Analysis of Bovine Viral Diarrhea Viruses-infected monocytes: identification of cytopathic and non-cytopathic biotype differences

<p>Abstract</p> <p>Background</p> <p>Bovine Viral Diarrhea Virus (BVDV) infection is widespread in cattle worldwide, causing important economic losses. Pathogenesis of the disease caused by BVDV is complex, as each BVDV strain has two biotypes: non-cytopathic (ncp) and cytopathic (cp). BVDV can cause a persistent latent infection and immune suppression if animals are infected with an ncp biotype during early gestation, followed by a subsequent infection of the cp biotype. The molecular mechanisms that underscore the complex disease etiology leading to immune suppression in cattle caused by BVDV are not well understood.</p> <p>Results</p> <p>Using proteomics, we evaluated the effect of cp and ncp BVDV infection of bovine monocytes to determine their role in viral immune suppression and uncontrolled inflammation. Proteins were isolated by differential detergent fractionation and identified by 2D-LC ESI MS/MS. We identified 137 and 228 significantly altered bovine proteins due to ncp and cp BVDV infection, respectively. Functional analysis of these proteins using the Gene Ontology (GO) showed multiple under- and over- represented GO functions in molecular function, biological process and cellular component between the two BVDV biotypes. Analysis of the top immunological pathways affected by BVDV infection revealed that pathways representing macropinocytosis signalling, virus entry via endocytic pathway, integrin signalling and primary immunodeficiency signalling were identified only in ncp BVDV-infected monocytes. In contrast, pathways like actin cytoskeleton signalling, RhoA signalling, clathrin-mediated endocytosis signalling and interferon signalling were identified only in cp BDVD-infected cells. Of the six common pathways involved in cp and ncp BVDV infection, acute phase response signalling was the most significant for both BVDV biotypes. Although, most shared altered host proteins between both BVDV biotypes showed the same type of change, integrin alpha 2b (ITGA2B) and integrin beta 3 (ITGB3) were down- regulated by ncp BVDV and up- regulated by cp BVDV infection.</p> <p>Conclusions</p> <p>This study shows that, as we expected, there are significant functional differences in the host proteins that respond to cp or ncp BVDV infection. The combined use of GO and systems biology network modelling facilitated a better understanding of host-pathogen interactions.</p

Proteomics-Based Systems Biology Modeling of Bovine Germinal Vesicle Stage Oocyte and Cumulus Cell Interaction

BACKGROUND: Oocytes are the female gametes which establish the program of life after fertilization. Interactions between oocyte and the surrounding cumulus cells at germinal vesicle (GV) stage are considered essential for proper maturation or 'programming' of oocytes, which is crucial for normal fertilization and embryonic development. However, despite its importance, little is known about the molecular events and pathways involved in this bidirectional communication. METHODOLOGY/PRINCIPAL FINDINGS: We used differential detergent fractionation multidimensional protein identification technology (DDF-Mud PIT) on bovine GV oocyte and cumulus cells and identified 811 and 1247 proteins in GV oocyte and cumulus cells, respectively; 371 proteins were significantly differentially expressed between each cell type. Systems biology modeling, which included Gene Ontology (GO) and canonical genetic pathway analysis, showed that cumulus cells have higher expression of proteins involved in cell communication, generation of precursor metabolites and energy, as well as transport than GV oocytes. Our data also suggests a hypothesis that oocytes may depend on the presence of cumulus cells to generate specific cellular signals to coordinate their growth and maturation. CONCLUSIONS/SIGNIFICANCE: Systems biology modeling of bovine oocytes and cumulus cells in the context of GO and protein interaction networks identified the signaling pathways associated with the proteins involved in cell-to-cell signaling biological process that may have implications in oocyte competence and maturation. This first comprehensive systems biology modeling of bovine oocytes and cumulus cell proteomes not only provides a foundation for signaling and cell physiology at the GV stage of oocyte development, but are also valuable for comparative studies of other stages of oocyte development at the molecular level

Detection of antiviral tissue responses and increased cell stress in the pancreatic islets of newly diagnosed Type 1 diabetes patients: results from the DiViD study

This is the final version. Available from Frontiers Media via the DOI in this record.DATA AVAILABILITY STATEMENT: The datasets presented in this article are not readily available because of the sensitive nature of the data and possible high risks associated with patient confidentiality. Requests to access the datasets should be directed to the authors.Aims/hypothesis: The Diabetes Virus Detection (DiViD) study has suggested the presence of low-grade enteroviral infection in pancreatic tissue collected from six of six live adult patients newly diagnosed with type 1 diabetes. The present study aimed to compare the gene and protein expression of selected virally induced pathogen recognition receptors and interferon stimulated genes in islets from these newly diagnosed type 1 diabetes (DiViD) subjects vs age-matched non-diabetic (ND) controls. Methods: RNA was extracted from laser-captured islets and Affymetrix Human Gene 2.0 ST arrays used to obtain gene expression profiles. Lists of differentially expressed genes were subjected to a data-mining pipeline searching for enrichment of canonical pathways, KEGG pathways, Gene Ontologies, transcription factor binding sites and other upstream regulators. In addition, the presence and localisation of specific viral response proteins (PKR, MxA and MDA5) were examined by combined immunofluorescent labelling in sections of pancreatic tissue. Results: The data analysis and data mining process revealed a significant enrichment of gene ontologies covering viral reproduction and infectious cycles; peptide translation, elongation and initiation, as well as oxidoreductase activity. Enrichment was identified in the KEGG pathways for oxidative phosphorylation; ribosomal and metabolic activity; antigen processing and presentation and in canonical pathways for mitochondrial dysfunction, oxidative phosphorylation and EIF2 signaling. Protein Kinase R (PKR) expression did not differ between newly diagnosed type 1 diabetes and ND islets at the level of total RNA, but a small subset of b-cells displayed markedly increased PKR protein levels. These PKR+ b-cells correspond to those previously shown to contain the viral protein, VP1. RNA encoding MDA5 was increased significantly in newly diagnosed type 1 diabetes islets, and immunostaining of MDA5 protein was seen in a- and certain b-cells in both newly diagnosed type 1 diabetes and ND islets, but the expression was increased in b-cells in type 1 diabetes. In addition, an uncharacterised subset of synaptophysin positive, but islet hormone negative, cells expressed intense MDA5 staining and these were more prevalent in DiViD cases. MxA RNA was upregulated in newly diagnosed type 1 diabetes vs ND islets and MxA protein was detected exclusively in newly diagnosed type 1 diabetes b-cells. Conclusion/interpretation: The gene expression signatures reveal that pathways associated with cellular stress and increased immunological activity are enhanced in islets from newly diagnosed type 1 diabetes patients compared to controls. The increases in viral response proteins seen in b-cells in newly diagnosed type 1 diabetes provide clear evidence for the activation of IFN signalling pathways. As such, these data strengthen the hypothesis that an enteroviral infection of islet b-cells contributes to the pathogenesis of type 1 diabetes.European UnionJuvenile Diabetes Research Foundation InternationalJuvenile Diabetes Research Foundation InternationalMedical Research CouncilNational Institute for Health ResearchJuvenile Diabetes Research FoundationJuvenile Diabetes Research FoundationLeona M & Harry B Helmsley Charitable TrustSouth-Eastern Norway Regional Health AuthorityNovo Nordisk Foundatio

Flyttbara strandbastun

Examensarbetet går ut på att planera och konstruera ett lämpligt övningsprojekt som andra årets studerande kunde utföra i yrkesskolan Inveon på husteknikenheten. Eftersom ett större eget projekt samt kundprojekt inte alltid är möjligt så planerade jag ett mindre projekt som ändå var tillräckligt krävande och som täcker de examensgrunder som finns för andra årets studerande. Examensarbetet inleds med att undersöka vilka olika typer av projekt som kan vara aktuella i detta sammanhang. Arbetet utgår från vilka examensgrunder som är bestämda för utbildningen i husteknik vid Inveon. Från examensgrunderna kunde jag bestämma vilka material samt vilka konstruktionslösningar som skulle användas. Med beställare Niclas Isaksson, enhetschef för husteknik vid Inveon bestämde vi att byggnaderna som jag planerade skulle vara flyttbara, samt att specialtransport skulle undvikas på grund av tilläggskostnader. I examensarbetet undersöker jag också vilka lagar och regler som gäller för denna typ av byggnader, med fokus på olika kommuners byggnadsordningar, markanvändnings- och bygglagen samt vilka regler trafikverket ställer på transporten. I examensarbetet ingick att planera tre olika modeller som yrkesskolan kunde tillverka, viktberäkning gällande transporten samt en kostnadsberäkning för den slutliga produkten.Opinnäytetyö koostuu kouluprojektin suunnittelemisesta sekä kehittämisestä, jota ammattikoulu Inveonin rakennuspuoli voisi käyttää toisen vuoden opiskelijoiden koulutuksessa. Koska oma isompi projekti tai asiakasprojekti ei aina ole mahdollista, niin suunnittelin pienemmän projektin, joka olisi tarpeeksi vaativa sekä kattaisi ne tutkinnon perusteet jotka on asetettu toisen vuoden opiskelijoille. Opinnäytetyössä selvitetään, minkä tyyppiset projektit sopivat toisen asteen koulutukseen. Työ perustuu tutkinnon perusteista, jotka on määrätty Inveonin rakennuspuolen opiskelijoille. Tutkinnon perusteista pystyin päättämään mitä materiaaleja ja suunnitteluratkaisuja tulisi käyttää. Opinnäytetyön tilaajan, Inveon rakennuspuolen yksikön päällikkö Niclas Isakssonin kanssa päätimme, että rakennukset tulisi olla siirrettäviä ja että erikoiskuljetuksia tulisi välttää lisäkustannuksien takia. Opinnäytetyössä tutkin myös lakeja sekä säädöksiä, jotka koskevat tämäntyyppisiä rakennuksia, erityisesti eri kuntien rakennusmääräyksiä, maankäyttö- ja rakennuslakia, sekä liikenneviraston määräyksiä koskien kuljetusta. Opinnäytetyöhön kuului kolmen erilaisen mallin suunnittelu, joita ammattikoulu voisi valmistaa, painolaskelma koskien kuljetusta, sekä kustannuslaskelma lopullisesta tuotteesta.The aim of this thesis is to plan and design an appropriate practical project for second-year students specializing in construction at the vocational school Inveon. Since large school or customer projects are not always available or possible I planned a small, but challenging enough project which meets the national qualification requirements for second-year students. The thesis begins by examining the types of projects that may be relevant in this context. The work is based on the qualification requirements that are defined for the Construction Builder study programme at Inveon. Based on these requirements I could determine what materials and design solutions that would be used. Together with the client Niclas Isaksson, the Head of the Construction Builder programme at Inveon, we decided that the buildings I planned would be removable, and that special transportation would be avoided because of the additional costs. This thesis also investigates the laws and regulations which apply to these types of buildings, focusing especially on different municipal building regulations, land use and construction law, as well as the requirements regarding transportation set by the traffic departments. The thesis includes the planning of three different models which could be produced by the vocational school students, weight calculations regarding transportation, and an estimate of the costs of the final product

The effect of gluten-free diet on Th1--Th2--Th3-associated intestinal immune responses in celiac disease

Objective. To study T-helper (Th)1--Th2--Th3 gene activation profile in the small intestine and peripheral blood of children with celiac disease (CD) with special interest in the response to the gluten-free diet (GFD) treatment in order to elucidate an immune dysregulation not triggered by gluten. Material and methods. Small intestinal biopsies and venous blood were taken from seven children with CD (mean age: 8 years, four girls) at presentation and after 1 year of strict GFD. The Th1--Th2--Th3 gene expression profile was examined by real-time PCR arrays. The findings were compared with the corresponding expressions in peripheral blood and small intestinal biopsies from six reference children without CD (mean age: 6 years, four girls). Results. The Th1 gene expression profile including interferon (IFN)-gamma gamma, signal transducer and activator of transcription (STAT) 1 and interferon regulatory factor (IRF) 1 together with reduced interleukin (IL)-2 expression was pronounced in small intestinal biopsies from children with untreated CD. A downregulation of IFN-gamma gamma transcripts was seen after 1 year of GFD, but there was still increased expression of STAT1 and IRF1 in association with low IL-2 expression in spite of eliminated exposure to wheat gluten. By contrast, the decreased intestinal expression of Th2 gene markers observed at presentation was normalized with GFD. The alterations in the mucosal gene expression profile were not reflected in peripheral blood. Conclusion. The GFD did not correct the increased activation of the IFN-gamma gamma signaling pathway related markers and reduced IL-2 expression, suggesting that they represent an immune dysregulation not dependent on gluten exposure