Search

Background Activating transcription factor 5 (ATF5) is a transcription factor that is highly expressed in undifferentiated neural progenitor/stem cells as well as a variety of human cancers including gliomas. Aims In this study, we examined the expression and localization of ATF5 protein in canine gliomas, and targeting of ATF5 function in canine glioma cell lines. Materials and Methods Paraffin-embedded canine brain glioma tissue sections and western blots of tumours and glioma cells were immunoassayed with anti-ATF5 antibody. Viability of glioma cells was tested with a synthetic cell-penetrating ATF5 peptide (CP-d/n ATF5) ATF5 antagonist. Results ATF5 protein expression was in the nucleus and cytoplasm and was present in normal adult brain and tumour samples, with significantly higher expression in tumours as shown by western immunoblotting. CP-d/n ATF5 was found to decrease cell viability in canine glioma cell lines in vitro in a dose-dependent manner. Conclusion Similarities in expression of ATF5 in rodent, dog and human tumours, and cross species efficacy of the CP-d/n ATF5 peptide support the development of this ATF5-targeting approach as a novel and translational therapy in dog gliomas.

Most people believe that their choices are based only on a rational analysis of available alternatives. Actually, emotions affect our choices and are activated as feedback during the decision process. In Recommender Systems research, preference learning is an important element to produce good recommendations. Preferences can be acquired by explicitly asking the user or by analyzing his implicit feedback that can be inferred from his behavior. Following the latter vein of research, we developed a novel approach to automatically extract these preferences by analyzing the gaze behavior and the facial expressions of the users while looking at item’s images form making a choice between them. Specifically, we focused on pairwise image comparisons in a preference elicitation experiment and exploited a Process Mining approach to learn preferences. In particular, we investigated whether behavior patterns can be learned and used to predict the user choice. We evaluated strategies based on gaze behavior and experienced emotions and results obtained so far show promising prediction performance.

Survivin (BIRC5, product of the BIRC5 gene) is highly expressed in many tumor types and has been widely identified as a potential target for cancer therapy. However, effective anti-survivin drugs remain to be developed. Here we report that both vector-delivered and cell-penetrating dominant-negative (dn) forms of the transcription factor ATF5 that promote selective death of cancer cells in vitro and in vivo cause survivin depletion in tumor cell lines of varying origins. dn-ATF5 decreases levels of both survivin mRNA and protein. The depletion of survivin protein appears to be driven at least in part by enhanced proteasomal turnover and depletion of the deubiquitinase USP9X. Survivin loss is rapid and precedes the onset of cell death triggered by dn-ATF5. Although survivin downregulation is sufficient to drive tumor cell death, survivin over-expression does not rescue cancer cells from dn-ATF5-promoted apoptosis. This indicates that dn-ATF5 kills malignant cells by multiple mechanisms that include, but are not limited to, survivin depletion. Cell-penetrating forms of dn-ATF5 are currently being developed for potential therapeutic use and the present findings suggest that they may pose an advantage over treatments that target only survivin.

The basic leucine zipper transcription factor ATF5 is overexpressed in many tumor types and interference with its expression or function inhibits cancer cell survival. As a potential therapeutic approach to exploit these findings, we created dominant-negative (DN) ATF5 forms lacking DNA-binding ability that retain the ATF5 leucine zipper, and thus associate with and sequester ATF5's requisite leucine zipper–binding partners. Preclinical studies with DN-ATF5, including a cell-penetrating form, show in vitro and in vivo efficacy in compromising cancer cell survival. However, DN-ATF5's targets, and particularly those required for tumor cell survival, have been unknown. We report that cells lacking ATF5 succumb to DN-ATF5, indicating that ATF5 itself is not DN-ATF5's obligate target. Unbiased pull-down assays coupled with mass spectrometry and immunoblotting revealed that DN-ATF5 associates in cells with the basic leucine zipper proteins CEBPB and CEBPD and coiled-coil protein CCDC6. Consistent with DN-ATF5 affecting tumor cell survival by suppressing CEBPB and CEBPD function, DN-ATF5 interferes with CEBPB and CEBPD transcriptional activity, while CEBPB or CEBPD knockdown promotes apoptotic death of multiple cancer cells lines, but not of normal astrocytes. We propose a two-pronged mechanism by which DN-ATF5 kills tumor cells. One is by inhibiting heterodimer formation between ATF5 and CEBPB and CDBPD, thus suppressing ATF5-dependent transcription. The other is by blocking the formation of transcriptionally active CEBPB and CEBPD homodimers as well as heterodimers with partners in addition to ATF5. Implications: This study indicates that the potential cancer therapeutic DN-ATF5 acts by associating with and blocking the transcriptional activities of CEBPB and CEBPD.

High manganese TWIP steels have a remarkable potential to produce impact resistant car body parts in the automotive industry. Hybrid laser arc welding (HLAW), i.e. simultaneous action of an arc and laser source, can be regarded as the most promising joining technique for such components due to, among other factors, the possibility to use filler metals to avoid improper microstructures and Mn chemical segregations in dissimilar welds of TWIP with other steel grades. In this regard, this work studies the influence of the main welding settings during HLAW of 1.4 mm thick TWIP and Dual Phase (DP) steel sheets used in the automotive industry, which were joined with the use of an austenitic steel filler. Quality of TWIP-DP welds was investigated based on welding parameters, tensile and hardness properties, and microstructures. TWIP and DP steel sheets can be properly joined with a linear heat input in the 50–75 kJ/m range. Welded samples exhibit an average tensile strength and elongation at fracture roughly of 650 MPa and 13%, respectively. These values are similar to those of DP steel, but significantly lower than TWIP steel. The tensile welded samples failed within the heat affected zone of DP steel, being weldments and TWIP steel more resistant. This work confirms that HLAW can be considered a promising solution to join vehicle components made of TWIP and DP sheets when fillers are needed.

In this paper, TWinning-Induced Plasticity (TWIP), Dual Phase (DP) and austenitic stainless (AISI 316) steels were joined together by hybrid laser/arc welding with an austenitic steel filler. Microstructural and mechanical characterization of welded joints was carried out by optical microscopy, microhardness, tensile and bend tests. The heat affected zone (HAZ) at the TWIP side was fully austenitic and exhibited a grain coarsening; at the DP side, new martensite formed close to the fusion zone and other phase transformations occurred moving toward the base metal; at the AISI 316 side, stringers of untreated ferrite δ inside the austenitic matrix was observed. All the tensile welded specimens broke within austenitic stainless steel. TWIP/AISI 316 welds exhibited a greater tensile strength than DP/AISI 316 ones. The bending tests confirmed the ductility of both sorts of joints.

The aim of this work was to evaluate the effects of aging treatments, in terms of microstructure and micro-hardness, on 18 Ni 300 Maraging steel samples processed by Selective Laser Melting (SLM) and to compare them with those of the same material made by casting. Optimized process parameters were used to build SLM parts in order to obtain almost full density (above 99%). For aging treatments, following the principles of the Design of Experiments, a full factorial plan with 4 levels of temperature and 6 levels of time was used. Before aging, the microstructure consisted of many overlapping fused/resolidified zones with a dendritic solidification morphology and epitaxial growth of the grains. After aging, the fused/resolidified zones disappeared and the hardening of the material was mainly governed by the mechanism of precipitation. The maximum value of hardness, on selective laser melted samples, was obtained, for a treatment temperature of 480 °C for 10 h: whereas, at the same temperature and after 10 h, on casting sample, it stabilized at 640 HV. Compared to samples produced by casting, SLM samples showed a lower increment in micro hardness at the aging temperature of 440 °C, while for 510 °C and 560 °C, trends were similar.

Glioblastoma, a high-grade diffuse glioma, carries a poor clinical prognosis despite decades of extensive research on the genetic and molecular features of disease and investigation of experimental therapeutics. Because spontaneous canine glioma and human glioblastoma share many clinicopathologic characteristics, recent efforts have focused on utilizing companion dogs as a preclinical model for both research and therapeutic development. A detailed investigation of the canine disease, with particular attention to the genetic and molecular profile, is important in order to allow translation of specific clinical findings from canines to humans and vice versa. In this chapter, we investigate the most common genetic, molecular, and epigenetic alterations associated with canine and human glioma. Appropriate implementation of the canine glioma model may provide valuable information to improve both human and veterinary patient care.

Glioblastoma Multiforme (GBM) tumors contain a small population of glioma stem-like cells (GSCs) among the various differentiated GBM cells (d-GCs). GSCs drive tumor recurrence, and resistance to Temozolomide (TMZ), the standard of care (SoC) for GBM chemotherapy. In order to investigate a potential link between GSC specific mitochondria function and SoC resistance, two patient-derived GSC lines were evaluated for differences in their mitochondrial metabolism. In both the lines, GSCs had significantly lower mitochondrial -content, and -function compared to d-GCs. In vitro, the standard mitochondrial-specific inhibitors oligomycin A, antimycin A, and rotenone selectively inhibited GSC proliferation to a greater extent than d-GCs and human primary astrocytes. These findings indicate that mitochondrial inhibition can be a potential GSC-targeted therapeutic strategy in GBM with minimal off-target toxicity. Mechanistically the standard mitochondrial inhibitors elicit their GSC-selective cytotoxic effects through the induction of apoptosis or autophagy pathways. We tested for GSC proliferation in the presence of 3 safe FDA-approved drugs--trifluoperazine, mitoxantrone, and pyrvinium pamoate, all of which are also known mitochondrial-targeting agents. The SoC GBM therapeutic TMZ did not trigger cytotoxicity in glioma stem cells, even at 100 μM concentration. By contrast, trifluoperazine, mitoxantrone, and pyrvinium pamoate exerted antiproliferative effects in GSCs about 30-50 fold more effectively than temozolomide. Thus, we hereby demonstrate that FDA-approved mitochondrial inhibitors induce GSC-selective cytotoxicity, and targeting mitochondrial function could present a potential therapeutic option for GBM treatment.

Direct laser powder deposition (DLPD) is a laser process that can produce fully dense components from powders for production and repairing purposes. Marages are high-strength steels that are hardened through ageing treatment. This work presents a study which integrated the numerical and experimental approaches, for the evaluation of the properties of 18 Ni (300) laser deposited marage powder. The powder was deposited onto AISI 304 substrate by a CO2 laser. During the experiments, laser power, scanning speed, powder flow rate and degree of overlapping varied according to a Taguchi plan. In particular, the degree of overlapping between adjacent layers and between successive tracks was calculated using an analytical model. Density, porosity, dilution, microstructure, adhesion to the substrate and microhardness of multilayer samples were evaluated using statistical algorithms. Both clad-substrate and clad-clad interfaces were characterised by metallographic and SEM analysis. High-density samples, with a satisfactory level of porosity, for successful additive and part repair applications, were fabricated through the method proposed and characterised in this paper.

Direct laser metal deposition (DLMD) is an innovative additive technology becoming of key importance in the field of repairing applications for industrial and aeronautical components. The performance of the repaired components is highly related to the intrinsic presence of defects, such as cracks, porosity, excess of dilution or debonding between clad and substrate. Usually, the quality of depositions is evaluated through destructive tests and microstructural analysis. Clearly, such methodologies are inapplicable in-process or on repaired components. The proposed work aims to evaluate the capability of ultrasonic techniques to perform the mechanical characterization of additive manufactured (AM) components. The tested specimens were manufactured by DLMD using a nickel-based superalloy deposited on an AISI 304 substrate. Ultrasonic goniometric immersion tests were performed in order to mechanical characterize the substrate and the new material obtained by AM process, consisting of the substrate and the deposition. Furthermore, the relationship was evaluated between the acoustic and the mechanical properties of the AM components and the deposition process parameters and the geometrical characteristics of multiclad depositions, respectively. Finally, the effectiveness of the proposed non-destructive experimental approach for the characterization of the created deposition anomalies has been investigated.

There has been growing interest in performing organocatalysis within a supramolecular system as a means of controlling reaction reactivity and stereoselectivity. Here, a protein is used as a host for iminium catalysis. A pyrrolidine moiety is covalently linked to biotin and introduced to the protein host streptavidin for organocatalytic activity. Whereas in traditional systems stereoselectivity is largely controlled by the substituents added to the organocatalyst, enantiomeric enrichment by the reported supramolecular system is completely controlled by the host. Also, the yield of the model reaction increases over 10-fold when streptavidin is included. A 1.1 angstrom crystal structure of the protein-catalyst complex and molecular simulations of a key intermediate reveal the chiral scaffold surrounding the organocatalytic reaction site. This work illustrates that proteins can be an excellent supramolecular host for driving stereoselective secondary amine organo-catalysis.

The Italian National Police started a research on vehicular traffic to improve road safety and reduce the number of theft victims. In order to support the discovery of anomalous behavior, this paper proposes a method for data analysis to automatically detect relevant hypotheses, a data mining technique to extract relevant information and a visualization technique. Traffic flow analysis is a challenging and complex task, due to the huge size of the data involved, thus falling in the realm of Big Data. Visual Analytics tools reduce and improve the search by representing a large amount of data in a small space through smart visualizations.

BackgroundActivating transcription factor 5 (ATF5) is a transcription factor that is highly expressed in undifferentiated neural progenitor/stem cells as well as a variety of human cancers including gliomas.AimsIn this study, we examined the expression and localization of ATF5 protein in canine gliomas, and targeting of ATF5 function in canine glioma cell lines.Materials and methodsParaffin-embedded canine brain glioma tissue sections and western blots of tumours and glioma cells were immunoassayed with anti-ATF5 antibody. Viability of glioma cells was tested with a synthetic cell-penetrating ATF5 peptide (CP-d/n ATF5) ATF5 antagonist.ResultsATF5 protein expression was in the nucleus and cytoplasm and was present in normal adult brain and tumour samples, with significantly higher expression in tumours as shown by western immunoblotting. CP-d/n ATF5 was found to decrease cell viability in canine glioma cell lines in vitro in a dose-dependent manner.ConclusionSimilarities in expression of ATF5 in rodent, dog and human tumours, and cross species efficacy of the CP-d/n ATF5 peptide support the development of this ATF5-targeting approach as a novel and translational therapy in dog gliomas.

Direct Laser Metal Deposition (DLMD) has been successfully applied for the coating or the repair of several kind of components, such as molds and dies. Recently, the aeronautical sector is also showing a high interest in this process for the repair of turbines and transmissions. However, technical requirements to be met for the repair of aeronautical components are much more stringent than standards of other industrial fields. Some of the deposited material defects that need to be carefully controlled are cracks and porosity, which largely depend on the temperature peaks and the cooling rates generated during the process. The aim of this work is to monitor the temperature field that was generated during the DLMD process, analyze its variation with some process parameters and study its effects on clad geometry and on dilution with the substrate. In this research, a number of experimental tests were designed for the deposition of single clads of a Nickel superalloy powder on an AISI 304 stainless steel substrate, using an Ytterbium fiber laser source. Temperature fields monitoring was carried out using a thermal camera capable of detecting temperatures up to 2500 °C.

In this study, AISI 304 and AISI 410 stainless steels were welded together by a fiber laser-TIG hybrid welding system. Weldability was evaluated through the characterization of the microstructure and mechanical properties. The effects of welding parameters, such as laser line energy (ratio between laser power and feed rate) and arc current, were analyzed. Welds microstructure and defects were assessed and related to the tensile test, type of fracture and residual stress. The latter was measured in the normal and longitudinal directions respect to the weld line. The difference observed in the two sides of the weld were highlighted and commented. FZ microstructure resulted predominantly martensitic, which increased the micro-hardness up to an average value of about 350 HV. Porosity reached a minimum, about 0.2%, when the maximum values were used for laser line energy and arc current. Maximum elongation was as far as 4.87% and the tensile strength as big as 391 MPa. Residual stresses were much lower than the ultimate tensile and yield stresses. Eventually, the paper demonstrated the good and easy weldability of dissimilar austenitic and martensitic stainless steels in annealed condition by fiber laser coupled with an electric arc.

International audience; Hospital networks are an emerging organizational form designed to face the new challenges of public health systems. Although the benefits introduced by network models in terms of rationalization of resources are known, evidence about stakeholders’ perspectives on hospital network performance from the literature is scanty. Using the Competing Values Framework of organizational effectiveness and its subsequent adaptation by Minvielle et al., we conducted in 2009 a survey in five hospitals of an Italian network for oncological care to examine and compare the views on hospital network performance of internal stakeholders (physicians, nurses and the administrative staff). 329 questionnaires exploring stakeholders’ perspectives were completed, with a response rate of 65.8%. Using exploratory factor analysis of the 66 items of the questionnaire, we identified 4 factors, i.e. Centrality of relationships, Quality of care, Attractiveness/Reputation and Staff empowerment and Protection of workers’ rights. 42 items were retained in the analysis. Factor scores proved to be high (mean score > 8 on a 10-item scale), except for Attractiveness/Reputation (mean score 6.79), indicating that stakeholders attach a higher importance to relational and health care aspects. Comparison of factor scores among stakeholders did not reveal significant differences, suggesting a broadly shared view on hospital network performance.

In this paper an experimental analysis on laser beam welding process of Ti6Al4 V titanium alloy was performed. Butt joints with a thickness of 2 mm were produced and assessed. A high brightness fiber laser was adopted to carry out the analysis. No filler wire and shaped grooves were used. An experimental plan was implemented keeping constant the laser power and varying the welding speed. A proper device was adopted to prevent oxidation phenomena that involve Ti at high temperatures reached during the thermal process. The geometric characteristics of the welding beads were studied through metallographic inspection of the cross sections. Vickers micro-hardness tests were conducted at the mid thickness of the cross sections of the joints and its variation in function of the distance from the welding line was achieved. Two tensile strength tests for each welding condition were performed in order to evaluate the mechanical properties of the welds. Results revealed a contained extent of the FZ and HAZ in the cross sections. The reduction of mechanical properties of the welds in comparison with those typical of the parent metal was about 20%, which is a good result in comparison with other previous research works.

Activating transcription factor 5 (ATF5) is a cellular prosurvival transcription factor within the basic leucine zipper (bZip) family that is involved in cellular differentiation and promotes cellular adaptation to stress. Recent studies have characterized the oncogenic role of ATF5 in the development of several different types of cancer, notably glioblastoma. Preclinical assessment of a systemically deliverable dominant-negative ATF5 (dnATF5) biologic has found that targeting ATF5 results in tumor regression and tumor growth inhibition of glioblastoma xenografts in mouse models. In this review, we comprehensively and critically detail the current scientific literature on ATF5 in the context of cellular differentiation, survival, and response to stressors in normal tissues. Furthermore, we will discuss how the prosurvival role of ATF5 aides in cancer development, followed by current advances in targeting ATF5 using dominant-negative biologics, and perspectives on future research.

Pterin-containing natural products have diverse functions in life, but an efficient and easy scheme for their in vitro synthesis is not available. Here we report a chemoenzymatic 14-step, one-pot synthesis that can be used to generate 13C- and 15N-labeled dihydrofolates (H2F) from glucose, guanine, and p-aminobenzoyl-l-glutamic acid. This synthesis stands out from previous approaches to produce H2F in that the average yield of each step is >91% and it requires only a single purification step. The use of a one-pot reaction allowed us to overcome potential problems with individual steps during the synthesis. The availability of labeled dihydrofolates allowed the measurement of heavy-atom isotope effects for the reaction catalyzed by the drug target dihydrofolate reductase and established that protonation at N5 of H2F and hydride transfer to C6 occur in a stepwise mechanism. This chemoenzymatic pterin synthesis can be applied to the efficient production of other folates and a range of other natural compounds with applications in nutritional, medical, and cell-biological research.

The lethality of the aggressive brain tumor glioblastoma multiforme (GBM) results in part from its strong propensity to invade surrounding normal brain tissue. Although oncogenic drivers such as epidermal growth factor receptor activation and Phosphatase and Tensin homolog inactivation are thought to promote the motility and invasiveness of GBM cells via phosphatidylinostitol 3-kinase activation, other unexplored mechanisms may also contribute to malignancy. Here we demonstrate that several components of the planar cell polarity (PCP) arm of non-canonical Wnt signaling including VANGL1, VANGL2 and FZD7 are transcriptionally upregulated in glioma and correlate with poorer patient outcome. Knockdown of the core PCP pathway component VANGL1 suppresses the motility of GBM cell lines, pointing to an important mechanistic role for this pathway in glioblastoma malignancy. We further observe that restoration of Nrdp1, a RING finger type E3 ubiquitin ligase whose suppression in GBM also correlates with poor prognosis, reduces GBM cell migration and invasiveness by suppressing PCP signaling. Our observations indicate that Nrdp1 physically interacts with the Vangl1 and Vangl2 proteins to mediate the K63-linked polyubiquitination of the Dishevelled, Egl-10 and Pleckstrin (DEP) domain of the Wnt pathway protein Dishevelled (Dvl). Ubiquitination hinders Dvl binding to phosphatidic acid, an interaction necessary for efficient Dvl recruitment to the plasma membrane upon Wnt stimulation of Fzd receptor and for the propagation of downstream signals. We conclude that the PCP pathway contributes significantly to the motility and hence the invasiveness of GBM cells, and that Nrdp1 acts as a negative regulator of PCP signaling by inhibiting Dvl through a novel polyubiquitination mechanism. We propose that the upregulation of core PCP components, together with the loss of the key negative regulator Nrdp1, act coordinately to promote GBM invasiveness and malignancy.

Pterin-containing natural products have diverse functions in life, but an efficient and easy scheme for their in vitro synthesis is not available. Here we report a chemoenzymatic 14-step, one-pot synthesis that can be used to generate

The expression of activating transcription factor 5 (ATF5) correlates negatively with patient survival in different types of cancer. ATF5 is important for the survival and proliferation of cancer cells, and can be targeted to selectively trigger cancer cell apoptosis while sparing normal cells. Cell-penetrating peptides combined with a dominant negative ATF5 cargo have recently shown efficacy against brain, breast, melanoma, and prostate cancers.

Mammalian dihydrofolate reductases (DHFRs) catalyze the reduction of folate more efficiently than the equivalent bacterial enzymes do, despite typically having similar efficiencies for the reduction of their natural substrate, dihydrofolate. In contrast, we show here that DHFR from the hyperthermophilic bacterium Thermotoga maritima can catalyze reduction of folate to tetrahydrofolate with an efficiency similar to that of reduction of dihydrofolate under saturating conditions. Nuclear magnetic resonance and mass spectrometry experiments showed no evidence of the production of free dihydrofolate during either the EcDHFR- or TmDHFR-catalyzed reductions of folate, suggesting that both enzymes perform the two reduction steps without release of the partially reduced substrate. Our results imply that the reaction proceeds more efficiently in TmDHFR than in EcDHFR because the more open active site of TmDHFR facilitates protonation of folate. Because T. maritima lives under extreme conditions where tetrahydrofolate is particularly prone to oxidation, this ability to salvage folate may impart an advantage to the bacterium by minimizing the squandering of a valuable cofactor.