LPS-induced systemic inflammation reveals an immunomodulatory role for the prion protein at the blood-brain interface

Abstract Background The cellular prion protein (PrPC) is an evolutionary conserved protein abundantly expressed not only in the central nervous system but also peripherally including the immune system. A line of Norwegian dairy goats naturally devoid of PrPC (PRNP Ter/Ter) provides a novel model for studying PrPC physiology. Methods In order to explore putative roles for PrPC in acute inflammatory responses, we performed a lipopolysaccharide (LPS, Escherichia coli O26:B6) challenge of 16 goats (8 PRNP +/+ and 8 PRNP Ter/Ter) and included 10 saline-treated controls (5 of each PRNP genotype). Clinical examinations were performed continuously, and blood samples were collected throughout the trial. Genome-wide transcription profiles of the choroid plexus, which is at the blood-brain interface, and the hippocampus were analyzed by RNA sequencing, and the same tissues were histologically evaluated. Results All LPS-treated goats displayed clinical signs of sickness behavior, which were of significantly (p < 0.01) longer duration in animals without PrPC. In the choroid plexus, a substantial alteration of the transcriptome and activation of Iba1-positive cells were observed. This response included genotype-dependent differential expression of several genes associated with the immune response, such as ISG15, CXCL12, CXCL14, and acute phase proteins, among others. Activation of cytokine-responsive genes was skewed towards a more profound type I interferon response, and a less obvious type II response, in PrPC-deficient goats. The magnitude of gene expression in response to LPS was smaller in the hippocampus than in the choroid plexus. Resting state expression profiles revealed a few differences between the PRNP genotypes. Conclusions Our data suggest that PrPC acts as a modulator of certain pathways of innate immunity signaling, particularly downstream of interferons, and probably contributes to protection of vulnerable tissues against inflammatory damage

Dynamic conservation and utilization of forest tree genetic resources: indicators for in situ and ex situ genetic conservation and forest reproductive material

This report offers a way to measure the conservation and use of the genetic diversity of forest trees across Europe in a standardized way. This indicator is based on an agreed revision of a commonly adopted framework allowing to measure this diversity and monitor its conservation and utilization over time. The Forest Europe process had adopted a general set of Criteria and Indicators for sustainable forest management as a tool to aid forest policy formulation and decision making, forest monitoring, and communication. Six criteria reflect complementary aspects of sustainable forest management in the pan-European region. These are currently being assessed by a set of 34 quantitative and 11 qualitative indicators. Indicator 4.6 is a quantitative indicator which contributes to Criterion 4 (Maintenance, conservation and appropriate enhancement of biological diversity in forest ecosystems) by focusing on the conservation and use of genetic resources. It is this indicator that has been revised, because it was not very informative on the conservation of genetic diversity nor fully standardised across countries

Loss of prion protein induces a primed state of type I interferon-responsive genes

<div><p>The cellular prion protein (PrP^C) has been extensively studied because of its pivotal role in prion diseases; however, its functions remain incompletely understood. A unique line of goats has been identified that carries a nonsense mutation that abolishes synthesis of PrP^C. In these animals, the PrP-encoding mRNA is rapidly degraded. Goats without PrP^C are valuable in re-addressing loss-of-function phenotypes observed in <i>Prnp</i> knockout mice. As PrP^C has been ascribed various roles in immune cells, we analyzed transcriptomic responses to loss of PrP^C in peripheral blood mononuclear cells (PBMCs) from normal goat kids (<i>n</i> = 8, <i>PRNP</i>^+/+) and goat kids without PrP^C (<i>n</i> = 8, <i>PRNP</i>^Ter/Ter) by mRNA sequencing. PBMCs normally express moderate levels of PrP^C. The vast majority of genes were similarly expressed in the two groups. However, a curated list of 86 differentially expressed genes delineated the two genotypes. About 70% of these were classified as interferon-responsive genes. In goats without PrP^C, the majority of type I interferon-responsive genes were in a primed, modestly upregulated state, with fold changes ranging from 1.4 to 3.7. Among these were <i>ISG15</i>, <i>DDX58</i> (RIG-1), <i>MX1</i>, <i>MX2</i>, <i>OAS1</i>, <i>OAS2 and DRAM1</i>, all of which have important roles in pathogen defense, cell proliferation, apoptosis, immunomodulation and DNA damage response. Our data suggest that PrP^C contributes to the fine-tuning of resting state PBMCs expression level of type I interferon-responsive genes. The molecular mechanism by which this is achieved will be an important topic for further research into PrP^C physiology.</p></div

Expression of interferon-responsive genes in blood leukocytes after <i>in vivo</i> lipopolysaccharide (LPS) challenge in goats without PrP^C.

<p>RNA was extracted from circulating blood leukocytes from both genotypes, and gene expression was analyzed by FLUIDIGM qPCR. (A) Basal expression level (0 h) of selected interferon-responsive genes and <i>STAT1</i> in <i>PRNP</i>^+/+ (<i>n</i> = 12) and <i>PRNP</i>^Ter/Ter (<i>n</i> = 13) animals. (B) Gene expression of interferon-responsive genes and <i>STAT1</i> after <i>in vivo</i> LPS challenge (1 h) from <i>PRNP</i>^+/+ (<i>n</i> = 7) and <i>PRNP</i>^Ter/Ter (<i>n</i> = 8) animals. Values are mean ± SEM. Statistical significance is indicated by *, p-value < 0.05, as assessed by multiple t-tests.</p

Validation of RNA sequencing data with quantitative PCR.

<p>Validation of 12 randomly chosen, differentially expressed genes was performed with qPCR using the original RNA. Expression data from the two methods are presented as relative expression between <i>PRNP</i>^Ter/Ter and <i>PRNP</i>^+/+ animals (RNA-seq data <i>n</i> = 8, qPCR <i>n</i> = 6; r = 0.9616, p < 0.0001, Pearson correlation).</p

PrP^C suppresses upregulation of <i>MX2</i> gene expression upon INF-α stimulation in SH-SY5Y cells.

<p>Untransfected human neuroblastoma SH-SY5Y cells and seven different clones transfected with a plasmid containing human <i>PRNP</i> to produce SH-SY5Y clones expressing human PrP^C, were stimulated for 3h with IFN-α (3 U/ml) (mean ± SEM, <i>n</i> = 4), and <i>MX2</i> gene expression was assessed. Six out of seven clones displayed a significantly lower response to IFN-α compared with the untransfected SH-SY5Y cells, using Dunnett’s post hoc test for multiple comparisons.</p

Mean unique reads of genes related to Interferon signaling from <i>PRNP</i>^+/+ (<i>n</i> = 8, ± SEM) and <i>PRNP</i>^Ter/Ter (<i>n</i> = 8, ± SEM) goats.

<p>Mean unique reads of genes related to Interferon signaling from <i>PRNP</i>^+/+ (<i>n</i> = 8, ± SEM) and <i>PRNP</i>^Ter/Ter (<i>n</i> = 8, ± SEM) goats.</p

Interferon-responsive genes dominate among the differentially expressed genes in goats lacking PrP^C.

<p>Graphical presentation of (A) the total number and percentage of differentially expressed genes (DEGs) between the two genotypes, compared to the average total number of genes expressed in peripheral blood mononuclear cells from both genotypes, and (B) the total number of upregulated and downregulated annotated DEGs. The fraction of upregulated (red) and downregulated (blue) interferon-responsive genes among the DEGs are also shown. (C) Hierarchical clustering of the interferon-responsive genes among the DEGs and expression data from all individual goats of both genotypes. Hierarchical clustering was performed using the ward algorithm on log2-normalized fold changes.</p

The effect of horizontal transmission on the spread of <i>Wolbachia</i> through populations.

<p>Simulations were performed to explore the effect of horizontal transmission on the prevalence of <i>Wolbachia</i>. The blue lines are where there was no horizontal transmission (<i>w</i> = 0), the dashed orange line was a low rate (<i>w</i> = 0.01), and the solid orange line a moderate rate of horizontal transmission (<i>w</i> = 0.06). A: The effect of horizontal transmission when there is not effect of <i>Wolbachia</i> on host fitness and no reproductive manipulation (<i>H</i> = 1, <i>F</i> = 1). B: <i>Wolbachia</i> induces cytoplasmic incompatibility (<i>H</i> = 0.1, <i>F</i> = 1). C: <i>Wolbachia</i> carries a fitness benefit (<i>H</i> = 1, <i>F</i> = 1.05). In all cases the starting prevalence was <i>p</i> = 0.01 and the rate of imperfect maternal transmission was <i>μ</i> = 0.03.</p