Optimization of γ-PGA biosynthesis supported by synthetic biology and metabolic engineering strategies

Poly-γ-glutamate (γ-PGA) is a natural polymer composed by glutamic acid residues, synthesized by the pgs operon of Bacillus subtilis. γ-PGA has a wide range of applications as food, cosmetic and pharmaceutical additive. However, to increase its industrial attractiveness, it is necessary to cut production costs utilizing cost-competitive feedstocks for fermentation. A low-cost by-product that can be used as feedstock is raw glycerol, that accounts for 10% (w/w) of the total biodiesel production. To achieve cost-competitive γ-PGA production from glycerol a multifaceted approach has been set up that includes: 1) improvement of pgs expression; 2) accumulation of γ-PGA precursors by metabolic engineering; 3) enhancement of glycerol metabolism. 1) The strength of the pgs operon regulatory elements has been analysed both by a synthetic biology approach, exploiting the well-characterized expression operating unit (EOU) inserted in amyE, and by a classical in-locus transcriptional fusion. Results from the two settings will be compared. These data will be then used to finely tune pgs expression and optimize γ-PGA yield. To this end, an inducible pgs operon has been constructed. 2) A genome-scale metabolic model was used to identify suitable targets for enhancing central carbon pathway flux toward γ-PGA synthesis. The first two B. subtilis strains, engineered following this analysis, showed enhanced polymer production. Other target genes are currently under investigation. 3) B. subtilis tolerance to raw glycerol obtained from a biodiesel plant (from both vegetable and animal origin) was verified. Further investigations are underway to improve glycerol uptake and consumption

Doubly heterozygous LMNA and TTN mutations revealed by exome sequencing in a severe form of dilated cardiomyopathy

Familial dilated cardiomyopathy (DCM) is a heterogeneous disease; although 30 disease genes have been discovered, they explain only no more than half of all cases; in addition, the causes of intra-familial variability in DCM have remained largely unknown. In this study, we exploited the use of whole-exome sequencing (WES) to investigate the causes of clinical variability in an extended family with 14 affected subjects, four of whom showed particular severe manifestations of cardiomyopathy requiring heart transplantation in early adulthood. This analysis, followed by confirmative conventional sequencing, identified the mutation p.K219T in the lamin A/C gene in all 14 affected patients. An additional variant in the gene for titin, p.L4855F, was identified in the severely affected patients. The age for heart transplantation was substantially less for LMNA:p.K219T/TTN:p.L4855F double heterozygotes than that for LMNA:p.K219T single heterozygotes. Myocardial specimens of doubly heterozygote individuals showed increased nuclear length, sarcomeric disorganization, and myonuclear clustering compared with samples from single heterozygotes. In conclusion, our results show that WES can be used for the identification of causal and modifier variants in families with variable manifestations of DCM. In addition, they not only indicate that LMNA and TTN mutational status may be useful in this family for risk stratification in individuals at risk for DCM but also suggest titin as a modifier for DCM

Differences between physician and caregiver evaluations in Alzheimer’s disease

In Alzheimer’s disease (AD) differences are often reported between caregivers’ and physicians’ aims and evaluations. The aim of this study was to investigate the presence of discrepancies between the evaluations of caregivers and physicians. Two hundred and eighteen caregivers of outpatients diagnosed with mild-to-moderate AD, referred to our Alzheimer’s Disease Assessment Unit, entered the study. Caregiver burden was investigated using the Caregiver Burden Inventory (CBI) and the Neuropsychiatric Inventory Caregiver Distress Scale (NPIDS). Caregivers were also administered a four-point scale to assess quantitatively whether their expectations in relation to the efficacy of the pharmacological treatment had been satisfied and a scale exploring clinical global impression of change. Caregivers whose expectations had not been satisfied had significantly higher CBI and NPIDS scores and a lower level of education; moreover, caregivers tended to overestimate the degree of cognitive decline during the course of the disease. These results underline the importance of correct and constant communication with AD patients and their familie

The effects of alcohol on cognition in the elderly : from protection to neurodegeneration

The effects of chronic alcohol abuse on cognition are well known. Memory and executive functions appear to be the cognitive domains primarily impaired, and prefrontal and frontal damage is reported on neuroimaging studies both at micro- and macrostructural levels. Abstinence can partially reverse these alterations through mechanisms of neuroplasticity. Alcohol acts in a dose-dependent fashion, and a light-to-moderate consumption indeed has protective effects on cardiovascular risk factors and promotes anti-inflammatory and anti-oxidative processes. In the elderly on such a regimen, several epidemiological studies have reported a decreased risk of both coronary and cerebrovascular disease and of dementia. However, because of data heterogeneity and the presence of several confounding variables, further studies are needed to clarify these findings. In addition, the complexity of alcohol neurobiology (interaction of alcohol effects with genetic predisposition and environmental factors) and the occurrence of age-related changes should also be taken into account. As dementia, stroke and cardiovascular disease are the leading causes of mortality in older people in developed countries, a better knowledge of the mechanisms underlying the effects of alcohol intake may be helpful from the perspective not only of medical management but also of social health polic

REM and NREM sleep enactment behaviors in Parkinson's disease, Parkinson's disease dementia, and dementia with Lewy bodies

none8Background/objective: Nocturnal sleep enactment behaviors (SEBs) are common in patients affected by Parkinson's disease (PD), dementia associated with Parkinson's disease (PDD), and dementia with Lewy bodies (DLB). We investigated the occurrence and neurobiological significance of abnormal SEBs in the context of PD without cognitive decline compared to PDD/DLB patients. Methods: We evaluated a sample of 139 patients with PD, PDD, or DLB in a cross-sectional survey. One hundred and seventeen patients showing either no cognitive impairment (PD group) or meeting the diagnostic requirements for dementia (PDD/DLB group) underwent video-polysomnography. Seventy subjects (42 males) in whom a clear-cut diagnosis of abnormal sleep-related motor-behavioral episodes was possible were included in the final analysis. Results: SEBs consisting of RBD or occurring on arousal from NREM or REM sleep were globally more frequent in the dementia group (PDD/DLB) than in the PD group (p= 0.001), the difference being statistically significant for arousal-related episodes (p= 0.002), while a trend emerged for RBD (p= 0.07). Male sex, daytime sleepiness, higher motor impairment, and lower mini-mental score were significantly more frequent with the occurrence of abnormal sleep-related motor-behavioral episodes. Conclusion: SEBs in PD, PDD, and DLB may consist of RBD episodes or of arousal-related NREM and REM episodes. These latter are more frequent in patients with PDD/DLB and seem to be mainly related to more advanced stages of disease with a higher degree of cognitive decline. © 2012 Elsevier B.V.noneRatti P.-L.; Terzaghi M.; Minafra B.; Repetto A.; Pasotti C.; Zangaglia R.; Pacchetti C.; Manni R.Ratti, P. -L.; Terzaghi, M.; Minafra, B.; Repetto, A.; Pasotti, C.; Zangaglia, R.; Pacchetti, C.; Manni, R

Nanoscale phase separation in coated Ag nanoparticles

In this paper we report the structural investigation of cysteine and glutathione capped Ag nanoparticles (NPs) by means of transmission electron microscopy (TEM), synchrotron X-ray diffraction (XRD) and pair distribution function (PDF) analysis. The combined use of these probes allowed us to observe the presence of two crystal structures in the coated AgNPs, i.e., the cubic and the hexagonal crystal structures of Ag. In particular, it was possible to demonstrate that the coated AgNPs are a nanoscale phase separated system where the two phases coexist within the single grain. In addition, the relative bulk amount of the fcc and hcp phases has been estimated and a possible correlation with the capping agent propose

A multi-component, adaptive Working Memory Assessment Battery (WoMAB): validation and norms in an Italian population sample

BACKGROUND: Working memory (WM) abilities are frequently impaired in neurological disorders affecting fronto-parietal cortical/sub-cortical structures. WM deficits negatively influence interventional outcomes and everyday functioning. This study thus aimed at the following: (a) developing and standardizing an ecologically valid task for WM assessment ( Ice Cream Test, ICT); (b) validating and norming a novel WM test (Digit Ordering Test, DOT), as well as providing updated norms for digit span (DS) tasks, in an Italian population sample; (c) introducing a novel scoring procedure for measuring WM. METHODS: One-hundred and sixty-eight Italian healthy participants—73 male, 95 females; age: 48.4 ± 19.1 (18–86); education: 12.1 ± 4.8 (4–21)—underwent a thorough WM assessment—DOT, ICT, and both forward and backward DS tasks (FDS, BDS). The ICT requires participants to act as waiters who have to keep track of customers’ orders. For each task, WM and total (T) outcomes were computed, i.e., the number of elements in the longest sequence and that of recalled sequences, respectively. Norms were derived via the equivalent score (ES) method. RESULTS: DS ratios (DSRs) were computed for both WM/S and T outcomes on raw DS measures (BDS divided by FDS). Age and education significantly predicted all WM tasks; sex affected FDS and DSR-T scores (males > females). WM measures were highly internally related. DISCUSSION: The present work provides Italian practitioners with a normatively updated, multi-component, adaptive battery for WM assessment (WoMAB) as well as with novel outcomes which capture different WM facets—WM capacity and attentive monitoring abilities

Antibacterial action of Glutathione coated Silver Nanoparticles against Gram positive and Gram negative bacteria

A study is presented on the bactericidal effect and its mechanism, of biomimetically-coated silver nanoparticles when interacting with E.Coli and S.Aureu

Assessing mood and cognitive functioning in acute stroke: clinical usability of a Visual Analogue Mood Scale (VAMS)

BACKGROUND: Patients suffering from stroke in the acute/post-acute phases often present with depressive mood — which negatively impacts on patients’ prognosis. However, psychometric evaluation of mood in acute stroke patients may be challenging due to cognitive deficits. Tools investigating emotional states via a vertical analogue line may overcome language/visuo-spatial disorders. This study thus aimed at (a) investigating the clinical usability of a Visual Analogue Mood Scale (VAMS) in acute stroke patients and (b) investigating the interplay between mood and cognition in this population. METHODS: Forty-one acute stroke patients were compared to 41 age-, education- and sex-matched healthy participants (HPs) on the VAMS and on cognitive measures (mental performance in acute stroke, MEPS). A control line bisection (LB) task was administered to control for potential visuo-spatial deficits in patients. RESULTS: Patients reported higher depression levels than HPs (lower VAMS scores); this between-group difference stayed significant when covarying for LB scores. MEPS scores discriminated patients from HPs; among cognitive measures, only the Clock drawing test (CDT) was positively associated with VAMS scores. Lesion side did not affect patients’ mood state; however, disease duration was inversely related to VAMS scores. DISCUSSION: The VAMS proved to be a suitable tool for assessing mood in acute stroke patients, as being independent from post-stroke cognitive sequelae. The CDT might represent an adequate measure of depression-induced, post-stroke cognitive efficiency decrease. Mood disorders might occur and thus should be adequately addressed also in post-acute phases — likely due to longer hospitalization times and regression of anosognosic features

Functionalization of nano-objects with Glutathione

Coating of Ag nanoparticles with glutathione results in a pH-controllable precipitation of the product. This is soluble at physiological ph (7-7.5, can be used in in-vivo like conditions and exerts a microbicidal effect against E.Col