Nano-imaging of environmental dust in human lung tissue by soft and hard X-ray fluorescence microscopy

It is well recognized that a large number of pulmonary diseases are induced by the effects of inhaled particulates. Anthracosis is defined as an asymptomatic, mild form of pneumoconiosis caused by the accumulation of \u201cblack carbon\u201d in the lungs due to repeated exposure to air pollution or inhalation of smoke or coal dust particles. Since the human population is progressively exposed to an increasing number and doses of anthropogenic micro and nano particles/compounds, there is a pressing urgency to explore toxicological impact arising from these exposures and the molecular mechanisms driving the body defense or possible related diseases. The toxicity mechanisms are clearly related to chemical composition and physical and surface properties of materials. A combination of synchrotron radiation-based (SR-based) nano X-ray fluorescence (XRF) imaging and soft X-ray microscopy was used to chemically characterize environmental particulates (anthracosis) in lung tissues from urban subjects with the aim of better understanding the complex nature of related lungs' deposits. High-resolution XRF analyses performed at ESRF and Elettra synchrotrons allowed discriminating single particles in the heterogeneous aggregates found in the lung tissue. The small particles have variable composition resulting from the different combinations of Ti with O, K and Si, Al and Si, or Zn and Fe with O. Interestingly, simultaneous absorption and phase contrast images showed the particulate morphology and allowed to predict the presence of very dense nanoparticles or high concentration of heavy elements

Mobile Daily Centre (Mdc) for Elder People with Cognitive Impairment: a Retrospective Observational Study

Introduction Trieste is a city characterized by a high mean age of the resident population, with 6,000 people with a cognitive impairment. Evidences show that is necessary to have a multidisciplinary approach, making alliances with the social network and families, while dealing with people with cognitive impairment. Because of this, the 3rd catchment district has developed a Mobile Daily Centre that aims to promote health, abilities and socialization giving the possibilities for these people to stay in a social context. Objectives Evaluating the impact of the Mobile Daily Centre on QoL of people with Cognitive Impairment. Aims Considering the rate of hospitalization and access to the first aid unit at the general hospital. Methods Retrospective Observational Study for the period between 01.01.2012 and 30.04.2014 on people in charge to the MDC. We have considered socio-demographic variables such as age, gender, care-givers; clinical variables such as psychopharmacotherapy and acetylcolinesterase-inhibitors drugs; rates of hospitalization, number of accesses to the first aid unit and of interventions of the MDC. Results in the period of the study 20 patients have been followed by the MDC; half of them had a psycho-pharmacological prescription. Very low rates of institutionalization have been detected. Conclusions MDC, in these small numbers, has shown to reduce the number of improper institutionalizations while guaranteeing to the elder people to maintain their abilities and socialization and to their care-givers periods of relief. Moreover, it promotes social inclusion and destigmatization. These results suggest that more territorial work and further studies should be done

Interaction of magnetic nanoparticles with U87MG cells studied by synchrotron radiation X-ray fluorescence techniques

International audienceSynchrotron radiation (SR) X-ray microscopy combined with X-ray fluorescence (XRF) microspectroscopy provides unique information that have pushed the frontiers of biological research, particularly when investigating intracellular mechanisms. This work reports an SR-XRF microspectroscopy investigation on the distribution and the potential toxicity of Fe 2 O 3 and CoFe 2 O 4 nanoparticles (NPs) in U87MG glioblastoma-astrocytoma cells. The U87MG cells exposed to NPs concentrations ranging from 5 to 250 mg/ml for 24 h were analyzed in order to monitor both morphological and chemical changes. The SR-XRF maps complemented with XRM absorption and phase contrast images have revealed different intracellular distribution patterns for the two nanoparticles types allowing different mechanism of toxicity to be deduced

A follow-up on psychiatric symptoms and post-traumatic stress disorders in Tuareg refugees in Burkina Faso

Introduction: The aim of this study was to carry out a 2-year follow-up of refugees in a camp in Burkina Faso who had been interviewed previously. We also aimed to verify whether the general conditions in which they lived (e.g., protection by international organizations and the conclusion of negotiations and new hope of returning to Mali and reunification with surviving family members) would affect their mental health state. Methods: This is a cross-sectional study repeated over time on a cohort of refugees. People living in the Subgandé camp who had participated in the first survey in 2012 were identified using informational chains and approached for follow-up. Those who agreed were interviewed using the Short Screening Scale for post-traumatic stress disorder (PTSD) and the K6 scale, French versions, to measure general psychopathology and the level of impairment. Results: The second survey shows a dramatic decrease in psychopathological symptoms (positivity at K6 scale). Improvement was also conspicuous in the frequency of people with stress symptoms (positivity at Short Screening Scale for PTSD and simultaneous positivity to K6 scale). The frequency of people screened positive at the Short Screening Scale for PTSD had also decreased, but the level of improvement was not pronounced. Conclusion: Our findings confirm that when physical conditions improve, psychological symptoms can also improve. Although in the studied sample psychological factors, such as the hope of returning to their own land and thus the possibility of maintaining ethnic cohesion, may have played a role, future research carried out with a proper methodology and sufficient resources to identify protective factors is needed

Impact of Sample Preparation Methods on Single-Cell X-ray Microscopy and Light Elemental Analysis Evaluated by Combined Low Energy X-ray Fluorescence, STXM and AFM

Background: Although X-ray fluorescence microscopy is becoming a widely used technique for single-cell analysis, sample preparation for this microscopy remains one of the main challenges in obtaining optimal conditions for the measurements in the X-ray regime. The information available to researchers on sample treatment is inadequate and unclear, sometimes leading to wasted time and jeopardizing the experiment's success. Many cell fixation methods have been described, but none of them have been systematically tested and declared the most suitable for synchrotron X-ray microscopy. Methods: The HEC-1-A endometrial cells, human spermatozoa, and human embryonic kidney (HEK-293) cells were fixed with organic solvents and cross-linking methods: 70% ethanol, 3.7%, and 2% paraformaldehyde; in addition, HEK-293 cells were subjected to methanol/ C3H6O treatment and cryofixation. Fixation methods were compared by coupling low-energy X-ray fluorescence with scanning transmission X-ray microscopy and atomic force microscopy. Results: Organic solvents lead to greater dehydration of cells, which has the most significant effect on the distribution and depletion of diffusion elements. Paraformaldehyde provides robust and reproducible data. Finally, the cryofixed cells provide the best morphology and element content results. Conclusion: Although cryofixation seems to be the most appropriate method as it allows for keeping cells closer to physiological conditions, it has some technical limitations. Paraformaldehyde, when used at the average concentration of 3.7%, is also an excellent alternative for X-ray microscopy

Charting DENR-dependent translation reinitiation uncovers predictive uORF features and links to circadian timekeeping via Clock.

The non-canonical initiation factor DENR promotes translation reinitiation on mRNAs harbouring upstream open reading frames (uORFs). Moreover, DENR depletion shortens circadian period in mouse fibroblasts, suggesting involvement of uORF usage and reinitiation in clock regulation. To identify DENR-regulated translation events transcriptome-wide and, in particular, specific core clock transcripts affected by this mechanism, we have used ribosome profiling in DENR-deficient NIH3T3 cells. We uncovered 240 transcripts with altered translation rate, and used linear regression analysis to extract 5' UTR features predictive of DENR dependence. Among core clock genes, we identified Clock as a DENR target. Using Clock 5' UTR mutants, we mapped the specific uORF through which DENR acts to regulate CLOCK protein biosynthesis. Notably, these experiments revealed an alternative downstream start codon, likely representing the bona fide CLOCK N-terminus. Our findings provide insights into uORF-mediated translational regulation that can regulate the mammalian circadian clock and gene expression at large

Identification of classical swine fever virus protein E2 as a target for cytotoxic T cells by using mRNA-transfected antigen-presenting cells

Vaccination of pigs against Classical swine fever virus (CSFV) by using live-virus vaccines induces early protection before detectable humoral immune responses. Immunological analyses indicate that this is associated with T-cell activation, underlining the importance of targeting cytotoxic T-lymphocyte (CTL) responses for vaccine improvement. Antigen-presenting cells (APCs) transfected with mRNA encoding structural protein E2 or non-structural viral proteins NS3¿NS4A were used to identify viral genes encoding CTL epitopes. Monocyte-derived dendritic cells (DCs) and fibrocytes served as the APCs. In vitro translation of the mRNA and microscopic analysis of transfected cells demonstrated that E2 and NS3¿NS4A could be identified. APCs transfected with either of the mRNA molecules restimulated CSFV-specific T cells to produce gamma interferon and specific cytotoxic activity against CSFV-infected target cells. The presence of CTL epitopes on E2 was confirmed by using d/d-haplotype MAX cells expressing E2 constitutively as target cells in d/d-haplotype CTL assays. A potent CTL activity against E2 was detected early (1¿3 weeks) after CSFV challenge. This work corroborates the existence of CTL epitopes within the non-structural protein domain NS3¿NS4A of CSFV. Furthermore, epitopes on the E2 protein can also now be classified as targets for CTLs, having important implications for vaccine design, especially subunit vaccines. As for the use of mRNA-transfected APCs, this represents a simple and efficient method to identify viral genes encoding CTL epitopes in outbred population

A Novel Liposome-Based Adjuvant CAF01 for Induction of CD8+ Cytotoxic T-Lymphocytes (CTL) to HIV-1 Minimal CTL Peptides in HLA-A*0201 Transgenic Mice

Background: Specific cellular cytotoxic immune responses (CTL) are important in combating viral diseases and a highly desirable feature in the development of targeted HIV vaccines. Adjuvants are key components in vaccines and may assist the HIV immunogens in inducing the desired CTL responses. In search for appropriate adjuvants for CD8+ T cells it is important to measure the necessary immunological features e.g. functional cell killing/lysis in addition to immunological markers that can be monitored by simple immunological laboratory methods. Methodology/Principal Findings: We tested the ability of a novel two component adjuvant, CAF01, consisting of the immune stimulating synthetic glycolipid TDB (Trehalose-Dibehenate) incorporated into cationic DDA (Dimethyldioctade-cylammonium bromide) liposomes to induce CD8+ T-cell restricted cellular immune responses towards subdominant minimal HLA-A0201-restricted CTL epitopes from HIV-1 proteins in HLA-A*0201 transgenic HHD mice. CAF01 has an acceptable safety profile and is used in preclinical development of vaccines against HIV-1, malaria and tuberculosis. Conclusions/Significance: We found that CAF01 induced cellular immune responses against HIV-1 minimal CTL epitopes in HLA-A*0201 transgenic mice to levels comparable with that of incomplete Freund’s adjuvant

Analogue peptides for the immunotherapy of human acute myeloid leukemia

Accepted manuscript. The final publication is available at: http://link.springer.com/article/10.1007%2Fs00262-015-1762-9The use of peptide vaccines, enhanced by adjuvants, has shown some efficacy in clinical trials. However, responses are often short-lived and rarely induce notable memory responses. The reason is that self-antigens have already been presented to the immune system as the tumor develops, leading to tolerance or some degree of host tumor cell destruction. To try to break tolerance against self-antigens, one of the methods employed has been to modify peptides at the anchor residues to enhance their ability to bind major histocompatibility complex molecules, extending their exposure to the T-cell receptor. These modified or analogue peptides have been investigated as stimulators of the immune system in patients with different cancers with variable but sometimes notable success. In this review we describe the background and recent developments in the use of analogue peptides for the immunotherapy of acute myeloid leukemia describing knowledge useful for the application of analogue peptide treatments for other malignancies