Valutazione degli effetti di alcuni inquinanti su diverse specie non convenzionali con particolare attenzione alle tartarughe marine (Caretta Caretta)

Il presente lavoro raccoglie i risultati di diversi studi inerenti la valutazione della presenza di metalli pesanti e mettalloidi (arsenico, piombo, cadmio, cromo, selenio, rame, ferro, zinco, mercurio), in alcuni tessuti di animali selvatici (cinghiale, anatidi selvatici e tartarughe marine), dei loro effetti sugli stessi ed ancora della presenza di bio-indicatori di inquinamento da interferenti endocrini (vitellogenina) nel sangue e nel fegato di pesce siluro (Silurus glanis) e nel sangue di tartaruga marina (Caretta caretta). Quello che emerge dagli studi effettuati è che le specie utilizzate (cinghiali, tartarughe marine) risultano essere per l’ambiente in cui vivono un valido indicatore di inquinamento e che ad oggi la presenza di metalli pesanti è ancora rilevante se pur a livelli accettabili. In alcuni casi (studio metalli nel sangue di tartarughe marine), tuttavia, la mancanza di letteratura inerente all’argomento rende necessario l’approfondimento dello studio. Il passo successivo potrebbe essere la comparazione tra i livelli di piombo in campioni di sangue e quelli in tessuti provenienti dallo stesso animale. Dato il basso grado di mortalità presso il centro recupero di Lampedusa, l’attuazione di tale ricerca richiede comunque un periodo lungo per poter ottenere risultati concreti. Per quanto riguarda la valutazione della tossicità delle differenti tipologie di pallino da caccia si può affermare come si dimostri la pericolosità dei pallini di piombo per gli Anatidi a differenza della “innocuità” dell’acciaio ferroso. Sarebbe quindi importante considerare l’eventualità di utilizzare queste leghe anche nei nostri distretti venatori al fine di non procurare sofferenze inutili ad animali che per abitudini etologiche sono coinvolti in questo problema. Infine dalla ricerca sulla vitellogenina è emerso che la metodica da noi utilizzata si è dimostrata essere veloce, di semplice applicazione, economica e, almeno per il plasma, affidabile. Vista la sua “aspecificità”, in quanto basata su processi di precipitazione, è potenzialmente applicabile a tutte quelle specie per le quali non esistono sistemi immunoenzimatici di rilevamento, potenzialità già da noi confermata dal suo utilizzo su due specie ovipare diverse. Anche la metodica SDS-PAGE per l’identificazione della Vtg si è dimostrata funzionale anche se la macanza di anticorpi specifici per le specie studiate non ne garantisce la sua specificità. Il passo successivo sarebbe quello di verificare l’esistenza di cross-reattività tra anticorpi anti-Vtg di specie diverse. Tale studio è in via di attuazione, almeno per quanto riguarda le tartarughe marine, grazie alla collaborazione con il Centro de Investigaciones Biológicas del Noreste (La Paz, México) che ha permesso di utilizzare un anticorpo anti-VTG di Chelonia mydas agasizii (Sifuentes et al., 2006) per verificare l’esistenza di una sua eventuale cross-reattività nei confronti di VTG di altre specie come la Caretta caretta, che porterebbe di conseguenza al miglioramento delle tecniche di monitoraggio degli effetti di interferenti endocrini, di cui la VTG rappresenta un ottimo biomarker, sulle tartarughe marine

New Insights into the Hepcidin-Ferroportin Axis and Iron Homeostasis in iPSC-Derived Cardiomyocytes from Friedreich’s Ataxia Patient

Iron homeostasis in the cardiac tissue as well as the involvement of the hepcidin-ferroportin (HAMP-FPN) axis in this process and in cardiac functionality are not fully understood. Imbalance of iron homeostasis occurs in several cardiac diseases, including iron-overload cardiomyopathies such as Friedreich’s ataxia (FRDA, OMIM no. 229300), a hereditary neurodegenerative disorder. Exploiting the induced pluripotent stem cells (iPSCs) technology and the iPSC capacity to differentiate into specific cell types, we derived cardiomyocytes of a FRDA patient and of a healthy control subject in order to study the cardiac iron homeostasis and the HAMP-FPN axis. Both CTR and FRDA iPSCs-derived cardiomyocytes express cardiac differentiation markers; in addition, FRDA cardiomyocytes maintain the FRDA-like phenotype. We found that FRDA cardiomyocytes show an increase in the protein expression of HAMP and FPN. Moreover, immunofluorescence analysis revealed for the first time an unexpected nuclear localization of FPN in both CTR and FRDA cardiomyocytes. However, the amount of the nuclear FPN was less in FRDA cardiomyocytes than in controls. These and other data suggest that iron handling and the HAMP-FPN axis regulation in FRDA cardiac cells are hampered and that FPN may have new, still not fully understood, functions. These findings underline the complexity of the cardiac iron homeostasis

Effects of tocotrienol supplementation in Friedreich's ataxia: A model of oxidative stress pathology

Friedreich’s Ataxia (FRDA) is an autosomal recessive disorder characterized by impaired mitochondrial function, resulting in oxidative stress. In this study, we aimed at evaluating whether tocotrienol, a phytonutrient that diffuses easily in tissues with saturated fatty layers, could complement the current treatment with idebenone, a quinone analogue with antioxidant properties. Methods. Five young FRDA patients received a low dose tocotrienol supplementation (5 mg/kg/day), while not discontinuing idebenone treatment. Several oxidative stress markers and of biological parameters related to oxidative stress were evaluated at time of initiation of treatment and two and twelve months post-treatment. Some oxidative stress-related parameters and some inflammation indices were altered in FRDA patients taking idebenone alone and tended to normal values following tocotrienol supplementation; likewise, a cardiac magnetic resonance study showed some improvement following 1-year tocotrienol treatment. The pathway by which tocotrienol affects the Nrf2 modulation of hepcidin gene expression, a peptide involved in iron handling and in inflammatory responses, is viewed in the light of the disruption of the iron intracellular distribution and of the Nrf2 anergy characterizing FRDA. This research provides a suitable model to analyze the efficacy of therapeutic strategies able to counteract the excess free radicals in FRDA, and paves the way to long-term clinical studies

Non-Coding RNAs in the Transcriptional Network That Differentiates Skeletal Muscles of Sedentary from Long-Term Endurance- and Resistance-Trained Elderly

In a previous study, the whole transcriptome of the vastus lateralis muscle from sedentary elderly and from age-matched athletes with an exceptional record of high-intensity, life-long exercise training was compared—the two groups representing the two extremes on a physical activity scale. Exercise training enabled the skeletal muscle to counteract age-related sarcopenia by inducing a wide range of adaptations, sustained by the expression of protein-coding genes involved in energy handling, proteostasis, cytoskeletal organization, inflammation control, and cellular senescence. Building on the previous study, we examined here the network of non-coding RNAs participating in the orchestration of gene expression and identified differentially expressed micro- and long-non-coding RNAs and some of their possible targets and roles. Unsupervised hierarchical clustering analyses of all non-coding RNAs were able to discriminate between sedentary and trained individuals, regardless of the exercise typology. Validated targets of differentially expressed miRNA were grouped by KEGG analysis, which pointed to functional areas involved in cell cycle, cytoskeletal control, longevity, and many signaling pathways, including AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR), which had been shown to be pivotal in the modulation of the effects of high-intensity, life-long exercise training. The analysis of differentially expressed long-non-coding RNAs identified transcriptional networks, involving lncRNAs, miRNAs and mRNAs, affecting processes in line with the beneficial role of exercise training

Skeletal Muscle Gene Expression in Long-Term Endurance and Resistance Trained Elderly

Physical exercise is deemed the most efficient way of counteracting the age-related decline of skeletal muscle. Here we report a transcriptional study by next-generation sequencing of vastus lateralis biopsies from elderly with a life-long high-level training practice (n = 9) and from age-matched sedentary subjects (n = 5). Unsupervised mixture distribution analysis was able to correctly categorize trained and untrained subjects, whereas it failed to discriminate between individuals who underwent a prevalent endurance (n = 5) or a prevalent resistance (n = 4) training, thus showing that the training mode was not relevant for sarcopenia prevention. KEGG analysis of transcripts showed that physical exercise affected a high number of metabolic and signaling pathways, in particular those related to energy handling and mitochondrial biogenesis, where AMPK and AKT-mTOR signaling pathways are both active and balance each other, concurring to the establishment of an insulin-sensitive phenotype and to the maintenance of a functional muscle mass. Other pathways affected by exercise training increased the efficiency of the proteostatic mechanisms, consolidated the cytoskeletal organization, lowered the inflammation level, and contrasted cellular senescence. This study on extraordinary individuals who trained at high level for at least thirty years suggests that aging processes and exercise training travel the same paths in the opposite direction

A new ligand for the urotensin II receptor

The cyclic peptide human urotensin II (U-II) has been recently recognized as the endogenous ligand of an orphan GPCR, subsequently named the UT receptor. No synthetic ligands are available for investigating this novel peptide-receptor system. A novel UT receptor ligand, [Orn(8)]U-II, was synthesized and evaluated in calcium functional assays performed on HEK293 cells expressing the recombinant rat and human UT receptor and in the rat aorta bioassay. [Orn(8)]U-II behaves as a full agonist (pEC(50)≈8) at both human and rat UT receptors in the FlipR calcium assay eliciting similar maximal effects as the natural ligand U-II. On the contrary, in the rat aorta bioassay, [Orn(8)]U-II behaves as a competitive and selective antagonist (pA(2)=6.56) showing however a small but consistent residual agonist activity. It is therefore proposed that [Orn(8)]U-II is a partial agonist at UT receptors

Increased excitability in tat-transgenic mice: role of tat in HIV-related neurological disorders

HIV-1 associated neurocognitive disorders (HAND) are a major complication of HIV-1 infection. The mechanism(s) underlying HAND are not completely understood but, based on in vitro studies, the HIV-1 Tat protein may play an important role. In this study, the effect of prolonged exposure to endogenously produced Tat in the brain was investigated using a tat-transgenic (TT) mouse model constitutively expressing the HIV-1 tat gene. We found that stimulus-evoked glutamate exocytosis in the hippocampus and cortex was significantly increased in TT as compared with wild-type control (CC) mice, while GABA exocytosis was unchanged in the hippocampus and decreased in the cortex. This suggests that Tat generates a latent hyper-excitability state, which favor the detrimental effects of neurotoxic and/or excitotoxic agents. To challenge this idea, TT mice were tested for susceptibility to kainate-induced seizures and neurodegeneration, and found to exhibit significantly greater responses to the convulsant agent than CC mice. These results support the concept that constitutive expression of tat in the brain generates a latent excitatory state, which may increase the negative effects of damaging insults. These events may play a key role in the development of HAND