A lymphofollicular microenvironment is required for pathological prion protein deposition in chronically inflamed tissues from scrapie-affected sheep

In sheep scrapie, pathological prion protein (PrPSc) deposition occurs in the lymphoreticular and central nervous systems. We investigated PrPSc distribution in scrapie- affected sheep showing simultaneous evidence of chronic lymphofollicular, lymphoproliferative/non-lymphofollicular,and/or granulomatous inflammations in their mammary gland, lung, and ileum. To do this, PrPSc detection was carried out via immunohistochemistry and Western Blotting techniques, as well as through inflammatory cell immunophenotyping. Expression studies of gene coding for biological factors modulating the host’s inflammatory response were also carried out. We demonstrated that ectopic PrPSc deposition occurs exclusively in the context of lymphofollicular inflammatory sites, inside newly formed and well-organized lymphoid follicles harboring follicular dendritic cells. On the contrary, no PrPSc deposition was detected in granulomas, even when they were closely located to newly formed lymphoid follicles. A significantly more consistent expression of lymphotoxin α and β mRNA was detected in lymphofollicular inflammation compared to the other two types, with lymphotoxin α and β signaling new lymphoid follicles’ formation and, likely, the occurrence of ectopic PrPSc deposition inside them. Our findings suggest that, in sheep co-affected by scrapie and chronic inflammatory conditions, only newly formed lymphoid follicles provide a suitable micro-environment that supports the scrapie agent’s replication in inflammatory sites, with an increased risk of prion shedding through body secretions/excretions.[...

Application of a New Method for GWAS in a Related Case/Control Sample with Known Pedigree Structure: Identification of New Loci for Nephrolithiasis

In contrast to large GWA studies based on thousands of individuals and large meta-analyses combining GWAS results, we analyzed a small case/control sample for uric acid nephrolithiasis. Our cohort of closely related individuals is derived from a small, genetically isolated village in Sardinia, with well-characterized genealogical data linking the extant population up to the 16th century. It is expected that the number of risk alleles involved in complex disorders is smaller in isolated founder populations than in more diverse populations, and the power to detect association with complex traits may be increased when related, homogeneous affected individuals are selected, as they are more likely to be enriched with and share specific risk variants than are unrelated, affected individuals from the general population. When related individuals are included in an association study, correlations among relatives must be accurately taken into account to ensure validity of the results. A recently proposed association method uses an empirical genotypic covariance matrix estimated from genome-screen data to allow for additional population structure and cryptic relatedness that may not be captured by the genealogical data. We apply the method to our data, and we also investigate the properties of the method, as well as other association methods, in our highly inbred population, as previous applications were to outbred samples. The more promising regions identified in our initial study in the genetic isolate were then further investigated in an independent sample collected from the Italian population. Among the loci that showed association in this study, we observed evidence of a possible involvement of the region encompassing the gene LRRC16A, already associated to serum uric acid levels in a large meta-analysis of 14 GWAS, suggesting that this locus might lead a pathway for uric acid metabolism that may be involved in gout as well as in nephrolithiasis

Intraepithelial and Interstitial Deposition of Pathological Prion Protein in Kidneys of Scrapie-Affected Sheep

Prions have been documented in extra-neuronal and extra-lymphatic tissues of humans and various ruminants affected by Transmissible Spongiform Encephalopathy (TSE). The presence of prion infectivity detected in cervid and ovine blood tempted us to reason that kidney, the organ filtrating blood derived proteins, may accumulate disease associated PrPSc. We collected and screened kidneys of experimentally, naturally scrapie-affected and control sheep for renal deposition of PrPSc from distinct, geographically separated flocks. By performing Western blot, PET blot analysis and immunohistochemistry we found intraepithelial (cortex, medulla and papilla) and occasional interstitial (papilla) deposition of PrPSc in kidneys of scrapie-affected sheep. Interestingly, glomerula lacked detectable signals indicative of PrPSc. PrPSc was also detected in kidneys of subclinical sheep, but to significantly lower degree. Depending on the stage of the disease the incidence of PrPSc in kidney varied from approximately 27% (subclinical) to 73.6% (clinical) in naturally scrapie-affected sheep. Kidneys from flocks without scrapie outbreak were devoid of PrPSc. Here we demonstrate unexpectedly frequent deposition of high levels of PrPSc in ovine kidneys of various flocks. Renal deposition of PrPSc is likely to be a pre-requisite enabling prionuria, a possible co-factor of horizontal prion-transmission in sheep

PrP(Sc) deposition in nervous tissues without lymphoid tissue involvement is frequently found in ARQ/ARQ Sarda breed sheep preclinically affected with natural scrapie

The pathogenesis of natural scrapie in Sarda breed sheep was investigated in 1050 asymptomatic and 49 sick sheep from scrapie-affected flocks.Central and peripheral nervous system, along with lymphoreticular system (LRS)tissues, were subjected to immunohistochemistry (IHC) and Western-blotting(WB) for detection of pathological isoform of the prion protein (PrPSc). A total of 69 of the 1050 clinically healthy sheep were found to be infected with scrapie, with PrPSc being detected in both the central nervous system (CNS) and enteric nervous system (ENS) plexuses of 60 of the sheep, while IHC and WB yielded evidence of PrPSc deposition only in lymphoid tissues of the remaining 9 clinically healthy sheep. PrPSc was also detected in the CNS, as well as in ENS plexuses from allof the 49 clinically affected sheep. Nevertheless, 18 of the 69 clinically healthyanimals (26%, 17 ARQ/ARQ and 1 ARQ/AHQ sheep), along with 3 ARQ/ARQ sheep (6%) of the clinically affected group, showed no IHC or WB evidenceof PrPSc in lymphoid tissues, but PrPSc could be still detected in their CNS and ENSplexuses. The study demonstrates dual CNS and ENS PrPSc deposition in Sardasheep with scrapie, in spite of an apparent lack of lymphoid tissue involvement ina number of cases.[...

Lipid profiles in brains from sheep with natural scrapie

Prion diseases are fatal neurodegenerative disorders affecting many mammals, ovine scrapie being the archetypal prion disease. Several independent studies in murine and cell-based models of scrapie have highlighted the presence of a link between prion generation and lipid alterations; yet, no data on natural disease are available. In this study we investigated levels of total lipids and cholesterol as well as profiles of fatty acids in brain homogenates from symptomatic and asymptomatic scrapie-infected sheep vs. healthy sheep, all belonging to the same flock. Lipid extracts were analyzed by means of gas chromatography and high performance liquid chromatography. Data of fatty acids were submitted to multivariate statistical analysis to give a picture of the brain lipid profiles of sheep. Interestingly, results revealed abnormalities in the brain fatty acid unsaturation of infected/symptomatic animals. Significant reduction of monoene 18:1 n-9 was detected in brain lipids from infected/symptomatic sheep, as compared to healthy and infected/asymptomatic animals, and this alteration occurred in combination with a significant increase in 18:0 level. The unsupervised Principal Component Analysis showed that infected/symptomatic and healthy sheep samples lie in two different regions of the plot, infected/asymptomatic lie mostly next to healthy. The increase of cerebral saturated fatty acids provides a rough indication of presumed alterations in lipid raft domains of nervous cells during scrapie, suggesting that they may exist in a notable viscous liquid-ordered state. Such physicochemical alteration would have a profound impact on the raft thermodynamic properties, its spatial organization, and signal transduction, all potentially relevant for prion generation

Pathology in Practice - Epitheliogenesis Imperfecta in a Sarda Breed Lamb

Morphologic Diagnosis and Case SummaryMorphologic diagnosis: Congenital, focal aplasia of the epidermis and adnexal structures, associated with superficial necrosis and extensive dermal inflammation.Case summary: Epitheliogenesis imperfecta (EI) in a lamb.[...

Gene expression profiling on sheep brain reveals differential transcripts in scrapie-affected/not-affected animals.

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