Redistribution of DAT/α-synuclein complexes visualized by “in situ” proximity ligation assay in transgenic mice modelling early Parkinson’s disease

Alpha-synuclein, the major component of Lewy bodies, is thought to play a central role in the onset of synaptic dysfunctions in Parkinson's disease (PD). In particular, α-synuclein may affect dopaminergic neuron function as it interacts with a key protein modulating dopamine (DA) content at the synapse: the DA transporter (DAT). Indeed, recent evidence from our "in vitro" studies showed that α-synuclein aggregation decreases the expression and membrane trafficking of the DAT as the DAT is retained into α-synuclein-immunopositive inclusions. This notwithstanding, "in vivo" studies on PD animal models investigating whether DAT distribution is altered by the pathological overexpression and aggregation of α-synuclein are missing. By using the proximity ligation assay, a technique which allows the "in situ" visualization of protein-protein interactions, we studied the occurrence of alterations in the distribution of DAT/α-synuclein complexes in the SYN120 transgenic mouse model, showing insoluble α-synuclein aggregates into dopaminergic neurons of the nigrostriatal system, reduced striatal DA levels and an altered distribution of synaptic proteins in the striatum. We found that DAT/α-synuclein complexes were markedly redistributed in the striatum and substantia nigra of SYN120 mice. These alterations were accompanied by a significant increase of DAT striatal levels in transgenic animals when compared to wild type littermates. Our data indicate that, in the early pathogenesis of PD, α-synuclein acts as a fine modulator of the dopaminergic synapse by regulating the subcellular distribution of key proteins such as the DAT

Multipoint temperature monitoring of microwave thermal ablation in bones through fiber bragg grating sensor arrays

Bones are a frequent site of metastases that cause intolerable cancer-related pain in 90% of patients, making their quality of life poor. In this scenario, being able to treat bone oncology patients by means of minimally invasive techniques can be crucial to avoid surgery-related risks and decrease hospitalization times. The use of microwave ablation (MWA) is gaining broad clinical acceptance to treat bone tumors. It is worth investigating temperature variations in bone tissue undergoing MWA because the clinical outcomes can be inferred from this parameter. Several feasibility studies have been performed, but an experimental analysis of the temperature trends reached into the bone during the MWA has not yet been assessed. In this work, a multi-point temperature study along the bone structure during such treatment is presented. The study has been carried out on ex vivo bovine femur and tibia, subjected to MWA. An overall of 40 measurement points covering a large sensing area was obtained for each configuration. Temperature monitoring was performed by using 40 fiber Bragg grating (FBGs) sensors (four arrays each housing 10 FBGs), inserted into the bones at specific distances to the microwave antenna. As result, the ability of this experimental multi-point monitoring approach in tracking temperature variations within bone tissue during MWA treatments was shown. This study lays the foundations for the design of a novel approach to study the effects of MWA on bone tumors. As consequence, the MWA treatment settings could be optimized in order to maximize the treatment effects of such a promising clinical application, but also customized for the specific tumor and patient

A multi-parametric wearable system to monitor neck movements and respiratory frequency of computer workers

Musculoskeletal disorders are the most common form of occupational ill-health. Neck pain is one of the most prevalent musculoskeletal disorders experienced by computer workers. Wrong postural habits and non-compliance of the workstation to ergonomics guidelines are the leading causes of neck pain. These factors may also alter respiratory functions. Health and safety interventions can reduce neck pain and, more generally, the symptoms of musculoskeletal disorders and reduce the consequent economic burden. In this work, a multi-parametric wearable system based on two fiber Bragg grating sensors is proposed for monitoring neck movements and breathing activity of computer workers. The sensing elements were positioned on the neck, in the frontal and sagittal planes, to monitor: (i) flexion-extension and axial rotation repetitions, and (ii) respiratory frequency. In this pilot study, five volunteers were enrolled and performed five repetitions of both flexion-extension and axial rotation, and ten breaths of both quite breathing and tachypnea. Results showed the good performances of the proposed system in monitoring the aforementioned parameters when compared to optical reference systems. The wearable system is able to well-match the trend in time of the neck movements (both flexion-extension and axial rotation) and to estimate mean and breath-by-breath respiratory frequency values with percentage errors ≤6.09% and ≤1.90%, during quiet breathing and tachypnea, respectively

The proximity ligation assay: an high throughput technique for protein analysis in neuroscience

In recent years, the ability to investigate proteins in cell cultures and tissue samples has expanded in order to offer profound insights into the dynamic interplay of molecules that occur within the cellular environment and their alterations in various disorders including neurodegenerative diseases. The proximity ligation assay (PLA) is a sensitive and specific technique that use sets of proximity probes to induce the formation of circular DNA strands when the probes are bound in close proximity. The DNA circles serve as templates for amplification reactions allowing the localized detection of individual, endogenous, interacting proteins in cells and tissue. Here, we describe the use of different variants of the PLA approach to analyze protein expression, post-translational modification and interaction in both liquid and tissue or cell culture samples

Alpha-synuclein synaptic pathology and its implications in the development of novel therapeutic approaches to cure Parkinson’s disease

Parkinson's disease (PD) is characterized by a progressive loss of dopamine (DA) neurons of the nigrostriatal system and by the presence of Lewy bodies (LB), proteinaceous inclusions mainly composed of filamentous alpha-synuclein aggregates. Alpha-synuclein is a natively unfolded protein which plays a central role in the control of dopaminergic neuronal functions and which is thought to be critically implicated in PD pathophysiology. Indeed, besides the fact that alpha-synuclein is the main protein component of LB, genetic studies showed that mutations and multiplications of the alpha-synuclein gene are responsible for the onset of familial forms of PD. A large body of evidence indicates that alpha-synuclein pathology at dopaminergic synapses may underlie the onset of neuronal cell dysfunction and degeneration in the PD brain. Thus, since the available therapeutic approaches to cure this disease are still limited, we hypothesized that the analysis of the alpha-synudein synaptic proteome/lipidome may represent a tool to identify novel potential therapeutic targets to cure this disorder. We thus performed a critical review of studies describing alpha-synuclein pathophysiology at synaptic sites in experimental models of PD and in this paper we outline the most relevant findings regarding the specific modulatory effects exerted by alpha-synuclein in the control of synaptic functions in physiological and pathological conditions. The conclusions of these studies allow to single out novel potential therapeutic targets among the alpha-synuclein synaptic partners. These targets may be considered for the development of new pharmacological and gene-based strategies to cure PD. (C) 2011 Elsevier B.V. All rights reserved