Expression of selected genes isolated from whole blood, liver and obex in lambs with experimental classical scrapie and healthy controls, showing a systemic innate immune response at the clinical end-stage.

BACKGROUND: Incubation period, disease progression, pathology and clinical presentation of classical scrapie in sheep are highly dependent on PRNP genotype, time and route of inoculation and prion strain. Our experimental model with pre-colostrum inoculation of homozygous VRQ lambs has shown to be an effective model with extensive PrPSc dissemination in lymphatic tissue and a short incubation period with severe clinical disease. Serum protein analysis has shown an elevation of acute phase proteins in the clinical stages of this experimental model, and here, we investigate changes in gene expression in whole blood, liver and brain. RESULTS: The animals in the scrapie group showed severe signs of illness 22 weeks post inoculation necessitating euthanasia at 23 weeks post inoculation. This severe clinical presentation was accompanied by changes in expression of several genes. The following genes were differentially expressed in whole blood: TLR2, TLR4, C3, IL1B, LF and SAA, in liver tissue, the following genes differentially expressed: TNF-α, SAA, HP, CP, AAT, TTR and TF, and in the brain tissue, the following genes were differentially expressed: HP, CP, ALB and TTR. CONCLUSIONS: We report a strong and evident transcriptional innate immune response in the terminal stage of classical scrapie in these animals. The PRNP genotype and time of inoculation are believed to contribute to the clinical presentation, including the extensive dissemination of PrPSc throughout the lymphatic tissue.

Presence of an acute phase response in sheep with clinical classical scrapie.

BACKGROUND: Work with experimental scrapie in sheep has been performed on-site for many years including studies on PrPSc dissemination and histopathology of organs and tissues both at preclinical and clinical stages. In this work serum was sampled at regular intervals from lambs which were infected immediately after birth and from parallel healthy controls, and examined for acute phase proteins. In contrast to earlier experiments, which extensively studied PrPSc dissemination and histopathology in peripheral tissues and brain, this experiment is focusing on examination of serum for non-PrPSc markers that discriminates the two groups, and give insight into other on-going processes detectable in serum samples. RESULTS: There was clear evidence of an acute phase response in sheep with clinical scrapie, both experimental and natural. All the three proteins, ceruloplasmin, haptoglobin and serum amyloid A, were increased at the clinical stage of scrapie. CONCLUSION: There was evidence of a systemic measurable acute phase response at the clinical terminal end-stage of classical scrapie.

Investigation of serum protein profiles in scrapie infected sheep by means of SELDI-TOF-MS and multivariate data analysis.

BACKGROUND: Classical scrapie in sheep is a fatal neurodegenerative disease associated with the conversion PrPC to PrPSc. Much is known about genetic susceptibility, uptake and dissemination of PrPSc in the body, but many aspects of prion diseases are still unknown. Different proteomic techniques have been used during the last decade to investigate differences in protein profiles between affected animals and healthy controls. We have investigated the protein profiles in serum of sheep with scrapie and healthy controls by SELDI-TOF-MS and LC-MS/MS. Latent Variable methods such as Principal Component Analysis, Partial Least Squares-Discriminant Analysis and Target Projection methods were used to describe the MS data. RESULTS: The serum proteomic profiles showed variable differences between the groups both throughout the incubation period and at the clinical end stage of scrapie. At the end stage, the target projection model separated the two groups with a sensitivity of 97.8%, and serum amyloid A was identified as one of the protein peaks that differed significantly between the groups. CONCLUSIONS: At the clinical end stage of classical scrapie, ten SELDI peaks significantly discriminated the scrapie group from the healthy controls. During the non-clinical incubation period, individual SELDI peaks were differently expressed between the groups at different time points. Investigations of differences in -omic profiles can contribute to new insights into the underlying disease processes and pathways, and advance our understanding of prion diseases, but comparison and validation across laboratories is difficult and challenging.

cDNA representational difference analysis of ileal Peyer's patches in lambs after oral inoculation with scrapie.

cDNA representational difference analysis (RDA) was used to study gene expression profiles in the ileal Peyer's patch of a lamb 1 week after oral inoculation with the scrapie agent. Twenty-five differentially expressed cDNA fragments were identified and cloned. Sequence analysis indicated seven novel gene sequences. Other clones shared sequence homology with genes encoding ribosomal and mitochondrial proteins, the translation initiation factor EIF4GII and the bovine pancreatic thread protein. Reverse Northern was used to confirm the differential expression in another four lambs inoculated with scrapie and the tissue distribution of the novel genes was examined using Northern blot analysis.