Ancient DNA extraction and amplification of human bone samples from the area of Delphi: a pilot case study

The present work is a preliminary effort, an experiment on the extraction of ancient DNA from human remains from a proto-Byzantine context in the area of Delphi. The first results are encouraging; however, the interpretation of such analyses needs to be very careful. DNA and other scientific methods have to take into consideration all historical and socio-economic characteristics of a past society before the proposal, for example, of the existence or migration of specific ethnic groups in an area. The theoretical and methodological thinking of Archaeology in the last decades suggest that all scientific analyses have to evaluate the specific context and the complex nature of human existence before the application of any general-based model

Effect of Aging Treatment on Surface Roughness, Mechanical Properties, and Fracture Behavior of 6xxx and 7xxx Aluminum Alloys

The effect of aging treatment on the surface roughness and mechanical properties of AA6061 and AA7075 alloys was studied. Microhardness and tensile tests were used to investigae the mechanical properties. X-ray diffraction analysis was used to investigate the surface of the specimens. Furthermore, after tensile tests fractured surfaces were examined with scanning electron microscopy. An atomic force microscope was employed for analysis of the effect of aging treatment on surface roughness. Higher surface roughness with an increase in the volume fraction of the precipitate was revealed.Исследовано влияние процесса старения на шероховатость поверхности и механические свойства алюминиевых сплавов AA6061 и AA7075. Механические свойства исследовали при испытаниях на микротвердость и растяжение. Поверхность образцов исследовали с помощью рентгеноструктурного анализа. После испытания на растяжение поверхность разрушения исследовали методом растровой электронной микроскопии. Влияние процесса старения на шероховатость поверхности изучали с помощью атомно-силового микроскопа. Показано, что с ростом шероховатости поверхности увеличивается количество выделившихся фаз.Досліджено вплив процесу старіння на шорсткість поверхні і механічні властивості алюмінієвих сплавів АА6061 та АА7075. Механічні властивості досліджували при випробуваннях на мікротвердість і розтяг. Поверхню зразків досліджували за допомогою рентгеноструктурного аналізу. Після випробувань на розтяг поверхню руйнування досліджували методом растрової електронної мікроскопії. Вплив процесу старіння на шорсткість поверхні вивчали за допомогою атомно-силового мікроскопа. Показано, що з ростом шорсткості поверхні збільшується кількість виділених фа

Effect of Demineralized Bone Matrix, Bone Marrow Mesenchymal Stromal Cells, and Platelet-Rich Plasma on Bone Tunnel Healing After Anterior Cruciate Ligament Reconstruction: A Comparative Micro-Computed Tomography Study in a Tendon Allograft Sheep Model

Background: The effect of demineralized bone matrix (DBM), bone marrow–derived mesenchymal stromal cells (BMSCs), and platelet-rich plasma (PRP) on bone tunnel healing in anterior cruciate ligament reconstruction (ACLR) has not been comparatively assessed. Hypothesis: These orthobiologics would reduce tunnel widening, and the effects on tunnel diameter would be correlated with tunnel wall sclerosis. Study Design: Controlled laboratory study. Methods: A total of 20 sheep underwent unilateral ACLR using tendon allograft and outside-in interference screw fixation. The animals were randomized into 4 groups (n = 5 per group): Group 1 received 4mL of DBM paste, group 2 received 10 million BMSCs in fibrin sealant, group 3 received 12 mL of activated leukocyte-poor platelet-rich plasma, and group 4 (control) received no treatment. The sheep were euthanized after 12 weeks, and micro-computed tomography scans were performed. The femoral and tibial tunnels were divided into thirds (aperture, midportion, and exit), and the trabecular bone structure, bone mineral density (BMD), and tunnel diameter were measured. Tunnel sclerosis was defined by a higher bone volume in a 250-µm volume of interest compared with a 4-mm volume of interest surrounding the tunnel. Results: Compared with the controls, the DBM group had a significantly higher bone volume fraction (bone volume/total volume [BV/TV]) (52.7% vs 31.8%; P = .020) and BMD (0.55 vs 0.47 g/cm3; P = .008) at the femoral aperture and significantly higher BV/TV at femoral midportion (44.2% vs 32.9%; P = .038). There were no significant differences between the PRP and BMSC groups versus controls in terms of trabecular bone analysis or BMD. In the controls, widening at the femoral tunnel aperture was significantly greater than at the midportion (46.7 vs 41.7 mm2; P = .034). Sclerosis of the tunnel was common and most often seen at the femoral aperture. In the midportion of the femoral tunnel, BV/TV (r = 0.52; P = .019) and trabecular number (r S = 0.50; P = .024) were positively correlated with tunnel widening. Conclusion: Only DBM led to a significant increase in bone volume, which was seen in the femoral tunnel aperture and midportion. No treatment significantly reduced bone tunnel widening. Tunnel sclerosis in the femoral tunnel midportion was correlated significantly with tunnel widening

A PMT-Block test bench

The front-end electronics of the ATLAS hadronic calorimeter (Tile Cal) is housed in a unit, called {\it PMT-Block}. The PMT-Block is a compact instrument comprising a light mixer, a PMT together with its divider and a {\it 3-in-1} card, which provides shaping, amplification and integration for the signals. This instrument needs to be qualified before being assembled on the detector. A PMT-Block test bench has been developed for this purpose. This test bench is a system which allows fast, albeit accurate enough, measurements of the main properties of a complete PMT-Block. The system, both hardware and software, and the protocol used for the PMT-Blocks characterisation are described in detail in this report. The results obtained in the test of about 10000 PMT-Blocks needed for the instrumentation of the ATLAS (LHC-CERN) hadronic Tile Calorimeter are also reported.Comment: 23 pages, 10 figure

MicroRNA-Restricted Transgene Expression in the Retina

Background: Gene transfer using adeno-associated viral (AAV) vectors has been successfully applied in the retina for the treatment of inherited retinal dystrophies. Recently, microRNAs have been exploited to fine-tune transgene expression improving therapeutic outcomes. Here we evaluated the ability of retinal-expressed microRNAs to restrict AAV-mediated transgene expression to specific retinal cell types that represent the main targets of common inherited blinding conditions. Methodology/Principal Findings: To this end, we generated AAV2/5 vectors expressing EGFP and containing four tandem copies of miR-124 or miR-204 complementary sequences in the 39UTR of the transgene expression cassette. These vectors were administered subretinally to adult C57BL/6 mice and Large White pigs. Our results demonstrate that miR-124 and miR-204 target sequences can efficiently restrict AAV2/5-mediated transgene expression to retinal pigment epithelium and photoreceptors, respectively, in mice and pigs. Interestingly, transgene restriction was observed at low vector doses relevant to therapy. Conclusions: We conclude that microRNA-mediated regulation of transgene expression can be applied in the retina to either restrict to a specific cell type the robust expression obtained using ubiquitous promoters or to provide an additiona

Circulating microRNAs Reveal Time Course of Organ Injury in a Porcine Model of Acetaminophen-Induced Acute Liver Failure

Acute liver failure is a rare but catastrophic condition which can progress rapidly to multi-organ failure. Studies investigating the onset of individual organ injury such as the liver, kidneys and brain during the evolution of acute liver failure, are lacking. MicroRNAs are short, non-coding strands of RNA that are released into the circulation following tissue injury. In this study, we have characterised the release of both global microRNA and specific microRNA species into the plasma using a porcine model of acetaminophen-induced acute liver failure. Pigs were induced to acute liver failure with oral acetaminophen over 19h±2h and death occurred 13h±3h thereafter. Global microRNA concentrations increased 4h prior to acute liver failure in plasma (P<0.0001) but not in isolated exosomes, and were associated with increasing plasma levels of the damage-associated molecular pattern molecule, genomic DNA (P<0.0001). MiR122 increased around the time of onset of acute liver failure (P<0.0001) and was associated with increasing international normalised ratio (P<0.0001). MiR192 increased 8h after acute liver failure (P<0.0001) and was associated with increasing creatinine (P<0.0001). The increase in miR124-1 occurred concurrent with the pre-terminal increase in intracranial pressure (P<0.0001) and was associated with decreasing cerebral perfusion pressure (P<0.002)

Metabolic Fingerprinting Links Oncogenic PIK3CA with Enhanced Arachidonic Acid-Derived Eicosanoids.

Oncogenic transformation is associated with profound changes in cellular metabolism, but whether tracking these can improve disease stratification or influence therapy decision-making is largely unknown. Using the iKnife to sample the aerosol of cauterized specimens, we demonstrate a new mode of real-time diagnosis, coupling metabolic phenotype to mutant PIK3CA genotype. Oncogenic PIK3CA results in an increase in arachidonic acid and a concomitant overproduction of eicosanoids, acting to promote cell proliferation beyond a cell-autonomous manner. Mechanistically, mutant PIK3CA drives a multimodal signaling network involving mTORC2-PKCζ-mediated activation of the calcium-dependent phospholipase A2 (cPLA2). Notably, inhibiting cPLA2 synergizes with fatty acid-free diet to restore immunogenicity and selectively reduce mutant PIK3CA-induced tumorigenicity. Besides highlighting the potential for metabolic phenotyping in stratified medicine, this study reveals an important role for activated PI3K signaling in regulating arachidonic acid metabolism, uncovering a targetable metabolic vulnerability that largely depends on dietary fat restriction. VIDEO ABSTRACT

Epigenetic potentiation of somatostatin-2 by guadecitabine in neuroendocrine neoplasias as a novel method to allow delivery of peptide receptor radiotherapy

Background Somatostatin receptor-2 (SSTR2) is expressed on cell surface of neuroendocrine neoplasias; its presence is exploited for the delivery of peptide receptor radionuclide therapy (PRRT). Patients with no or low expression of SSTR2 are not candidates for PRRT. SSTR2 promotor undergoes epigenetic modification, known to regulate gene expression. We investigated whether the demethylation agent, guadecitabine, could enhance the expression of SSTR2 in NET models, using radioligand uptake/PET imaging as a biomarker of epigenetic modification. Methods The effects of guadecitabine on the transcriptional, translational, and functional regulation of SSTR2 both in vitro and in vivo using low (QGP-1) and high (BON-1) methylated neuroendocrine neoplasia models was characterised. Promotor region methylation profiling of clinical samples (n = 61) was undertaken. Safety of combination guadecitabine and PRRT was assessed in vivo. Results Pyrosequencing of cell lines illustrated differential methylation indices – BON: 1 94%, QGP: 1 21%. Following guadecitabine treatment, a dose-dependent increase in SSTR2 in BON-1 at a transcriptional, translational, and functional levels using the SSTR2-directed radioligand, 18F-FET-βAG-TOCA ([18F]-FETO) (150% increase [18F]-FETO uptake, p < 0.05) was observed. In vivo, guadecitabine treatment resulted in a 70% increase in [18F]-FETO uptake in BON-1 tumour models compared models with low baseline percentage methylation (p < 0.05). No additive toxicity was observed with the combination treatment of PRRT and guadecitabine in vivo. Methylation index in clinical samples was 10.5% compared to 5.2% in controls (p = 0.03) and correlated with SSTR2 expression (Wilcoxon rank sign −3.75,p < 0.01). Conclusion Guadecitabine increases SSTR2 expression both in vitro and in vivo. The combination of demethylation agents with PRRT warrants further investigation

Cell-Specific DNA Methylation Patterns of Retina-Specific Genes

Many studies have demonstrated that epigenetic mechanisms are important in the regulation of gene expression during embryogenesis, gametogenesis, and other forms of tissue-specific gene regulation. We sought to explore the possible role of epigenetics, specifically DNA methylation, in the establishment and maintenance of cell type-restricted gene expression in the retina. To assess the relationship between DNA methylation status and expression level of retinal genes, bisulfite sequence analysis of the 1000 bp region around the transcription start sites (TSS) of representative rod and cone photoreceptor-specific genes and gene expression analysis were performed in the WERI and Y79 human retinoblastoma cell lines. Next, the homologous genes in mouse were bisulfite sequenced in the retina and in non-expressing tissues. Finally, bisulfite sequencing was performed on isolated photoreceptor and non-photoreceptor retinal cells isolated by laser capture microdissection. Differential methylation of rhodopsin (RHO), retinal binding protein 3 (RBP3, IRBP) cone opsin, short-wave-sensitive (OPN1SW), cone opsin, middle-wave-sensitive (OPN1MW), and cone opsin, long-wave-sensitive (OPN1LW) was found in the retinoblastoma cell lines that inversely correlated with gene expression levels. Similarly, we found tissue-specific hypomethylation of the promoter region of Rho and Rbp3 in mouse retina as compared to non-expressing tissues, and also observed hypomethylation of retinal-expressed microRNAs. The Rho and Rbp3 promoter regions were unmethylated in expressing photoreceptor cells and methylated in non-expressing, non-photoreceptor cells from the inner nuclear layer. A third regional hypomethylation pattern of photoreceptor-specific genes was seen in a subpopulation of non-expressing photoreceptors (Rho in cones from the Nrl −/− mouse and Opn1sw in rods). These results demonstrate that a number of photoreceptor-specific genes have cell-specific differential DNA methylation that correlates inversely with their expression level. Furthermore, these cell-specific patterns suggest that DNA methylation may play an important role in modulating photoreceptor gene expression in the developing mammalian retina