Tilstedeværelsen av en akutt fase-reaksjon hos lam med eksperimentell klassisk skrapesjuke indikerer et skifte mot en pro-inflammatorisk tilstand i det kliniske endestadiet

Abstract

Classical scrapie in sheep is a transmissible and fatal neurodegenerative disease caused by the self-replicating and infectious prion protein, PrPSc, which is a conformational variant of the normal cellular prion protein, PrPC. The prion protein is a highly conserved glycoprotein encoded by the PRNP gene and therefore within the same host both PrPC and PRPSC have the same unique amino acid sequence and they only differ in their three-dimensional folded structure. Specific mutations at codons 136, 154 and 171 of the PRNP gene leads to single amino acid substitutions, and the most common polymorphisms give rise to five possible alleles and 15 PRNP genotypes found in sheep. The different alleles are highly associated with levels of susceptibility to classical scrapie, where A136R154R171 allele provides high genetic resistance and V136R154Q171 allele results in highly susceptible animals. On the basis of this association between PRNP genotype and susceptibility, many EU MSs have implemented national breeding for resistance programme with the aim of increasing distribution of ARR allele and reducing the distribution of VRQ allele. For almost 20 years, the EU TSE regulation has required surveillance within each country to establish prevalence of prion diseases and the different PRNP genotypes. Classical scrapie has a widespread distribution and incidence rate fluctuates due to the complex interaction between prion and host factors, and prevalence can only be estimated by ante mortem testing through active and passive surveillance. Transmission between sheep occurs through direct and indirect contact, and PrPSc can remain infective in the environment for years. The most common route of infection is the oral route, and infected animals can excrete PrPSc through foetal membranes and fluids, saliva, urine, faeces, and milk. Pathogenesis is highly influenced by PRNP genotype, as animals of the most susceptible genotypes have the most effective uptake of PrPSc across small intestine followed by an extensive dissemination and involvement of the SLOs, and an early neuroinvasion with spread of PrPSc within the CNS. The susceptible genotypes will contribute the most to spread of infectivity and environmental contamination. This work describes the results from experimental classical scrapie where homozygous VRQ lambs were inoculated orally at birth with homogenated brain material from either healthy sheep or from natural cases of classical scrapie. This resulted in a worst-case scenario type of classical scrapie with sudden onset of severe clinical signs at 22 wpi followed by a rapid deterioration and euthanasia at 23 wpi. Serum samples were collected at regular intervals and tissue samples from brain and liver were sampled at post mortem examination. Proteomic examinations of serum revealed a downregulation of several protein peaks during the pre-symptomatic incubation period in the scrapie affected group compared to the control group, and a shift to upregulation of protein peaks onwards from 22 wpi. Genomic examinations of serum samples showed a slight downregulation IL1B and TLR4 at 16 wpi, followed by a change at 22 wpi with upregulation of genes encoding TLRs, C3 and APPs. Genomic examination of liver and brain tissues showed an alteration in gene expression of APPs in accordance with an APR. Serum analyses of different APPs showed increased levels of the positive APPs and a reduced concentration of negative APPs. These findings are indicative of a shift from anti-inflammatory to pro-inflammatory systemic innate immune response that coincide with the onset of debilitating clinical disease. In neurodegenerative diseases, the innate immune response in the CNS has a key role in both onset and progression of disease and resolution of inflammation. The accumulation of PrPSc in the CNS has been associated with a chronic activation of the innate immune response, pro-inflammatory activation of microglia, neuroinflammation, and neurodegeneration. The diseases phenotype registered in this work is a result of PRNP genotype, and time and dose of inoculation, which can occur naturally if the right circumstances are in place. New-born homozygous VRQ lambs from an infected dam can get infected at birth. These cases could develop a similar disease progression as described in this work, resulting in an efficient and fast uptake and widespread peripheral and central dissemination of PrPSc, and clinical disease at a young age. These cases would present as a diagnostic challenge and easily missed as classical scrapie. Due to their young age, these cases would not be sampled through active surveillance. If incubation period extends commercial lifespan, these lambs would be slaughtered for human consumption, and due to their PRNP genotype, prions would enter the food chain. Control of classical scrapie can probably not be achieved by absence of infectivity, but absence of clinical disease is possible through breeding for resistance which will provide flock immunity to classical scrapie.Klassisk skrapesyke hos sau er en overførbar og dødelig nevrodegenerativ sykdom forårsaket av det selvrepliserende og smittsomme prionproteinet, PrPSc, som er en variant av det normale cellulære prionproteinet, PrPC. Prionproteinet er et glykoprotein som er kodet for av PRNP-genet. Dette betyr at PrPC og PRPSC hos samme verten, har den samme unike aminosyresekvensen og det er kun den tredimensjonale strukturen som skiller dem. Spesifikke mutasjoner ved kodonene 136, 154 og 171 i PRNP-genet fører til substitusjoner av enkelte aminosyrer, og de vanligste polymorfismer gir opphav til fem mulige alleler, og 15 PRNP-genotyper hos sau. De forskjellige allelene er assosiert med nivå av mottakelighet for klassisk skrapesyke, og A136R154R171-allel fører til genetisk resistens, og V136R154Q171-allel gir høy mottagelighet. På bakgrunn av denne sammenhengen mellom PRNP-genotype og mottakelighet, har mange EU medlemsland innført nasjonale avlsprogram som har mål om å øke utbredelsen av ARR-allel, og samtidig en reduksjon av VRQ-allel. I snart 20 år har EUs TSE-regelverk krevd nasjonale overvåkingsprogram for å bestemme forekomsten av prionsykdommer og kartlegge utbredelsen av de forskjellige PRNP-genotypene. Klassisk skrapesyke er utbredt, men forekomsten vil variere med bakgrunn i det komplekse samspillet mellom prionprotein og vertsfaktorer. Prevalens kan estimeres gjennom ante mortem testing i forbindelse med aktivt og passivt overvåkingsprogram. Smitteoverføring mellom sau skjer ved direkte og indirekte kontakt, og PrPSc er smittsomt i flere år i miljøet. Den vanligste infeksjonsveien er gjennom oralt inntak, og dyr kan skille ut smittsomt PrPSc via fosterhinner og væsker, spytt, urin, feces og melk, og nivå er styrt av PRNP genotype.Research Council of Norwa