ageing and/or anti-aggregation properties or reveals additional beneficial properties on already known bioactive compounds

Neuron-specific proteasome activation exerts cell non-autonomous protection against amyloid-beta (Aβ) proteotoxicity in Caenorhabditis elegans

Proteostasis reinforcement is a promising approach in the design of therapeutic interventions against proteinopathies, including Alzheimer's disease. Understanding how and which parts of the proteostasis network should be enhanced is crucial in developing efficient therapeutic strategies. The ability of specific tissues to induce proteostatic responses in distal ones (cell non-autonomous regulation of proteostasis) is attracting interest. Although the proteasome is a major protein degradation node, nothing is known on its cell non-autonomous regulation. We show that proteasome activation in the nervous system can enhance the proteasome activity in the muscle of Caenorhabditis elegans. Mechanistically, this communication depends on Small Clear Vesicles, with glutamate as one of the neurotransmitters required for the distal regulation. More importantly, we demonstrate that this cell non-autonomous proteasome activation is translated into efficient prevention of amyloid-beta (Αβ)-mediated proteotoxic effects in the muscle of C. elegans but notably not to resistance against oxidative stress. Our in vivo data establish a mechanistic link between neuronal proteasome reinforcement and decreased Aβ proteotoxicity in the muscle. The identified distal communication may have serious implications in the design of therapeutic strategies based on tissue-specific proteasome manipulation