A Beckwith–Wiedemann-Associated CDKN1C Mutation Allows the Identification of a Novel Nuclear Localization Signal in Human p57Kip2

p57^Kip2 protein is a member of the CIP/Kip family, mainly localized in the nucleus where it exerts its Cyclin/CDKs inhibitory function. In addition, the protein plays key roles in embryogenesis, differentiation, and carcinogenesis depending on its cellular localization and interactors. Mutations of CDKN1C, the gene encoding human p57^Kip2, result in the development of different genetic diseases, including Beckwith–Wiedemann, IMAGe and Silver–Russell syndromes. We investigated a specific Beckwith–Wiedemann associated CDKN1C change (c.946 C>T) that results in the substitution of the C-terminal amino acid (arginine 316) with a tryptophan (R316W-p57^Kip2). We found a clear redistribution of R316W-p57^Kip2, in that while the wild-type p57^Kip2 mostly occurs in the nucleus, the mutant form is also distributed in the cytoplasm. Transfection of two expression constructs encoding the p57^Kip2 N- and C-terminal domain, respectively, allows the mapping of the nuclear localization signal(s) (NLSs) between residues 220–316. Moreover, by removing the basic RKRLR sequence at the protein C-terminus (from 312 to 316 residue), p57^Kip2 was confined in the cytosol, implying that this sequence is absolutely required for nuclear entry. In conclusion, we identified an unreported p57^Kip2 NLS and suggest that its absence or mutation might be of relevance in CDKN1C-associated human diseases determining significant changes of p57^Kip2 localization/regulatory roles.     

High SPARC Expression Starting from Dysplasia,Associated with Breast Carcinoma,Is Predictive for Bone Metastasis without Enhancement of Plasma Levels

In order to become established in the skeleton, metastatic cells disseminating from the breast carcinoma need to acquire organ-specific traits. There are no effective predictors for who will develop bone metastasis to guide long-term predictive therapy. Our purpose was to individuate events critical for bone colonization to make a molecular classification of breast carcinoma useful for bone-metastasis outcome. In dysplasia adjacent to carcinoma and in pair-matched specimens of bone metastasis we examined SPARC expression and localization as well as Endothelin 1/ET_AR signals by immunohistochemistry, and the evaluation of plasma levels of SPARC by ELISA was also performed. In patients with breast carcinoma metastasizing to bone, SPARC and Endothelin 1/ET_AR axis were highly expressed from dysplasia until bone metastasis, but the SPARC plasma level was as low as that of normal women, in contrast to patients that never develop bone metastasis, suggesting that circulating SPARC was counter adhesive. Altogether, the early identification of SPARC/Endothelin 1/ET_AR in dysplastic lesions would be important to devise therapies preventing metastasis engraftment, since often carcinoma cells spread to distant organs at the time or even before patients present with cancer.     

Ejection chain moves for automatic neighborhood synthesis in constrained cardinality‐minimization problems

In this paper, we deal with the problem of automatically synthesizing “good” neighborhoods for a specific class of problems, namely constrained cardinality‐minimization problems. Exploiting the peculiarity of the objective function of such problems, we develop automatic ejection chain moves that define neighborhood structures to be explored with a black‐box solver. In particular, starting from a formulation of a cardinality‐minimization problem and a feasible solution, our procedure automatically detects the “entities” involved in the problem and learns the strength of the relationships among them. This information is then used to define the characteristics of our moves that consist in ejecting one entity at a time from the solution. If one of such moves results in an infeasible solution, then feasibility is recovered by performing an additional step based on the solution of an auxiliary problem. The computational results show that, when assessed on four well‐known constrained cardinality‐minimization problems, our approach outperforms both a black‐box mixed integer programming solver and a state‐of‐the‐art model‐based neighborhood search procedure with respect to both solution quality and computing times.     

Reliability of RF MEMS switches due to charging effects and their circuital modelling

The reliability of RF MEMS switches is typically reduced by charging effects occurring in the dielectrics. The aim of this paper is to discuss these effects, and to propose analytical and equivalent circuit models which account for most of the physical contributions present in the structure.     

A comparison of antioxidant properties between artisan‐made and factory‐produced chocolate

The antioxidant capacities and the total phenolic content in cocoa liquor directly manufactured chocolate from an artisan manufacturer were measured using different in vitro methods (BR, TEAC, and Folin–Ciocalteu Reagent). These parameters were then compared with those of a chocolate made by a leading manufacturing company producing chocolate and cocoa‐containing products. A statistical analysis of the collected data showed that the antioxidant properties of the artisan‐made chocolate are significantly better than those of the factory‐produced one. These results were ascribed to the fact that all the bioactive components in the cocoa beans are better preserved in the artisan‐made chocolate.     

Study of ITER equatorial port plug handling system and vacuum sealing interface

In the field of the ITER port plug engineering and integration task, CEA has contributed to define proposals concerning the port plugs vacuum sealing interface with the vessel flange and the equatorial plug handling.The 2001 baseline vacuum flange sealing consisted of TIG welding of a 316L strip plate on to U shapes. This arrangement presented some issues like welding access, implementation of tools, lip consumption, complex local leak test, continuous leak checking. Therefore, an alternate sealing solution based on the use of metallic gaskets is proposed. The different technical aspects are discussed to explain how this design can simplify the maintenance and deal with safety and vacuum requirements.The design of the mechanical attachment and vacuum sealing of the plug has constantly evolved, but the associated remote handling equipment was not systematically reviewed. An update of the cask and maintenance procedure was studied in order to design it in accordance with the last generic plug flange design. This includes a concept of a gripping system that uses the plug flange bolting area and, to help the remote handling process, a cantilever assisting system is suggested to increase the reliability of the transfer operation between vacuum vessel and cask.     

Unraveling the Link between Interferon-α and Systemic Lupus Erythematosus: From the Molecular Mechanisms to Target Therapies

Systemic lupus erythematosus (SLE) is a chronic, systemic autoimmune disease with a wide range of clinical expressions. The kidney is often affected, usually within 5 years of the onset of SLE, and lupus nephropathy (LN) carries a high risk for increased morbidity. The clinical heterogeneity of the disease is accompanied by complex disturbances affecting the immune system with inflammation and tissue damage due to loss of tolerance to nuclear antigens and the deposition of immune complexes in tissues. Several studies have reported that in human SLE, there is an important role of the Type-I-interferons (INF) system suggested by the upregulation of INF-inducible genes observed in serial gene expression microarray studies. This review aims to describe the transduction pathways of Type-I-interferons, in particular INFα, and its immune-regulatory function in the pathogenesis of SLE and, in particular, in LN. In addition, recent novelties concerning biologic therapy in LN will be discussed.     

A Food-Grade Method for Enhancing the Levels of Low Molecular Weight Proanthocyanidins with Potentially High Intestinal Bioavailability

Proanthocyanidins (PACs) are a group of bioactive molecules found in a variety of plants and foods. Their bioavailability depends on their molecular size, with monomers and dimers being more bioavailable than those that have a higher polymerization degree. This study aimed to develop a method to convert high-molecular-weight PACs to low-molecular-weight ones in a grape seed extract (GSE) from Vitis vinifera L. Therefore, GSE was subjected to alkaline treatment (ATGSE), and its difference in chemical composition, compared to GSE, was evaluated using a molecular networking (MN) approach based on results obtained from HPLC-ESI HRMS/MS characterization analysis. The network analysis mainly noted the PAC cluster with about 142 PAC compounds identified. In particular, the obtained results showed a higher content of monomeric and dimeric PACs in ATGSE compared to GSE, with 58% and 49% monomers and 31% and 24% dimers, respectively. Conversely, trimeric (9%), polymeric (4%), and galloylated PACs (14%) were more abundant in GSE than in ATGSE (6%, 1%, and 4%, respectively). Moreover, in vitro antioxidant and anti-inflammatory activities were investigated, showing the high beneficial potential of both extracts. In conclusion, ATGSE could represent an innovative natural matrix rich in bioavailable and bioaccessible PACs for nutraceutical applications with potential beneficial properties.     

An Ultra-Low Dose of ∆9-Tetrahydrocannabinol Alleviates Alzheimer’s Disease-Related Cognitive Impairments and Modulates TrkB Receptor Expression in a 5XFAD Mouse Model

Alzheimer’s disease (AD) is the most common form of dementia, but there is still no available treatment. Δ9-tetrahydrocannabinol (THC) is emerging as a promising therapeutic agent. Using THC in conventional high doses may have deleterious effects. Therefore, we propose to use an ultra-low dose of THC (ULD-THC). We previously published that a single injection of ULD-THC ameliorated cognitive functioning in several models of brain injuries as well as in naturally aging mice. Here, 5xFAD AD model mice received a single treatment of ULD-THC (0.002 mg/kg) after disease onset and were examined in two separate experiments for cognitive functions, neurotropic, and inflammatory factors in the hippocampus. We show that a single injection of ULD-THC alleviated cognitive impairments in 6- and 12-month-old 5xFAD mice. On the biochemical level, our results indicate an imbalance between the truncated TrkB receptor isoform and the full receptor, with AD mice showing a greater tendency to express the truncated receptor, and ULD-THC improved this imbalance. We also investigated the expression of three AD-related inflammatory markers and found an ameliorating effect of ULD-THC. The current research demonstrates for the first time the beneficial effects of a single ultra-low dose of THC in a mouse model of AD after disease onset.     

Effects of Physiological and Pathological Urea Concentrations on Human Microvascular Endothelial Cells

Urea is the uremic toxin accumulating with the highest concentration in the plasma of chronic kidney disease (CKD) patients, not being completely cleared by dialysis. Urea accumulation is reported to exert direct and indirect side effects on the gastrointestinal tract, kidneys, adipocytes, and cardiovascular system (CVS), although its pathogenicity is still questioned since studies evaluating its side effects lack homogeneity. Here, we investigated the effects of physiological and pathological urea concentrations on a human endothelial cell line from the microcirculation (Human Microvascular Endothelial Cells-1, HMEC-1). Urea (5 g/L) caused a reduction in the proliferation rate after 72 h of exposure and appeared to be a potential endothelial-to-mesenchymal transition (EndMT) stimulus. Moreover, urea induced actin filament rearrangement, a significant increase in matrix metalloproteinases 2 (MMP-2) expression in the medium, and a significant up- or down-regulation of other EndMT biomarkers (keratin, fibrillin-2, and collagen IV), as highlighted by differential proteomic analysis. Among proteins whose expression was found to be significantly dysregulated following exposure of HMEC-1 to urea, dimethylarginine dimethylaminohydrolase (DDAH) and vasorin turned out to be down-regulated. Both proteins have been directly linked to cardiovascular diseases (CVD) by in vitro and in vivo studies. Future experiments will be needed to deepen their role and investigate the signaling pathways in which they are involved to clarify the possible link between CKD and CVD.