Optimization of the wavesax device: numerical modelling and ocean wave basin tests

The Wavesax device has been conceived to be installed in ports and harbours, in the Mediterranean sea. Therefore, two aspects are quite important: flexibility of the device to fit in different structural configurations and replication in a large number of units. Preliminary numerical modelling of the fixed component of the device has been performed using Computational Fluid Dynamics analysis (RANS-CFD model) and considering four regular wave conditions typical of the Mediterranean sea. Main issues to be considered in the first stage scale modelling analysis are the effective functionality of the device conception (scale 1:20), the optimization of the design and position of the device in terms of generated velocity gradients in the working section where the turbine blades are installed. The main parameters to be investigated, both with numerical modelling and ocean wave basin tests, are the pressure field in different sections of the device, water levels for different wave conditions and device sinking. Following the scale model test, the numerical model was calibrated and validated. The paper presents the results of the numerical simulations related to different configurations of the device, under typical Mediterranean wave climates

Heterogeneity of neuroinflammatory responses in amyotrophic lateral sclerosis: A challenge or an opportunity?

Amyotrophic Lateral Sclerosis (ALS) is a complex pathology: (i) the neurodegeneration is chronic and progressive; it starts focally in specific central nervous system (CNS) areas and spreads to different districts; (ii) multiple cell types further than motor neurons (i.e., glial/immune system cells) are actively involved in the disease; (iii) both neurosupportive and neurotoxic neuroinflammatory responses were identified. Microglia cells (a key player of neuroinflammation in the CNS) attracted great interest as potential target cell population that could be modulated to counteract disease progression, at least in preclinical ALS models. However, the heterogeneous/multifaceted microglia cell responses occurring in different CNS districts during the disease represent a hurdle for clinical translation of single-drug therapies. To address this issue, over the past ten years, several studies attempted to dissect the complexity of microglia responses in ALS. In this review, we shall summarize these results highlighting how the heterogeneous signature displayed by ALS microglia reflects not only the extent of neuronal demise in different regions of the CNS, but also variable engagement in the attempts to cope with the neuronal damage. We shall discuss novel avenues opened by the advent of single-cell and spatial transcriptomics technologies, underlining the potential for discovery of novel therapeutic targets, as well as more specific diagnostic/prognostic not-invasive markers of neuroinflammation

Low intensity laser therapy accelerates muscle regeneration in aged rats

Background: Elderly people suffer from skeletal muscle disorders that undermine their daily activity and quality of life; some of these problems can be listed as but not limited to: sarcopenia, changes in central and peripheral nervous system, blood hypoperfusion, regenerative changes contributing to atrophy, and muscle weakness. Determination, proliferation and differentiation of satellite cells in the regenerative process are regulated by specific transcription factors, known as myogenic regulatory factors (MRFs). In the elderly, the activation of MRFs is inefficient which hampers the regenerative process. Recent studies found that low intensity laser therapy (LILT) has a stimulatory effect in the muscle regeneration process. However, the effects of this therapy when associated with aging are still unknown. Objective: This study aimed to evaluate the effects of LILT (λ=830 nm) on the tibialis anterior (TA) muscle of aged rats. Subjects and methods: The total of 56 male Wistar rats formed two population sets: old and young, with 28 animals in each set. Each of these sets were randomly divided into four groups of young rats (3 months of age) with n=7 per group and four groups of aged rats (10 months of age) with n=7 per group. These groups were submitted to cryoinjury + laser irradiation, cryoinjury only, laser irradiation only and the control group (no cryoinjury/no laser irradiation). The laser treatment was performed for 5 consecutive days. The first laser application was done 24 h after the injury (on day 2) and on the seventh day, the TA muscle was dissected and removed under anesthesia. After this the animals were euthanized. Histological analyses with toluidine blue as well as hematoxylin-eosin staining (for counting the blood capillaries) were performed for the lesion areas. In addition, MyoD and VEGF mRNA was assessed by quantitative polymerase chain reaction. Results: The results showed significant elevation (p<0.05) in MyoD and VEGF genes expression levels. Moreover, capillary blood count was more prominent in elderly rats in laser irradiated groups when compared to young animals. Conclusion: In conclusion, LILT increased the maturation of satellite cells into myoblasts and myotubes, enhancing the regenerative process of aged rats irradiated with laser.National Institutes of Health (U.S.) (grant RO1AI050875

Alteration in Superoxide Dismutase 1 Causes Oxidative Stress and p38 MAPK Activation Following RVFV Infection

Rift Valley fever (RVF) is a zoonotic disease caused by Rift Valley fever virus (RVFV). RVFV is a category A pathogen that belongs to the genus Phlebovirus, family Bunyaviridae. Understanding early host events to an infectious exposure to RVFV will be of significant use in the development of effective therapeutics that not only control pathogen multiplication, but also contribute to cell survival. In this study, we have carried out infections of human cells with a vaccine strain (MP12) and virulent strain (ZH501) of RVFV and determined host responses to viral infection. We demonstrate that the cellular antioxidant enzyme superoxide dismutase 1 (SOD1) displays altered abundances at early time points following exposure to the virus. We show that the enzyme is down regulated in cases of both a virulent (ZH501) and a vaccine strain (MP12) exposure. Our data demonstrates that the down regulation of SOD1 is likely to be due to post transcriptional processes and may be related to up regulation of TNFα following infection. We also provide evidence for extensive oxidative stress in the MP12 infected cells. Concomitantly, there is an increase in the activation of the p38 MAPK stress response, which our earlier published study demonstrated to be an essential cell survival strategy. Our data suggests that the viral anti-apoptotic protein NSm may play a role in the regulation of the cellular p38 MAPK response. Alterations in the host protein SOD1 following RVFV infection appears to be an early event that occurs in multiple cell types. Activation of the cellular stress response p38 MAPK pathway can be observed in all cell types tested. Our data implies that maintaining oxidative homeostasis in the infected cells may play an important role in improving survival of infected cells

Unraveling transcriptional and translational control in plant energy homeostasis : a bioinformatic approach

Due to their sessile nature, plants require a tight regulation of energy homeostasis in order to survive and reproduce in changing environmental conditions. Regulation of gene expression is controlled at several levels, from transcription to translation and beyond. Sugars themselves can act directly as signaling molecules, accelerating molecular adaptation processes. Understanding these processes is relevant both for fundamental research in plant biology and for agricultural applications, such as targeted crop improvement programs. In this thesis, we investigated specific transcriptional and translational regulatory systems in plant energy homeostasis using different bioinformatic approaches. In Chapter 2 and 3, we discuss specific examples of transcriptional regulators whose activity is affected by sugars: ABI4 and the C/S1 bZIP transcription factor network. Both chapters investigate these regulators from an evolutionary and comparative genomics perspective. In Chapter 4, we moved our focus from transcriptional to translational control, also changing our analysis approach from phylogenetics to integrative bioinformatics. In Chapter 2, we showed that ABI4, well known for its role in seed development and germination, is conserved across land plants predominantly as a single copy gene, which suggests a strictly conserved function. We also discovered a highly conserved ‘ABI4’ motif, which is likely necessary for ABI4 activity. In Chapter 3, we investigated the evolution and conservation of members of the C and S1 families of bZIP transcription factors. Our results show that the two ortholog groups likely originated from a duplication event in algae, indicating that these genes are much more ancient than previously thought, and could have played a crucial role in the adaptation of plants to new environments. The discovery also provides a plausible scenario for the origin of heterodimerization between C and S1 bZIPs, which we discuss thoroughly. Importantly, the chapter includes experimental work showing the translational repression by sucrose (SIRT) of newly discovered gymnosperm S1 sequences, indicating that this regulatory mechanism emerged early on in the evolution of S1 bZIPs, and is likely conserved in other plant species. In Chapter 4, we focused exclusively on translational regulation, trying to identify sequence features responsible for the differential translational efficiency of specific subsets of transcripts during sucrose treatment and seed germination. Our results indicate that both general sequence properties and specific mRNA features, such as motifs and structured elements, play a role in translational control; in particular, it appears that a combination of signals rather than a single one is required to achieve condition-dependent changes in translational efficiency. Overall, our work clarified in detail the evolution of long-known energy-responsive transcriptional regulators which still lacked a phylogenetic description, and revealed precious clues on the much less explored field of translational regulation, providing directions for future studies

Synthesis and characterization of a “clickable” PBR28 TSPO-selective ligand derivative suitable for the functionalization of biodegradable polymer nanoparticles

Reactive microgliosis is a pathological hallmark that accompanies neuronal demise in many neurodegenerative diseases, ranging from acute brain/spinal cord injuries to chronic diseases, such as amyotrophic lateral sclerosis (ALS), Alzheimer’s disease (AD) and age-related dementia. One strategy to assess and monitor microgliosis is to use positron emission tomography (PET) by exploiting radioligands selective for the 18 kDa translocator protein (TSPO) which is highly upregulated in the brain in pathological conditions. Several TSPO ligands have been developed and validated, so far. Among these, PBR28 has been widely adopted for PET imaging at both preclinical and clinical levels, thanks to its high brain penetration and high selectivity. For this reason, PBR28 represents a good candidate for functionalization strategies, where this ligand could be exploited to drive selective targeting of TSPO-expressing cells. Since the PBR28 structure lacks functional moieties that could be exploited for derivatization, in this work we explored a synthetic pathway for the synthesis of a PBR28 derivative carrying an alkyne group (PBR-alkyne), enabling the fast conjugation of the ligand through azide-alkyne cycloaddition, also known as click-chemistry. As a proof of concept, we demonstrated in silico that the derivatized PBR28 ligand maintains the capability to fit into the TSPO binding pocked, and we successfully exploited PBR-alkyne to decorate zwitterionic biodegradable polymer nanoparticles (NPs) resulting in efficient internalization in cultured microglia-like cell lines

Simultaneous flow cytometric characterization of multiple cell types retrieved from mouse brain/spinal cord through different homogenization methods

Recent advances in viral vector and nanomaterial sciences have opened the way for new cutting-edge approaches to investigate or manipulate the central nervous system (CNS). However, further optimization of these technologies would benefit from methods allowing rapid and streamline determination of the extent of CNS and cell-specific targeting upon administration of viral vectors or nanoparticles in the body. Here, we present a protocol that takes advantage of the high throughput and multiplexing capabilities of flow cytometry to allow a straightforward quantification of different cell subtypes isolated from mouse brain or spinal cord, namely microglia/macrophages, lymphocytes, astrocytes, oligodendrocytes, neurons and endothelial cells. We apply this approach to highlight critical differences between two tissue homogenization methods in terms of cell yield, viability and composition. This could instruct the user to choose the best method depending on the cell type(s) of interest and the specific application. This method is not suited for analysis of anatomical distribution, since the tissue is homogenized to generate a single-cell suspension. However, it allows to work with viable cells and it can be combined with cell-sorting, opening the way for several applications that could expand the repertoire of tools in the hands of the neuroscientist, ranging from establishment of primary cultures derived from pure cell populations, to gene-expression analyses and biochemical or functional assays on well-defined cell subtypes in the context of neurodegenerative diseases, upon pharmacological treatment or gene therapy