Low-copper diet as a preventive strategy for Alzheimer's disease

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Correction: Single nucleotide polymorphisms in the human ATP7B gene modify the properties of the ATP7B protein

Correction for 'Single nucleotide polymorphisms in the human ATP7B gene modify the properties of the ATP7B protein' by Courtney J. McCann et al., Metallomics, 2019, 11, 1128-1139

Ceruloplasmin/Transferrin Ratio Changes in Alzheimer's Disease

The link between iron and Alzheimer's disease (AD) has been mainly investigated with a focus on the local accumulation of this metal in specific areas of the brain that are critical for AD. In the present study, we have instead looked at systemic variations of markers of iron metabolism. We measured serum levels of iron, ceruloplasmin, and transferrin and calculated the transferrin saturation and the ceruloplasmin to transferrin ratio (Cp/Tf). Cp/Tf and transferrin saturation increased in AD patients. Cp/Tf ratios also correlated positively with peroxide levels and negatively with serum iron concentrations. Elevated values of ceruloplasmin, peroxides, and Cp/Tf inversely correlated with MMSE scores. Isolated medial temporal lobe atrophy positively correlated with Cp/Tf and negatively with serum iron. All these findings indicate that the local iron accumulation found in brain areas critical for AD should be viewed in the frame of iron systemic alterations

ATP7B Variants as Modulators of Copper Dyshomeostasis in Alzheimer's Disease

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Oxidative Stress Related to Iron Metabolism in Relapsing Remitting Multiple Sclerosis Patients With Low Disability

Oxidative status may play a role in chronic inflammation and neurodegeneration which are considered critical etiopathogenetic factors in Multiple Sclerosis (MS), both in the early phase of the disease and in the progressive one. The aim of this study is to explore oxidative status related to iron metabolism in peripheral blood of stable Relapsing-Remitting MS with low disability. We studied 60 Relapsing-Remitting MS patients (age 37.2 ± 9.06, EDSS median 1.0), and 40 healthy controls (age 40.3 ± 10.86). We measured total hydroperoxides (dROMs test) and Total Antioxidant Status (TAS), along with the iron metabolism biomarkers: Iron (Fe), ferritin (Ferr), transferrin (Tf), transferrin saturation (Tfsat), and ceruloplasmin (Cp) panel biomarkers [concentration (iCp) and enzymatic activity (eCp), copper (Cu), ceruloplasmin specific activity (eCp:iCp), copper to ceruloplasmin ratio (Cu:Cp), non-ceruloplasmin copper (nCp-Cu)]. We computed also the Cp:Tf ratio as an index of oxidative stress related to iron metabolism. We found lower TAS levels in MS patients than in healthy controls (CTRL) and normal reference level and higher dROMs and Cp:Tf ratio in MS than in healthy controls. Cp and Cu were higher in MS while biomarkers of iron metabolism were not different between patients and controls. Both in controls and MS, dROMs correlated with iCp (CTRL r = 0.821, p < 0.001; MS r = 0.775 p < 0.001) and eCp (CTRL r = 0.734, p < 0.001; MS r = 0.820 p < 0.001). Moreover, only in MS group iCp correlated negatively with Tfsat (r = -0.257, p = 0.047). Dividing MS patients in “untreated” group and “treated” group, we found a significant difference in Fe values [F(2, 97) = 10.136, p < 0.001]; in particular “MS untreated” showed higher mean values (mean = 114.5, SD = 39.37 μg/dL) than CTRL (mean 78.6, SD = 27.55 μg/dL p = 0.001) and “MS treated” (mean = 72.4, SD = 38.08 μg/dL; p < 0.001). Moreover, “MS untreated” showed significantly higher values of Cp:Tf (mean = 10.19, SD = 1.77∗10-2; p = 0.015), than CTRL (mean = 9.03, SD = 1.46 ∗10-2). These results suggest that chronic oxidative stress is relevant also in the remitting phase of the disease in patients with low disability and short disease duration. Therefore, treatment with antioxidants may be beneficial also in the early stage of the disease to preserve neuronal reserve

Strutture cristallografiche di complessi tra l’Acetilcolinesterasi da Torpedo Californica e quattro nuovi inibitori, potenziali farmaci per l’Alzheimer

E’ stato dimostrato che i sintomi del morbo di Alzheimer sono associati ad un deficit della funzione colinergica a livello celebrale e pertanto l’impiego di inibitori dell’acetilcolinesterasi (AChE) è ancora oggi considerato l’approccio farmacologico più promettente nella terapia sintomatica di questa malattia. Oggetto dello studio è stata la determinazione della struttura cristallografica della TcAChE in complesso con quattro nuovi inibitori, potenziali farmaci per l’Alzheimer. L’AChE è stata isolata e purificata dagli organi elettrici della Torpedo californica, e cristallizzata con il metodo dell’ “hanging drop”. I cristalli sono stati sottoposti alla procedura di “soaking”(incubazione) con quattro nuovi inibitori dell’AChE: la Ganstigmina (CHF-2819; Chiesi farmaceutici S.p.a., Parma, Italia) e l’N1,N8-bisnorcimserina (National Institute of Health, U.S.A.), entrambi inibitori carbammici e l’SPH1371e l’SPH-1373 (Sanochemia Pharmazeutica, Vienna, Austria) entrambi inibitori reversibili. I cristalli dei quattro complessi sono stati caratterizzati mediante un’analisi cristallografica utilizzando come sorgente di raggi X, la luce di Sincrotrone presso “Elettra”, Trieste. I dati di diffrazione sono stati elaborati utilizzando i programmi biocristallografici DENZO, SCALEPACK e CCP4. La struttura 3D è stata determinata mediante il metodo della sostituzione molecolare utilizzando il programma AMoRe. L’affinamento della struttura è stato eseguito utilizzando il programma CNS. Lo studio del complesso con la ganstigmina è stata dettato dal buon profilo farmacodinamico e farmacocinetico del nuovo inibitore. Nella struttura cristallografica si osserva che il gruppo carbammico è legato covalentemente alla serina catalitica, mentre la geneserolina, il gruppo uscente, non è presente in tutto l’enzima, confermando l’ipotesi dell’esistenza di una “porta di servizio” implicata nell’eliminazione dei prodotti di reazione. Questo risultato è stato già suggerito dall’analisi della struttura del complesso con il MF268, un altro inibitore carbammico. I risultati ottenuti hanno permesso di studiare da un punto di vista strutturale le interazioni esistenti tra il gruppo carbammico della ganstigmina ed il sito attivo. La risoluzione della struttura cristallografica ha permesso di studiare da un punto di vista strutturale le interazioni esistenti tra il gruppo carbammico della ganstigmina e la triade catalitica, in particolare l’His440. La lunga durata di azione di questo inibitore può essere attribuita ad una alterazione della funzionalità della triade catalitica. Questa osservazione ha permesso di studiare come l’acetilcolinesterasi possa subire delle modificazioni strutturali importanti ad opera degli inibitori carbammici. L’N1,N8-bisnorcimserina, derivato della fisostigmina è un inibitore ben tollerato in vivo, oltrepassa la barriera emato-encefalica e, saggiata sui roditori, ha rivelato un aumento delle risposte cognitive. Come nel caso del MF268 e della ganstigmina, ci si aspettava di trovare nel sito catalitico una densità elettronica residua attribuibile al gruppo paraisopropilfenilcarbammico legato covalentemente alla serina catalitica. Inaspettatamente, invece, all’interno della gola enzimatica è stata riscontrata una densità residua corrispondente al gruppo uscente, la nor-eserolina, mentre non è stato identificato il gruppo carbammico. Il risultato ottenuto si è rivelato di grande interesse in quanto per la prima volta è stata determinata la struttura della TcAChE in complesso con il gruppo uscente degli inibitori carbammici analoghi della fisostigmina. L’SPH-1371 e l’SPH-1373 sono derivati della galantamina , farmaco già in uso terapeutico con il nome di Reminyl®. Essi sono stati disegnati e sintetizzati con lo scopo di identificare un candidato che presenti migliori capacità inibitorie e proprietà farmacodinamiche e farmacocinetiche superiori a quelle della stessa galantamina. Le strutture cristallografiche dei complessi tra la TcAChE e queste due nuove molecole, hanno evidenziato un differente orientamento dei due gruppi funzionali sostituenti sull’azoto dell’anello tetraidroazepinico della galantamina, fornendo una base razionale per l’interpretazione delle loro rispettive proprietà farmacologiche. Il sostituente propilpiperidinico dell’SPH-1371, ha mostrato un orientamento nella gola catalitica dell’AChE del tutto inatteso e in disaccordo con le previsioni ottenute da studi di molecular modelling. La struttura del complesso con l’inibitore SPH-1373, ha mostrato che la catena alchilica, costituita da sei gruppi metilenici e funzionalizzata con un gruppo saccarinico, assume una conformazione estesa all’interno della gola enzimatica e presenta una interazione di tipo π-π con il Trp279 del Peripheral Anionic Site, situato all’ingresso della cavità enzimatica. Con l’analisi strutturale è stato possibile interpretare razionalmente i buoni valori delle attività inibitorie mostrati da questi due composti. Le strutture cristallografiche dei complessi TcAChE-inibitori presentate hanno permesso di razionalizzare le proprietà farmacologiche delle molecole utilizzate e nel loro complesso costituiscono una solida base di partenza per i successivi studi di molecular modelling e drug design.It has been demonstrated that the Alzheimer’s disease symptoms are connected with a deficit of cerebral cholinergic action. For this reason the use of AChE inhibitors it still considered promising in the symptomatic therapy of this disease. Object of this PhD thesis is the determination of crystallographic structures of complexes between TcAChE and four new inhibitors, examined as potential drugs in the symptomatic therapy of Alzheimer’s disease. The AChE was extracted and purified from Torpedo californica electric tissues; the crystallization conditions were explored using the hanging drop method. The native crystals were soaked with four different inhibitors of AChE: ganstigmine (CHF-2819; Chiesi farmaceutici S.p.a., Parma, Italy) and N1,N8-bisnorcymserine (National Institute of Health, U.S.A.), both carbammic inhibitors, as well as SPH-1371 and SPH-1373 (Sanochemia Pharmazeutica, Vienna, Austria), two reversible inhibitors. The X-ray diffraction data of the four crystal complexes, were collected at the XRD-1 beam line of the Italian Synchrotron facility ELETTRA, Trieste, and were processed with the biocrystallography software DENZO, SCALEPACK and CCP4. The crystal structures were solved by molecular replacement method, using the software AMoRe and were refined using the software CNS. The good pharmacodynamic profile of ganstigmine prompted us to determine the crystal structure of this inhibitor in complex with TcAChE. The structure revealed a residual electron density given by the ethylphenylcarbamoyl moiety, which is covalently bound to the catalytic serine. Geneseroline, the leaving group, is not retained in the anionic site, confirming the hypothesis of the existence of a back door implied by the elimination of reaction products. This had already been suggested in the analysis of the structure of the complex with MF268, another carbammic inhibitor. The crystal structure of this complex allowed us to study the interactions between the carbammic group of gangstigmine and the catalytic residues, in particular His440. The long duration of action of this inhibitor could be due to a modification of the functionality of the catalytic triad. This evidence highlights that AChE undergoes important structural modifications by carbammic inhibitors. The phisostigmine derivative N1,N8-bisnorcymserine is well-tolerated in vivo, enters the blood-brain barrier, and improves cognitive performance in rodents. In analogy to MF268 and ganstigmine, it was expected that the residual electron density found in the catalytic site was due to the paraisopropylphenylcarbammic moiety, covalently bound to the catalytic serine. Surprisingly, the residual density observed at the bottom of the enzymatic gorge, corresponds to the leaving group, the nor-eseroline. We were not able to locate the carbammic moiety anywhere in the enzyme. The particular interest of this structure lies in the fact that it is the first one which shows only the leaving group of carbammic inhibitors analogues of phisostigmine. SPH-1371 and SPH-1373 are both derivatives of galanthamine -a therapeutic drug commercialized as Reminyl®-. These compounds were designed and synthesized with the aim to obtain a molecule showing higher inhibition properties and a better pharmacodynamic and pharmacokinetic profile than galanthamine itself. The crystal structures of the complexes between TcAChE and these new molecules revealed that the functional groups linked to the nitrogen of the tetrahydroazepine ring of galanthamine, had different orientations. This explains the different pharmacologic data showed by the two molecules. The propylpiperidine group of SPH-1371 shows an unexpected orientation in the enzymatic gorge. This result is in contrast with the molecular modelling predictions. The higher activity shown by SPH-1371 over galanthamine appears to be caused by the rigidity of the tetracyclic structure (low entropic cost), by the fact that a larger volume of the gorge is occupied by the inhibitor itself, and by a number of moderate and weak interactions with the residues lining the enzymatic gorge. The crystal structure of the complex with SPH-1373 shows that the six –methylene- alkyl chain ending with the saccharine moiety, is fully extended in the enzymatic gorge and forms a π-π stacking with Trp279 of the Peripheral Anionic Site, located at the entrance of catalytic pocket. This added interaction explains the higher activity of SPH-1373 over galanthamine. The crystal structures of the complexes between TcAChE and these inhibitors allowed us to rationalize their pharmacologic properties and represent an important starting point for future molecular modelling and drug design studies

Nutrition in Post-Stroke Subjects during Rehabilitation

Correct and appropriate nutrition after a stroke insult appears to exert an essential influence on, and play a key role in, the recovery of patients [...

Pet Presence Can Reduce Anxiety in the Elderly: The Italian Experience during COVID-19 Lockdown Assessed by an Electronic Survey

The lockdown imposed in Italy due to the COVID-19 outbreak required restrictions that severely limited individual freedom to protect the population and reduce virus diffusion. This situation psychologically challenged the entire Italian population but mostly the elderly. The “Digital mental health approach” employs digital tools to evaluate and prevent increasing mental health problems. “Anonymous online electronic surveys” are digital tools that assess rates of mental health outcomes (using for example self-assessment/awareness tools). Immediately at the beginning of restrictions, we designed an electronic survey a) to remotely investigate the psychological impact of the lockdown and b) to compare the anxiety between pet owners and not-pet owners. A total of 3905 subjects filled out the survey; we focused our study on 781 (20%) elderly subjects. Dividing elderly patients between pet-owners (n = 405) and not-pet owners (n = 376), the pet owners showed a Zung scale score significantly lower in respect to the not-pet owners. We observed that, during the COVID-19 outbreak, the pet presence could have a positive effect on anxiety in the elderly subject. These results: (A) encourage the use of mobile technologies for the assessment of psychological disorders that can be promptly employed in emergencies such as the COVID-19 outbreak; (B) highlight the positive effect of pet interaction to mitigate the psychological distress in elderly people