Identification and sequence comparison of a cuticular collagen of Brugia pahangi

The cuticle of filarial nematodes is a specialized extracellular matrix that covers the parasite and protects it from adverse conditions of the environment. As a surface structure it is in direct contact with the host defence mechanisms and therefore plays an important role in the molecular host-parasite relationship. Using polyclonal antisera raised against the insoluble components of the cuticle of the adult filarial parasite Brugia pahangi, we have isolated cDNA clones encoding collagen molecules of the cuticle. The protein domain structure of cDNA clone Bpcol-1 was compared with the known structures of cuticular collagens of the nematodes Brugia malayi, Caenorhabditis elegans, Ascaris suum and Haemonchus contortus, confirming interspecies similarities. Using affinity-purified anti-Bpcol-1 antibodies we identified Bpcol-1 antigenic determinants in different nematode extracts, and determined the localization of such epitopes within the cuticle of B. pahang

Summary Report on Live Demos and Videos with results

Within this deliverable, reports on the results of Live Demos showcasing the AEROWORKS robotic team potential towards autonomous infrastructure inspection and maintenance operations will be provided. Special care will be taken in outlining the feedback from the end-users during the most important public demonstrations, while video results will be available to emphasize the achieved capabilities

Progress in the design and construction of SPES at INFN-LNL

INFN-LNL is constructing an ISOL (Isotope Separation On Line) facility delivering neutron rich ion beams at 10 A MeV or beyond, making use of the linear accelerator ALPI as the secondary accelerator. The facility includes a direct ISOL target based on UCx and able to reach 1013 fissions/s. In parallel, an applied physics facility will be developed, with applications in medicine and neutron production. The SPES project is a national facility, approved and funded. Commissioning with the first exotic species is expected in 2019. The primary accelerator is a commercial cyclotron, which will send a 40 MeV, 200 μA proton beam onto an UCx target, connected to SIS, PIS and LIS ion sources. The extracted beam is purified through a Low Resolution Mass Separator (LMRS, i.e. a Wien filter and a dispersive dipole), a beam cooler and a High Resolution Mass Separator (HRMS) and sent to an ECR charge breeder to boost the exotic beam charge state. The highly charged exotic beam is further separated in a MRMS (Medium Resolution Mass Separator) and injected into a 100% duty cycle RFQ and into the existing superconducting linac ALPI, which will be refurbished and upgraded to be an efficient exotic beam accelerator. The upgrade of ALPI will give ∼10 A MeV energy to 132Sn19+, taken as the reference ion beam. The paper presents the status of the design and construction of the SPES facility

Human African trypanosomiasis in travellers to Kenya

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Prostate Field Cancerization -- Thinking Outside the Tumor

Prostate field cancerization (or field effect) is characterized by the presence of molecular alterations in histologically normal tissues adjacent to adenocarcinomas. Accordingly, our research indicates deregulated expression of several proteins that define this type of molecular pathology. The scope of the present study was to determine the expression of the key transcription factor and potential marker of field cancerization early growth response 1 (EGR-1) in human prostate tissues derived from prostatectomies and biopsy cores. EGR-1 was detected by immunofluorescence using a polyclonal anti-human EGR-1 and Alexa Fluor 488-conjugated secondary antibodies. EGR-1 expression was quantitated by determining the pixel count per area (signal intensity) in digitized images using ImageJ software. In this proof-of-concept study, a total of 4 cases consisting each of prostatectomy and matched biopsy material, either containing or devoid of cancerous cells (malignant or benign) were analyzed. Expression levels and data distribution of EGR-1 were similar by region of interest analysis between malignant and benign prostatectomies (p = 0.28), but different between malignant and benign biopsies (p \u3c 0.05). EGR-1 protein expression is similar in cancerous (malignant) and in histologically normal adjacent (benign) tissues from both prostatectomy and biopsy specimens. This supports the concept of field cancerization and indicates a potential organ-wide molecular change, regardless of the presence or absence of cancer cells. Markers of field cancerization, such as EGR-1, could be exploited as pre-surgical disease indicators in false negative biopsies, identify areas of repeat biopsy, and add molecular information to surgical margins

Exploring EGR-1 as a Master Regulator of Prostate Field Cancerization

Field cancerization denotes the presence of molecular aberrations (genetic, epigenetic, biochemical) in structurally intact cells residing in histologically normal tissues adjacent to tumors. Markers of field cancerization in prostate tissues have the potential to improve the clinical management of this malignancy through their potential to act as indicators of early disease and to serve as molecular targets for early intervention. However, for this, a detailed understanding of the functional pathways underlying field cancerization is necessary. We have recently identified four protein markers of prostate field cancerization, i.e. the key transcription factor early growth response 1 (EGR-1), the lipogenic enzyme fatty acid synthase (FASN), and the secreted growth factors platelet derived growth factor A (PDGF-A) and macrophage inhibitory cytokine 1 (MIC-1). In this study, we provide for the first time a comprehensive association analysis between these factors, especially a potentially regulatory role of EGR-1 for the other factors, using cell models of prostate cancer and expression data in human prostate tissues. Our results indicate a potential discrepancy between research in vitro and observations in situ. More importantly, our detailed tissue expression analyses reveal novel functional pathways of prostate cancerization with a central regulatory role for EGR-1

Structure Activity Relationship Studies of Novel Diarylpentanoid Analogs Targeting The Androgen Receptor in Prostate Cancer Cells

The development of prostate cancer (PCa) relies strongly on the activation of the androgen receptor (AR) signaling pathway by its natural ligand dihydrotestosterone. Furthermore, PCa progression to metastatic disease represents oncogene addiction to AR activity. Androgen ablation therapy is thus a mainstay therapy against this disease, but the development of ligand-independent AR activation and persisting AR expression eventually leads to castration resistant PCa (CRPC). Therefore, down-regulation of AR expression in PCa cells may be an effective therapeutic modality. The diarylpentanoid ca27 has previously been shown to down-regulate AR expression by an unknown mechanism of action. The present work represents a preliminary structure activity relationship (SAR) study addressing the contribution of the hydroxyl (OH) groups and Michael acceptors of ca27 to the down-regulation of AR protein expression. Accordingly, LNCaP human PCa cells were treated with ca27 and a selection of analogs differing with respect to the position of the OH groups and the presence/absence of the Michael acceptors. AR expression was determined by Western blotting using specific antibodies against the AR and β-actin as a loading control and quantified using Image J analysis. The dose-dependent effect of ca27 and its analogs was visualized by bright field light microscopy. Our data shows that the presence of OH groups and Michael acceptors are major contributors of AR down-regulation. In addition, this observation was confirmed by the dose-responsive nature of our results. Our studies aim at identifying active pharmacophores of diarylpentanoids that down-regulate AR expression. When targeted to prostate cancer cells, this could lead to the development of novel therapeutics against CRPC

Molecular Insights into Prostate Field Cancerization: Telomere Length, EGR-­‐1 Expression, and Regulation of MIC-­‐1, PDGF-­‐A, and FAS

The diagnosis of prostate cancer (adenocarcinoma) relies on screening for elevated prostate-specific antigen (PSA) in blood samples and on digital rectal examination (DRE). With high PSA levels and/or abnormal DRE, physicians recommend a biopsy, which often misses the location of the adenocarcinoma and results in false negatives. Previous studies have shown expression of the key transcription factor early growth response 1 (EGR-1), the pro-survival factor macrophage inhibitor cytokine 1 (MIC-1), and the growth stimulatory platelet derived growth factor A (PDGF-A) to be up-regulated in histologically normal tissues 1 centimeter adjacent to prostate adenocarcinomas. We hypothesize that tumors emerge from “field cancerized” tissues, and while such tissues appear normal under gross histological examination, further analysis reveals a molecular history that adds insight into development of oncogenesis. We aim to explore EGR-1, MIC-1, and PDGF-A as essential markers of field cancerization and prostate oncogenesis that may also provide an early detection of premalignant cells by biochemical and molecular biological methods. We present here data from our ongoing investigations into the effect of tissue microenvironmental factors involved in tumorigenesis, for example oxidative stress and telomere-mediated genomic instability in human prostate cancer cell models. We also present data on the regulation of expression of MIC-1 and PDGF-A by EGR-1 using human prostate cancer cell models

Identification and Characterization of Epicuticular Proteins of Nematodes Sharing Motifs with Cuticular Proteins of Arthropods

Specific collagens and insoluble proteins called cuticlins are major constituents of the nematode cuticles. The epicuticle, which forms the outermost electron-dense layer of the cuticle, is composed of another category of insoluble proteins called epicuticlins. It is distinct from the insoluble cuticlins localized in the cortical layer and the fibrous ribbon underneath lateral alae. Our objective was to identify and characterize genes and their encoded proteins forming the epicuticle. The combination between previously obtained laboratory results and recently made available data through the whole-genome shotgun contigs (WGS) and the transcriptome Shotgun Assembly (TSA) sequencing projects of Ascaris suum allowed us to identify the first epicuticlin gene, Asu-epic-1, on the chromosome VI. This gene is formed of exon1 (55 bp) and exon2 (1067 bp), separated by an intron of 1593 bp. Exon 2 is formed of tandem repeats (TR) whose number varies in different cDNA and genomic clones of Asu-epic-1. These variations could be due to slippage of the polymerases during DNA replication and RNA transcription leading to insertions and deletions (Indels). The deduced protein, Asu-EPIC-1, consists of a signal peptide of 20 amino acids followed by 353 amino acids composed of seven TR of 49 or 51 amino acids each. Three highly conserved tyrosine motifs characterize each repeat. The GYR motif is the Pfam motif PF02756 present in several cuticular proteins of arthropods. Asu-EPIC-1 is an intrinsically disordered protein (IDP) containing seven predicted molecular recognition features (MoRFs). This type of protein undergoes a disorder-to-order transition upon binding protein partners. Three epicuticular sequences have been identified in A. suum, Ascaris lumbricoides, and Toxocara canis. Homologous epicuticular proteins were identified in over 50 other nematode species. The potential of this new category of proteins in forming the nematode cuticle through covalent interactions with other cuticular components, particularly with collagens, is discussed. Their localization in the outermost layer of the nematode body and their unique structure render them crucial candidates for biochemical and molecular interaction studies and targets for new biotechnological and biomedical applications