PapRIV, a BV-2 microglial cell activating quorum sensing peptide

Quorum sensing peptides (QSPs) are bacterial peptides produced by Gram-positive bacteria to communicate with their peers in a cell-density dependent manner. These peptides do not only act as interbacterial communication signals, but can also have effects on the host. Compelling evidence demonstrates the presence of a gut-brain axis and more specifically, the role of the gut microbiota in microglial functioning. The aim of this study is to investigate microglial activating properties of a selected QSP (PapRIV) which is produced by Bacillus cereus species. PapRIV showed in vitro activating properties of BV-2 microglia cells and was able to cross the in vitro Caco-2 cell model and reach the brain. In vivo peptide presence was also demonstrated in mouse plasma. The peptide caused induction of IL-6, TNFα and ROS expression and increased the fraction of ameboid BV-2 microglia cells in an NF-κB dependent manner. Different metabolites were identified in serum, of which the main metabolite still remained active. PapRIV is thus able to cross the gastro-intestinal tract and the blood–brain barrier and shows in vitro activating properties in BV-2 microglia cells, hereby indicating a potential role of this quorum sensing peptide in gut-brain interaction

Absence of microglia promotes diverse pathologies and early lethality in Alzheimer’s disease mice

Microglia are strongly implicated in the development and progression of Alzheimer's disease (AD), yet their impact on pathology and lifespan remains unclear. Here we utilize a CSF1R hypomorphic mouse to generate a model of AD that genetically lacks microglia. The resulting microglial-deficient mice exhibit a profound shift from parenchymal amyloid plaques to cerebral amyloid angiopathy (CAA), which is accompanied by numerous transcriptional changes, greatly increased brain calcification and hemorrhages, and premature lethality. Remarkably, a single injection of wild-type microglia into adult mice repopulates the microglial niche and prevents each of these pathological changes. Taken together, these results indicate the protective functions of microglia in reducing CAA, blood-brain barrier dysfunction, and brain calcification. To further understand the clinical implications of these findings, human AD tissue and iPSC-microglia were examined, providing evidence that microglia phagocytose calcium crystals, and this process is impaired by loss of the AD risk gene, TREM2

Tissue Effect on Genetic Control of Transcript Isoform Variation

Current genome-wide association studies (GWAS) are moving towards the use of large cohorts of primary cell lines to study a disease of interest and to assign biological relevance to the genetic signals identified. Here, we use a panel of human osteoblasts (HObs) to carry out a transcriptomic survey, similar to recent studies in lymphoblastoid cell lines (LCLs). The distinct nature of HObs and LCLs is reflected by the preferential grouping of cell type–specific genes within biologically and functionally relevant pathways unique to each tissue type. We performed cis-association analysis with SNP genotypes to identify genetic variations of transcript isoforms, and our analysis indicates that differential expression of transcript isoforms in HObs is also partly controlled by cis-regulatory genetic variants. These isoforms are regulated by genetic variants in both a tissue-specific and tissue-independent fashion, and these associations have been confirmed by RT–PCR validation. Our study suggests that multiple transcript isoforms are often present in both tissues and that genetic control may affect the relative expression of one isoform to another, rather than having an all-or-none effect. Examination of the top SNPs from a GWAS of bone mineral density show overlap with probeset associations observed in this study. The top hit corresponding to the FAM118A gene was tested for association studies in two additional clinical studies, revealing a novel transcript isoform variant. Our approach to examining transcriptome variation in multiple tissue types is useful for detecting the proportion of genetic variation common to different cell types and for the identification of cell-specific isoform variants that may be functionally relevant, an important follow-up step for GWAS

Next-generation sequencing-based genome diagnostics across clinical genetics centers: Implementation choices and their effects

Implementation of next-generation DNA sequencing (NGS) technology into routine diagnostic genome care requires strategic choices. Instead of theoretical discussions on the consequences of such choices, we compared NGS-based diagnostic practices in eight clinical genetic centers in the Netherlands, based on genetic testing of nine pre-selected patients with cardiomyopathy. We highlight critical implementation choices, including the specific contributions of laboratory and medical specialists, bioinformaticians and researchers to diagnostic genome care, and how these affect interpretation and reporting of variants. Reported pathogenic mutations were consistent for all but one patient. Of the two centers that were inconsistent in their diagnosis, one reported to have found 'no causal variant', thereby underdiagnosing this patient. The other provided an alternative diagnosis, identifying another variant as causal than the other centers. Ethical and legal analysis showed that informed consent procedures in all centers were generally adequate for diagnostic NGS applications that target a limited set of genes, but not for exome- and genome-based diagnosis. We propose changes to further improve and align these procedures, taking into account the blurring boundary between diagnostics and research, and specific counseling options for exome- and genome-based diagnostics. We conclude that alternative diagnoses may infer a certain level of 'greediness' to come to a positive diagnosis in interpreting sequencing results. Moreover, there is an increasing interdependence of clinic, diagnostics and research departments for comprehensive diagnostic genome care. Therefore, we invite clinical geneticists, physicians, researchers, bioinformatics experts and patients to reconsider their role and position in future diagnostic genome care

Hair Cortisol in Twins : Heritability and Genetic Overlap with Psychological Variables and Stress-System Genes

A. Palotie on työryhmän jäsen.Hair cortisol concentration (HCC) is a promising measure of long-term hypothalamus-pituitary-adrenal (HPA) axis activity. Previous research has suggested an association between HCC and psychological variables, and initial studies of inter-individual variance in HCC have implicated genetic factors. However, whether HCC and psychological variables share genetic risk factors remains unclear. The aims of the present twin study were to: (i) assess the heritability of HCC; (ii) estimate the phenotypic and genetic correlation between HPA axis activity and the psychological variables perceived stress, depressive symptoms, and neuroticism; using formal genetic twin models and molecular genetic methods, i.e. polygenic risk scores (PRS). HCC was measured in 671 adolescents and young adults. These included 115 monozygotic and 183 dizygotic twin-pairs. For 432 subjects PRS scores for plasma cortisol, major depression, and neuroticism were calculated using data from large genome wide association studies. The twin model revealed a heritability for HCC of 72%. No significant phenotypic or genetic correlation was found between HCC and the three psychological variables of interest. PRS did not explain variance in HCC. The present data suggest that HCC is highly heritable. However, the data do not support a strong biological link between HCC and any of the investigated psychological variables.Peer reviewe

Achieving information-seeking skills: exploring information-seeking emotions through an appraisal theory framework

Introduction. The paper explores the proposed appraisal theory framework in a study of primary teacher students´ emotions in the process of learning and achieving information-seeking skills. Studying this process is important for teacher students’ academic success and future research-informed practice. The relevance of applying the framework, Scherer´s (2005) semantic space of emotions and Pekrun´s (2006) control-value theory of achievement emotions, is presented, elucidated and discussed. Method. Concepts and elements of the semantic space of emotions and control-value theory of achievement emotions are presented. Analysis and results. Viewed together, Scherer´s (2005) semantic space of emotions and Pekrun´s (2006) control-value theory of achievement emotions constitute an appropriate analytical tool to investigate teacher students´ emotions in learning and achieving information-seeking skills. It offers an understanding and explanation of the nature of information-seeking emotions and their interplay with cognitive appraisals. Conclusion. Given the limited body of literature on affective behaviours in library and information science research in general and in relation to appraisal theories in an educational context in particular, it is hoped that the paper provides a valuable conceptual and theoretical contribution to the field

Generating tissue-resident macrophages from pluripotent stem cells: Lessons learned from microglia

Over the past decades, the importance of the immune system in a broad scope of pathologies, has drawn attention towards tissue-resident macrophages, such as microglia in the brain. To enable the study of for instance microglia, it is crucial to recreate in vitro (and in vivo) assays. However, very fast loss of tissue-specific features of primary tissue resident macrophages, including microglia, upon in vitro culture has complicated such studies. Moreover, limited knowledge of macrophage developmental pathways and the role of local 'niche factors', has hampered the generation of tissue-resident macrophages from pluripotent stem cells (PSC). Recent data on the ontogeny of tissue-resident macrophages, combined with bulk and single cell RNAseq studies have identified the distinct origins and gene profile of microglia compared to other myeloid cells. As a result, over the past years, protocols have been published to create hPSC-derived microglia-'like' cells, as these cells are considered potential new therapeutic targets for therapies to treat neurodegenerative diseases. In this review we will provide an overview of different approaches taken to generate human microglia in vitro, taking into account their origin, and resemblance to their in vivo counterpart. Finally, we will discuss cell-extrinsic (culture conditions) and intrinsic factors (transcriptional machinery and epigenetics) that we believe can improve future differentiation protocols of tissue-resident macrophages from stem cells.status: publishe

Generation of Self-Assembling 3D Neural Micro-Organoids using Human iPSC-Derived Neuronal Progenitors and Microglia

INTRODUCTION: Recent studies showed that microglia (MG) play critical roles in neurodegeneration, and mutations of specific genes in MG increase the risk of neurodegenerative diseases, including Alzheimer’s disease (AD). Although murine models have started to shed light on the role of MG in AD, the lack of human models hampers the study of MG-related neurodegeneration. With the advent of induced pluripotent stem cell (iPSC) technology, and knowing that in-vitro 3D co-culture better approximates the complex in-vivo cellular organization than 2D monolayer co-culture, we set out to generate self-assembling 3D neural micro-organoids of human origin as potential in-vitro 3D cellular models for unravelling neuronal-MG crosstalk relevant to neurodegeneration. MATERIALS AND METHODS: To facilitate lineage tracing, human iPSCs were genetically engineered to constitutively express either TdTomato or GFP fluorescent protein from the safe harbor AAVS1 locus. Using in-house protocols, the cells were differentiated into human neuronal progenitor cells (hNPCs-TdTomato) (Garcia Leon, et al., Alzheimer’s & Dementia, 2018) or human microglia-like cells (hMGs-GFP) (Claes et al., Alzheimer’s & Dementia, in press) respectively. At a hNPCs:hMGs ratio of 2:1, the cells were mixed and seeded onto agarose substrates imprinted with 121 microwells (Ф500 µm x H500 µm). The cells were cultured in neural maintenance medium to allow self-assembling of neural micro-organoids. Micro-organoid formation was monitored by brightfield and fluorescent microscopy. Differentiation and maturation of hNPCs and hMGs was analyzed by qRT-PCR. As an alternative approach, cerebral brain organoids were generated via a well-accepted published protocol, followed by seeding of hMGs onto the organoids. RESULTS AND DISCUSSION: hiPSC-derived hNPC:hMG mixtures started to self-assemble into micro-organoids after 9 days of culture (Fig. 1a). The micro-organoids consisted of hNPCs-dTomato (red) and hMGs-GFP cells, and discrete hNPCs and hMGs submicro-organoids with distinctive spheroid morphologies (Fig. 1b). Alternatively, hiPSCs were differentiated towards cerebral brain organoids, and the neuronal marker, Nestin, was detected after 21 days (Fig. 2a & 2f). Interestingly, after seeding hMGs (Fig. 2b) on the organoids, clear morphological changes were observed in the co-culture (Fig. 2c–2e). CONCLUSION: By harnessing the self-assembling property of hiPSC-derived hNPCs and hMGs, and assisted by robust differentiation protocols, neural micro-organoids or cerebral brain organoids were generated. These organoids represent unprecedented in-vitro 3D models essential for unravelling neuron-microglia crosstalk linked to neurodegeneration. ACKNOWLEDGEMENT: This research project is supported by the IWT_SBO-iPSCAF (IWT 150031 iPSC), KU Leuven C1-3DMuSyC grant (C14/17/111), and IWT Aspirant grant (SC/1S1071N).status: publishe