Neuropsychological effects of the CSMD1 genome‐wide associated schizophrenia risk variant rs10503253

The single‐nucleotide polymorphism (SNP) rs10503253, located within the CUB and Sushi multiple domains‐1 (CSMD1) gene on 8p23.2, was recently identified as genome‐wide significant for schizophrenia (SZ), but is of unknown function. We investigated the neurocognitive effects of this CSMD1 variant in vivo in patients and healthy participants using behavioral and imaging measures of brain structure and function. We compared carriers and non‐carriers of the risk ‘A’ allele on measures of neuropsychological performance typically impaired in SZ (general cognitive ability, episodic and working memory and attentional control) in independent samples of Irish patients (n = 387) and controls (n = 171) and German patients (205) and controls (n = 533). Across these groups, the risk ‘A’ allele at CSMD1 was associated with deleterious effects across a number of neurocognitive phenotypes. Specifically, the risk allele was associated with poorer performance on neuropsychological measures of general cognitive ability and memory function but not attentional control. These effects, while significant, were subtle, and varied between samples. Consistent with previous evidence suggesting that CSMD1 may be involved in brain mechanisms related to memory and learning, these data appear to reflect the deleterious effects of the identified ‘A’ risk allele on neurocognitive function, possibly as part of the mechanism by which CSMD1 is associated with SZ risk.     

Characterization of Cytomegalovirus Disease in Solid Organ Transplant Recipients by Markers of Inflammation in Plasma

Background

While several studies have examined the general inflammatory responses in relation to cytomegalovirus infection, the identification of the various inflammatory mediators as well as their relative importance is far from clear.

Patients and Methods

Solid organ recipients enrolled in an international multicenter trial of cytomegalovirus disease treatment (the VICTOR study) were analyzed (n = 289) (ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00431353","term_id":"NCT00431353"}}NCT00431353). Plasma markers of inflammation and endothelial cell activation were assessed at baseline by enzyme immunoassays.

Results

The major findings were: (i) Plasma levels of the CXC-chemokine interferon-inducible protein-10 (P<0.001) and C-reactive protein (P = 0.046) were independently associated with the presence of cytomegalovirus DNAemia above lower level of quantification. (ii) High levels of CC-chemokine ligand 21 (P = 0.027) and pentraxin 3 (P = 0.033) were independently associated with tissue invasive cytomegalovirus disease as opposed to cytomegalovirus syndrome.

Conclusion

Our findings illustrate the complex interaction between cytomegalovirus and the immune system, involving a wide range of inflammatory mediators that could be associated to disease manifestations in cytomegalovirus related disease.     

A Human In Vitro Whole Blood Assay to Predict the Systemic Cytokine Response to Therapeutic Oligonucleotides Including siRNA

Therapeutic oligonucleotides including siRNA and immunostimulatory ligands of Toll-like receptors (TLR) or RIG-I like helicases (RLH) are a promising novel class of drugs. They are in clinical development for a broad spectrum of applications, e.g. as adjuvants in vaccines and for the immunotherapy of cancer. Species-specific immune activation leading to cytokine release is characteristic for therapeutic oligonucleotides either as an unwanted side effect or intended pharmacology. Reliable in vitro tests designed for therapeutic oligonucleotides are therefore urgently needed in order to predict clinical efficacy and to prevent unexpected harmful effects in clinical development. To serve this purpose, we here established a human whole blood assay (WBA) that is fast and easy to perform. Its response to synthetic TLR ligands (R848: TLR7/8, LPS: TLR4) was on a comparable threshold to the more time consuming peripheral blood mononuclear cell (PBMC) based assay. By contrast, the type I IFN profile provoked by intravenous CpG-DNA (TLR9 ligand) in humans in vivo was more precisely replicated in the WBA than in stimulated PBMC. Since Heparin and EDTA, but not Hirudin, displaced oligonucleotides from their delivery agent, only Hirudin qualified as the anticoagulant to be used in the WBA. The Hirudin WBA exhibited a similar capacity as the PBMC assay to distinguish between TLR7-activating and modified non-stimulatory siRNA sequences. RNA-based immunoactivating TLR7/8- and RIG-I-ligands induced substantial amounts of IFN-α in the Hirudin-WBA dependent on delivery agent used. In conclusion, we present a human Hirudin WBA to determine therapeutic oligonucleotide-induced cytokine release during preclinical development that can readily be performed and offers a close reflection of human cytokine response in vivo.     

Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses

The IFNL4 gene is a recently discovered type III interferon, which in a significant fraction of the human population harbours a frameshift mutation abolishing the IFNλ4 ORF. The expression of IFNλ4 is correlated with both poor spontaneous clearance of hepatitis C virus (HCV) and poor response to treatment with type I interferon. Here, we show that the IFNL4 gene encodes an active type III interferon, named IFNλ4, which signals through the IFNλR1 and IL‐10R2 receptor chains. Recombinant IFNλ4 is antiviral against both HCV and coronaviruses at levels comparable to IFNλ3. However, the secretion of IFNλ4 is impaired compared to that of IFNλ3, and this impairment is not due to a weak signal peptide, which was previously believed. We found that IFNλ4 gets N‐linked glycosylated and that this glycosylation is required for secretion. Nevertheless, this glycosylation is not required for activity. Together, these findings result in the paradox that IFNλ4 is strongly antiviral but a disadvantage during HCV infection.     

Development of an analytical approach for identification and quantification of 5-α-androst-16-en-3-one in human milk

A method was developed for the quantification of 5-α-androst-16-en-3-one in human breast milk based on application of a stable isotope dilution assay using 5α-androst-16-en-3-one-6, 6-d₂. The procedure includes extraction of the human milk by hexane with subsequent clean-up of the obtained extract by gel permeation and silica gel column chromatography. The extracted samples were analyzed by gas chromatography–mass spectrometry. Using this method 5-α-androst-16-en-3-one could be identified and for the first time quantified in a concentration range of 26–155 ng/kg in human milk.     

Feeding the Probiotic Enterococcus faecium Strain NCIMB 10415 to Piglets Specifically Reduces the Number of Escherichia coli Pathotypes That Adhere to the Gut Mucosa

Feed supplementation with the probiotic Enterococcus faecium for piglets has been found to reduce pathogenic gut microorganisms. Since Escherichia coli is among the most important pathogens in pig production, we performed comprehensive analyses to gain further insight into the influence of E. faecium NCIMB 10415 on porcine intestinal E. coli. A total of 1,436 E. coli strains were isolated from three intestinal habitats (mucosa, digesta, and feces) of probiotic-supplemented and nonsupplemented (control) piglets. E. coli bacteria were characterized via pulsed-field gel electrophoresis (PFGE) for clonal analysis. The high diversity of E. coli was reflected by 168 clones. Multilocus sequence typing (MLST) was used to determine the phylogenetic backgrounds, revealing 79 sequence types (STs). Pathotypes of E. coli were further defined using multiplex PCR for virulence-associated genes. While these analyses discerned only a few significant differences in the E. coli population between the feeding groups, analyses distinguishing clones that were uniquely isolated in either the probiotic group only, the control group only, or both groups (shared group) revealed clear effects at the habitat level. Interestingly, extraintestinal pathogenic E. coli (ExPEC)-typical clones adhering to the mucosa were significantly reduced in the probiotic group. Our data show a minor influence of E. faecium on the overall population of E. coli in healthy piglets. In contrast, this probiotic has a profound effect on mucosa-adherent E. coli. This finding further substantiates a specific effect of E. faecium strain NCIMB 10415 in piglets against pathogenic E. coli in the intestine. In addition, these data question the relevance of data based on sampling fecal E. coli only.     

TRAP assists membrane protein topogenesis at the mammalian ER membrane

Membrane protein insertion and topogenesis generally occur at the Sec61 translocon in the endoplasmic reticulum membrane. During this process, membrane spanning segments may adopt two distinct orientations with either their N- or C-terminus translocated into the ER lumen. While different topogenic determinants in membrane proteins, such as flanking charges, polypeptide folding, and hydrophobicity, have been identified, it is not well understood how the translocon and/or associated components decode them. Here we present evidence that the translocon-associated protein (TRAP) complex is involved in membrane protein topogenesis in vivo. Small interfering RNA (siRNA)-mediated silencing of the TRAP complex in HeLa cells enhanced the topology effect of mutating the flanking charges of a signal-anchor, but not of increasing signal hydrophobicity. The results suggest a role of the TRAP complex in moderating the ‘positive-inside’ rule.     

High levels of Cellular Prion Protein improve astrocyte development

Prion protein (PrP^C) has neuroprotective functions and herein we demonstrate that astrocytes from PrP^C-over-expressing mice are more resistant to induced cell death than wild-type astrocytes. The Stress-Inducible-Protein 1 (STI1), a PrP^C ligand, prevents cell death in both wild-type and PrP^C-over-expressing astrocytes through the activation of protein-kinase-A. Cultured embryonic astrocytes and brain extracts from PrP^C-over-expressing mice show higher glial fibrillary acidic protein expression and reduced vimentin and nestin levels when compared to wild-type astrocytes, suggesting faster astrocyte maturation in the former mice. Our data indicate that PrP^C levels modulate astrocyte development, and that PrP^C–STI1 interaction contributes to protect against astrocyte death.