1,720,993 research outputs found
Characterization of natural killer cells in healthy goats (capra aegagrus hircus)
Get PDFNK cells are essential cells of the innate immune system characterized by their ability to spontaneously eliminate virus-infected cells, tumor cells and all cells that are missing evidence of self. In addition to their immunoprotective properties, they have immunoregulatory functions, such as contributing to a tolerant immune environment during the establishment of pregnancy. NK cells have been thoroughly characterized in humans, mice, pigs, and cattle as well as to some extend in sheep. However, NK cells in goats have not been characterized before. In a world of virus- induced chronic infections, tumor manifestations and complex immune interactions, it is necessary to establish basal knowledge of these important immune cells, to meet the growing demand for immunological model organisms. The main objective of this project was to establish essential methods for describing the phenotype and function of goat NK cells. Several cell selection approaches were performed. One finding was that the anti-ovine NKp46 EC1.1 antibody seemed to result in a higher cell outcome than the anti-bovine NKp46 AKS6 antibody, when performing cell selection with magnetic beads. A functional effector cell assay, including cytotoxicity based on CD107a expression as well as IFN-γ production, was established in goats. Using this method, NKp46+CD3- cells showed typical NK-cell like properties like cytotoxic activity and cytokine production. An observed circulating population of NKp46+CD3+ cells were suspected to be γδT cells. Furthermore, a population of cells that were frequently contaminating NKp46+ cell cultures were CD3+ and were also hypothesized to be γδT cells. To resolve these hypotheses, cell sorting strategies were attempted with the aim to subsequently confirm genes specific for certain cell types by PCR. Time constraints of the research track project did not allow for the fulfillment of the cell sorting tasks and PCR; nevertheless, these data will provide a useful basis for the optimalization of these methods, and the verification of these heterogenous cell populations in goats
Immunresponser i atlantisk laks (Salmo salar) etter vaksinasjon mot og smitte med Salmonid alphavirus
Get PDFNorway has perfect conditions for Atlantic salmon (Salmo salar) production and is one of the largest producers of salmon for food consumption worldwide, but rearing fish at high densities also means that infectious disease agents can easily transmit between individuals. Vaccines have been an essential contributor to the steady increase in Atlantic salmon production over the last decades by reducing the severity and cases of diseases. To further increase vaccine efficacy and fish welfare, an increased knowledge about immune responses after vaccination and challenge is essential. This PhD work aimed to describe immune responses in Atlantic salmon following vaccination with a multivalent vaccine and challenge with Salmonid alpha virus (SAV).
Our results show that vaccination induces the production of vaccine specific antibodies, and that seroconversion occurs at around 2-4 weeks post vaccination. Additionally, both vaccination and infection with SAV induce the production of nonspecific, presumably polyreactive, antibodies. After we compared the serology assays data and gene transcript data from head kidney in paper I, a dissimilarity in the levels of immunoglobulin (Ig) transcripts and soluble Ig protein were evident, and in context with Ig sequencing of the CDR3 region (IgSeq) indicated a traffic of Ig producing B cells from head kidney in response to vaccination. Later results indicated a transfer of B cells from spleen to heart in response to SAV infection, and that this traffic is earlier in vaccinated individuals. Furthermore, results showed a colocalization of B cells and SAV transcribing cells, indicating that B cells traffic to the site of SAV replication to perform their function.
Lastly, the effect of constant light (CL) relative to vaccination were evaluated. Our results showed that the CL exposure highly influenced the gene transcript profile, and the results indicated that all fish experienced an immune suppression shortly after onset of CL. Gene transcript profiles also indicated that the vaccinated groups were more similar in their immune responses against SAV, regardless of the CL treatment prior to vaccination. In summary, our results indicate that vaccinated fish has an advantage against SAV infection and that they are less affected by the duration of CL prior to vaccination.Norge har perfekte forhold for produksjon av Atlantisk laks (Salmo salar) og er en av verdens største produsenter av laks, men oppdrett av fisk ved høy tetthet betyr imidlertid også at smittsomme sykdommer lett sprer seg mellom individer. Det har vært en jevn økning i produksjon av laks de siste tiårene, og vaksinering har vært en viktig bidragsyter til dette. Vaksiner har bidratt ved å redusere alvorlighetsgraden og antall tilfeller av ulike sykdommer. For å ytterligere øke vaksineeffektivitet og fiskevelferden er det viktig med mer kunnskap om immunresponser etter vaksinering og infeksjoner. Dette doktorgradsarbeidet hadde derav som mål å beskrive immunresponser i Atlantisk laks etter vaksinering med en multivalent vaksine og etter smitte med Salmonid alphavirus (SAV).
Våre resultater viste at vaksinasjon fører til produksjon av vaksinespesifikke antistoffer, og at serokonvertering skjer omtrent 2-4 uker etter vaksinasjon. I tillegg fører både vaksinasjon og SAV infeksjon til produksjon av ikke-spesifikke, antagelig polyreaktive, antistoffer. Etter å ha sammenlignet serologianalyse- og gentranskripsjon resultater fra hodenyrene i artikkel I så vi en forskjell i nivåene av immunglobulin (Ig) transkripter og løselig Ig-protein, som i sammenheng med Igsekvensering av CDR3-regionen (IgSeq) indikerte en trafikk av Ig-produserende Bceller vekk fra hodenyren i respons på vaksinasjon. Senere viste vi der er en trafikk av B-celler fra milt til hjerte i respons på SAV-infeksjon, og at det foregikk tidligere hos vaksinerte individer. En samlokalisering av Ig produserende B-celler og SAVtranskriberende celler innad i hjerte-vevet indikerte også at B-celler flytter seg til stedet for SAV-replikering for å utføre deres funksjon.
Til slutt ble effekten av konstant lys (CL) i forhold til vaksinasjonstidspunkt evaluert. Resultatene våre viste at CL-eksponeringen i stor grad påvirket gentranskript-profilen til fisken og at alle fisker opplevde en immunsuppresjon kort tid etter CL eksponering. Gentranskripsjon resultater fra de vaksinerte fiskene viste fisk med lignende transkripsjonsprofiler, som indikerer at vaksinert fisk hadde en mer lik immunrespons mot SAV, uavhengig av CL-eksponeringen før vaksinasjon. Kort oppsummert indikerer resultatene at vaksinert fisk har en fordel mot SAV og at de på sikt er mindre påvirket av lengden av konstant lys før vaksinasjon
A review on the use of Interleukin 2 and 12 as treatment for canine melanoma, osteosarcoma, and mast cell tumours
Get PDFLitteraturstudie 2025Melanom, osteosarkom og mastcelletumor er viktige kreftformer hos hund. De tradisjonelle behandlingspilarene: kirurgi, strålingsterapi og kjemoterapi; gir per i dag ikke tilfredsstillende resultater. Derfor har immunterapi blitt sett på som en mulig fjerde pilar innen kreftbehandling. Kreft har mange metoder for å unngå immunsystemet. Derfor har cytokiner blitt undersøkt på grunn av deres evne til å regulere immunrespons, og potensial for å rette immunsystemet mot tumorcellene. Cytokinbehandling er derimot ikke problemfritt, da de terapeutiske dosene nødvendig, gir unødvendig mange bivirkninger.
I denne litteraturstudien undersøkes bruken av IL-2 og IL-12 på melanom, osteosarkom og mastcelletumor hos hund; der det blir sett på ulike interleukinformuleringer, administrasjonsmetoder, behandlingsregimer, antitumoreffekt og uønskede virkninger. Det ble gjennomgått 13 artikler fordelt på kreftformene, som brukte IL-2 og/ eller IL-12 i ulike behandlingsregimer.
Det er ikke mulig å konkludere med bruken av cytokiner til kreftbehandling, verken på tvers av cytokiner eller kreftformer, men potensialet er lovende. Det er behov for mer forskning på feltet, for optimalisering av behandlingsregimer. Det trengs også studier med større studieutvalg for bedre statistisk grunnlag. Selv om det ikke er mulig å dra en definitiv konklusjon, kan cytokiner ha en lovende rolle som adjuvans til kreftbehandlingsregimer for hunder med melanom, osteosarkom og mastcelletumor.Melanoma, osteosarcoma and mast cell tumours are important canine cancers. The traditional pillars of treatment: surgery, radiation and chemotherapy; do not, as of today, yield satisfying results. Immunotherapy has therefore been recognized as a possible fourth pillar. Cancer has many methods for evading the immune system. Cytokines have therefore been researched, because of their ability to regulate immune responses, and potentially direct the immune system against tumour cells. However, cytokine treatment is not flawless; the necessary therapeutic doses result in unnecessary adverse effects.
This review examines the use of IL-2 and IL-12 on canine melanoma, osteosarcoma and mast cell tumours, assessing different interleukin formulations, methods of administration, treatment protocols, anti-tumour effect, and adverse events.
13 articles on the different cancer types and cytokines, used in different treatment protocols, were reviewed.
Definitive conclusions cannot be made; neither across different cytokines nor cancer types, though the potential remains promising. Additional studies are needed for optimization of treatment protocols. There is also a need for studies with a larger study population, for better statistical basis. Although definitive conclusions cannot be drawn, cytokines may have a promising role as an adjuvant for cancer treatment protocols for canine melanoma, osteosarcoma and mast cell tumours.VE
Activation and recirculation of bovine natural killer cells in steady state and inflammation
Get PDFNatural killer (NK) cells are innate lymphocytes that act as early responders in the immune response by means of cytotoxicity and production of immunoregulatory cytokines. NK cells in cattle were described a decade ago, and since then, significant progress has been made in the characterization of NK cells. However, NK cell activation and recirculation in vivo remains largely unknown. This thesis describes the properties of activated NK cells at steady-state and inflammation, and investigates routes of NK cell recirculation in the bovine model.NK-celler (naturlige dreperceller) er lymfocytter i det medfødte immunforsvaret som bidrar tidlig i en immunrespons ved hjelp av celle-drap og produksjon av immunregulatoriske cytokiner. NK-celler hos storfe ble beskrevet for et tiår siden, og siden den gang har det vært en betydelig fremgang i karakteriseringen av disse cellene. Aktiveringen og resirkuleringen av NK
celler in vivo er likevel i stor grad fortsatt ukjent. Denne avhandlingen beskriver egenskaper ved aktiverte NK-celler under steady-state (stabil tilstand) og ved inflammasjon, og undersøker NK celle resirkulering i en storfe modell
Effect of different modes of naturalization on expression of barrier-related genes in the murine intestine
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A comparison between the immune response in macrophages when exposed to two different strains of Streptococcus agalactiae
Get PDFAbstract of thesis Mastitis is the most common disease in the dairy industry today. In the mammary gland, macrophages play a leading role in the immune response as first line defence against invading udder pathogens. Different candidate genes associated with the response against udder pathogens are detected, where some of these candidate genes represent pro-inflammatory cytokines produced by macrophages. However, recent studies from our group indicate that macrophages infected with bacteria such as Staphylococcus aureus also produce anti-inflammatory cytokines in an alternative response. The alternatively activated macrophages counteract many pro- inflammatory mechanisms, and this may be a strategy for udder pathogens to evade the host immune response. A key question is whether macrophages exposed to Streptococcus agalactiae will have a greater inclination towards alternative than classical activation, and if this diversify between the different sequence types (ST) of the bacteria. We investigated the early phase response of bovine monocyte-derived macrophages infected in vitro with two different sequence types of live Streptococcus agalactiae (ST103 and ST12) in vitro, by examining the transcription level of six macrophage-associated cytokines. First, we isolated monocyte-derived macrophages from six healthy Norwegian Red (NR) cows aged 2,5-7 years, and for each individual animal the immature macrophages were divided into four classes. Two classes were infected in vitro with either of the two S. agalactiae strains with a multiplicity of infection (MOI) of 1, then incubated for 1 hour before penicillin/streptomycin was added in each well, and further incubated for 5 hours (a total of 6 hours). The third cell class was exposed to Lipopolysaccharides (LPS) (positive control) and the last class was left uninfected (negative control), and both positive and negative control were treated equally as the infected cells with penicillin/streptomycin and incubated for a total of 6 hours. Originally, we planned to compare the early phase response also with monocyte-derived macrophages infected with Staphylococcus aureus in vitro, but we were not able to continue this work due to a non-reproducible method when infecting the cells with S. aureus. Consequently, this part of the study was abandoned. Further we isolated total RNA from the cells infected with S. agalactiae ST12 and ST103, and measured the transcript levels of Tumor Necrosis Factor α (TNFα), Interleukin 1β (IL-1β), Interleukin 6 (IL-6), Interleukin 8 (IL-8), Interleukin 10 (IL-10) and Transforming growth factor β1 (TGFβ1) by Reverse transcription-quantitative Polymerase Chain Reaction (RT-qPCR). TNF-α, IL-1β, IL-6, IL-8 and IL-10 were significantly up-regulated by ST12, ST103 and LPS compared to the negative control. IL-6 and IL-10 displayed different responses both between ST103 and LPS, and between ST12 and LPS, with high levels of IL-10 (anti-inflammatory cytokine) and low levels of IL-6 (pro-inflammatory cytokine) in the cells infected with S. agalactiae compared to cells activated by LPS. TGFβ1 were significantly down-regulated only in the macrophages infected with ST12. When comparing the transcription levels of the cytokines between the macrophages infected with the two strains of S. agalactiae, we did not observe significantly different expression of any of the six cytokines. Thus, we know there is activation of both pro-inflammatory and anti-inflammatory cytokines when infected with S. agalactiae, and that there is a decrease in the anti-inflammatory signal of TGFβ1 only in macrophages infected with ST12. We also propose that the macrophages infected with bacteria might be activated in the alternative pathway compared to macrophages activated by LPS, but this field of study needs further investigation compared to macrophages activated by LPS, but this field of study needs further investigation
Oppstalling av laboratoriemus i et naturlig habitat : virkning på immunsystem, tarmmikrobiota og utvikling av kolorektal kreft
Get PDFThe natural habitat for the house mouse is on the ground, typically close to humans and their livestock and hence surrounded by a rich microbial diversity that throughout evolutionary history has driven the adaptation of the house mouse. It is thus paradoxical that almost without exception, experimental disease studies using this mammalian model take place in perfect isolation from the outer microbial world. The end goal for preclinical research, humans, rarely live in microbial isolation, although lifestyles can arguably be said to vary on a scale. To develop a preclinical model that better resemble realistic lifestyles of mammals, we have established a system where laboratory mice are raised under a full set of environmental conditions present in a typical farmyard habitat for the house mouse. We call the process feralization, and the first paper covered by this thesis show the resulting mammal display more functionally mature states of immune cells and a diverse gut microbiota, likely surpassing conventional laboratory mice in resembling responses of free-living mice.
Furthermore, we demonstrated the use of this animal modelling approach that recapitulates realistic disease responses in a naturalized mammal. We first established a protocol of the AOM/DSS model for colorectal cancer (CRC) induction in mice using a lower-than-usual dose of DSS that is presented in paper II. In paper III, we employed the AOM/DSS model, as well as a previously established genetic Min/+ model of CRC, in a feralization system. We showed that the mice feralized in a farmyard-type habitat were protected against colorectal carcinogenesis compared to conventionally reared laboratory mice. Moreover, our feralization model allows for full control of the timing of microbial exposure. We took advantage of this by including groups of mice that were either born in the farmyard habitat or introduced to it in later life, demonstrating that neonatal microbial exposure was not essential for the CRC protection. The findings were supported by changes in gut microbiota profiles, as well as immunophenotypes indicative of antigenic experience in the feralized mice.
In currently unpublished work, we aimed to narrow in on mechanisms for the protective effects conveyed by feralization in the intestines. Assays investigating mucus layer properties showed no differences following feralization, yet a few genes in the colon mucosa related to barrier function were found to be significantly upregulated between feralized and conventional laboratory mice. Further assessments are needed to elaborate on the mechanisms underlying the beneficial effects of feralization.Det naturlige habitatet for husmus er typisk nær mennesker og deres husdyr, og husmusa er gjennom evolusjon tilpasset et slikt rikt mikrobielt levemiljø. Det er derfor paradoksalt at eksperimentelle studier der mus brukes for å studere sykdomsmekanismer, omtrent uten unntak foregår i perfekt isolasjon fra den ytre, mikrobielle verden. Målet for prekliniske studier ved bruk av forsøksmus er typisk å overføre funnene til relevans for mennesker, men mennesker lever sjeldent i mikrobiell isolasjon, selv om individuelle livsstiler varierer stort. For å utvikle en preklinisk musemodell som mer realistisk representerer naturlige livstiler hos pattedyr, har vi etablert et system der laboratoriemus fostres opp i en mer naturlig situasjon, der habitatet deres er beriket med elementer som er tilstede i et typisk gårdsmiljø. Vi har kalt denne prosessen «feralisering», og den første artikkelen som omfattes av denne avhandlingen demonstrerer at de «feraliserte» musene viser tegn til mer funksjonelt modne immunceller og en rikere tarmmikrobiota sammenliknet med laboratoriemus oppstallet under tradisjonelle, rene forhold.
Videre har vi demonstrert bruken av feraliserings-systemet som kan benyttes til å studere realistiske sykdomsresponser i et naturalisert pattedyr. Først etablerte vi en protokoll for kjemisk induksjon av kolorektalkreft hos mus, ved å bruke en lavere-enn-normal dose av DSS i en AOM/DSS modell. Dette arbeidet er presentert i artikkel II. I artikkel III benyttet vi AOM/DSS modellen, så vel som en tidligere etablert genetisk Min/+ musemodell for kolorektalkreft, i feraliseringssystemet. Vi demonstrerte at musene som ble feralisert i et naturalistisk gårdsmiljø var beskyttet mot tykktarmskreft, sammenliknet med laboratoriemus oppstallet under tradisjonelle, rene forhold. Videre tillater feraliserings-systemet full kontroll av timingen for mikrobiell eksponering. Vi utnyttet dette ved å inkludere en gruppe av mus som enten var født i gårdsmiljøet, eller introdusert dit senere i livet. Med dette viste vi at neonatal mikrobiell eksponering ikke var essensiell for beskyttelse mot kolorektal kreft. Funnene ble støttet av endringer i tarmmikrobiotaprofiler, samt immunofenotyper som kan indikere at de har blitt eksponert for antigener.
I nåværende upublisert arbeid hadde vi som mål å undersøke mulige mekanismer for hvordan feralisering i et gårdsmiljø kunne gi beskyttelse mot kolorektal kreft. Undersøkelser av slimlaget i tarm viste ingen forskjeller etter feralisering. Likevel var det none få gener i tykktarmsslimhinnen relatert til barrierefunksjon som var oppregulert i feraliserte mus sammenliknet med konvensjonelt oppstallede laboratoriemus. Videre undersøkelser er nødvendig for å kunne utdype mer når det gjelder mekanismene bak feraliserings beskyttende effekt
Deteksjon av antistoffer i atlantisk laks med et kulebasert multipleks immunoassay
Get PDFViral diseases are among of the main challenges in aquaculture in Norway today. Heart- and skeletal muscle inflammation, caused by Piscine orthoreovirus (PRV), and pancreas disease (PD), caused by Salmonid alphavirus (SAV), both cause huge losses of farmed Atlantic salmon. Vaccination is a possible solution that is already contributing to the control of bacterial diseases. Unfortunately, making effective vaccines against viral diseases has been challenging and information about which immune mechanisms that are involved in protection is lacking.
Antibodies can mediate complete protection against some diseases, and antibody levels after vaccination can correlate with protection, even if the protection is mediated by other mechanisms. In addition, antibody detection can be used in disease surveillance to determine if a fish population is or has been infected with a virus. Unfortunately, measuring antibody levels in salmon is not straightforward. Such measurements have not been widely used, and often show high levels of background binding. An important reason for this is that fish antibodies are of the IgM type. IgM is less specific than IgG, the dominating antibody in mammals. Therefore, other methods, like PCR and histology, are more used in diagnostics in Atlantic salmon. In this work, we have for the first time used an assay based on microscopic magnetic beads conjugated with antigen to measure antibody levels in Atlantic salmon. This method has a high sensitivity and can be used to measure antibodies against several proteins simultaneously in the same sample.
By using this method, we have detected antibodies against PRV and SAV. In paper I and II, plasma from two PRV challenge trials was used to detect antibodies against the PRV proteins μ1c, μNS and σ1. We also detected antibodies against PRV1- σ1 in plasma from PRV-3-infected rainbow trout. There have been no previous publications detecting antibodies against PRV. The peak antibody level coincided with decreased pathology in the heart. In addition to showing virus-specific antibodies, our results show an increase in non-specific antibodies in PRV-infected salmon. This unspecific binding, but not the virus-specific binding, was decreased by heat treatment of samples. The nonspecific antibodies could be so-called polyreactive antibodies. Polyreactive antibodies could be crucial in protecting the salmon against infections before the adaptive immune system has had time to react, but their function, as well as the function of virus-specific antibodies during a PRV infection is unclear.
In paper III, we detected antibodies against whole SAV particles disrupted with Triton-X. The antibody binding increased from between week three and week six after the introduction of SAVinjected shedder fish. The SAV particles also worked well with little background binding when analyzing plasma from a PD outbreak in the field. Antibodies were detected in most fish from four weeks after the start of the outbreak, and the antibody level stayed elevated until the last sampling point at 15 weeks. At this time point, most samples were negative for virus when analyzed with RTqPCR, showing that serology has a longer window of detection compared to detection of viral RNA by RT-qPCR.
These results show that both virus-specific and non-specific antibodies can be produced after infection in Atlantic salmon. The method used is well suited for antibody detection in salmon but can be complicated by the presence of non-specific antibodies. It is therefore crucial to optimize the antigens for detection of specific antibodies.Virussykdommer er en av de store utfordringene i oppdrettsnæringa i Norge i dag. Hjerte- og skjelettmuskelbetennelse, forårsaket av Piscine orthoreovirus (PRV), og pankreas sykdom (PD), forårsaket av Salmonid alphavirus (SAV), fører begge til store tap av atlantisk laks i oppdrett. Vaksinering er en mulig løsning som allerede har bidratt til god kontroll på bakterielle infeksjoner hos laks. Dessverre er det utfordrende å lage vaksiner som virker godt mot virusinfeksjoner. Informasjon om hvilke immunfunksjoner som er involvert i beskyttelse er også mangelfull.
Antistoffer kan gi full beskyttelse mot noen sykdommer, og antistoffnivået etter vaksinering kan korrelere med beskyttelsen, også om beskyttelsen skyldes andre immunmekanismer. I tillegg kan antistoffdeteksjon brukes i sykdomsovervåkning for å finne ut om fiskepopulasjoner er eller har vært infisert med et virus. Dessverre er det ikke rett fram å måle antistoffnivå hos laks. Antistoffmåling er lite brukt og det er ofte mye bakgrunnsbinding. En viktig grunn til dette er at fiskens antistoff er av typen IgM, som er mindre spesifikk enn IgG i pattedyr. Derfor er andre metoder, som PCR og histologi, mest brukt til påvisning av virussykdom hos laks. I dette arbeidet har vi for første gang brukt et assay basert på mikroskopiske magnetiske kuler konjugert med antigen til å måle antistoffnivå i laks. Denne metoden har høy sensitivitet og gir muligheten til å måle antistoffer mot mange proteiner i samme prøve samtidig
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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