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Schlagworte
- induced pluripotent stem cells (3)
- ADHD (2)
- Stammzelle (2)
- cardiomyocytes (2)
- fatty acid (2)
- hiPSC-CM (2)
- stem cell therapy (2)
- tracer (2)
- CRISPR-Cas Systems (1)
- Cadherin-13 (CDH13) (1)
- Cell reprogramming (1)
- Embryonic stem cell (1)
- Induced Pluripotent Stem Cells (1)
- N-Myc (1)
- Nanog protein (1)
- Oct4 (1)
- PARK2 (1)
- Pluripotency (1)
- Protein transduction (1)
- SLC2A3 (1)
- Senescence (1)
- T-cadherin (1)
- blood-brain barrier (BBB) model (1)
- c-Myc (1)
- cadherin-13 (CDH13) (1)
- cadherins (1)
- cell penetrating peptides (1)
- cellular internalization (1)
- disease modelling (1)
- dorsal raphe (1)
- embryonic stem cells (1)
- female (1)
- gene expression (1)
- hematopoietic stem cells (1)
- heterozygote (1)
- hiPSC (1)
- homeodomain transcription factors (1)
- human induced pluripotent stem cell (hiPSC) (1)
- human induced pluripotent stem cells (hiPSCs)human induced pluripotent stem cells (hiPSCs) (1)
- humans (1)
- leukemia inhibitory factor (1)
- median and dorsal raphe (1)
- middle aged (1)
- mitochondria (1)
- multipotent fetal neural stem cells (fNSCs) (1)
- mutations (1)
- neurodevelopment (1)
- neurodevelopmental disorders / genetics (1)
- neuropsychiatric disorders (1)
- neurovascular unit in vitro (1)
- p27(KIP1) (1)
- penetratin (1)
- prefrontal cortex (1)
- protein synthesis (1)
- psychiatric disorders (1)
- radial glia (1)
- reprogramming (1)
- self-renewal (1)
- serotonin (1)
- serotonin-specific neurons (1)
- synapse formation (1)
Institut
- Institut für Anatomie und Zellbiologie (11)
- Institut für Humangenetik (3)
- Klinik und Poliklinik für Psychiatrie, Psychosomatik und Psychotherapie (3)
- Deutsches Zentrum für Herzinsuffizienz (DZHI) (2)
- Klinik und Poliklinik für Nuklearmedizin (2)
- Lehrstuhl für Molekulare Psychiatrie (2)
- Physiologisches Institut (2)
- Theodor-Boveri-Institut für Biowissenschaften (2)
- Frauenklinik und Poliklinik (1)
- Institut für Klinische Neurobiologie (1)
Sonstige beteiligte Institutionen
- Department of Biomedical Imaging, National Cerebral and Cardiovascular Research Center, Suita, Japan (2)
- Division of Medical Technology and Science, Department of Medical Physics and Engineering, Course of Health Science, Osaka University Graduate School of Medicine, Suita Japan (2)
- Institut for Molecular Biology and CMBI, Department of Genomics, Stem Cell Biology and Regenerative Medicine, Leopold-Franzens-University Innsbruck, Innsbruck, Austria (2)
- Johns Hopkins School of Medicine, The Russell H Morgan Department of Radiology and Radiological Science, Baltimore, MD, USA (2)
Fibroblasts were isolated from a skin biopsy of a clinically diagnosed 51-year-old female attention-deficit/hyperactivity disorder (ADHD) patient carrying a duplication of SLC2A3, a gene encoding neuronal glucose transporter-3 (GLUT3). Patient fibroblasts were infected with Sendai virus, a single-stranded RNA virus, to generate transgene-free human induced pluripotent stem cells (iPSCs). SLC2A3-D2-iPSCs showed expression of pluripotency-associated markers, were able to differentiate into cells of the three germ layers in vitro and had a normal female karyotype. This in vitro cellular model can be used to study the role of risk genes in the pathogenesis of ADHD, in a patient-specific manner.
BACKGROUND:
Oct4 is a transcription factor that plays a major role for the preservation of the pluripotent state in embryonic stem cells as well as for efficient reprogramming of somatic cells to induced pluripotent stem cells (iPSC) or other progenitors. Protein-based reprogramming methods mainly rely on the addition of a fused cell penetrating peptide. This study describes that Oct4 inherently carries a protein transduction domain, which can translocate into human and mouse cells.
RESULTS:
A 16 amino acid peptide representing the third helix of the human Oct4 homeodomain, referred to as Oct4 protein transduction domain (Oct4-PTD), can internalize in mammalian cells upon conjugation to a fluorescence moiety thereby acting as a cell penetrating peptide (CPP). The cellular distribution of Oct4-PTD shows diffuse cytosolic and nuclear staining, whereas penetratin is strictly localized to a punctuate pattern in the cytoplasm. By using a Cre/loxP-based reporter system, we show that this peptide also drives translocation of a functionally active Oct4-PTD-Cre-fusion protein. We further provide evidence for translocation of full length Oct4 into human and mouse cell lines without the addition of any kind of cationic fusion tag. Finally, physico-chemical properties of the novel CPP are characterized, showing that in contrast to penetratin a helical structure of Oct4-PTD is only observed if the FITC label is present on the N-terminus of the peptide.
CONCLUSIONS:
Oct4 is a key transcription factor in stem cell research and cellular reprogramming. Since it has been shown that recombinant Oct4 fused to a cationic fusion tag can drive generation of iPSCs, our finding might contribute to further development of protein-based methods to generate iPSCs. Moreover, our data support the idea that transcription factors might be part of an alternative paracrine signalling pathway, where the proteins are transferred to neighbouring cells thereby actively changing the behaviour of the recipient cell.
Background: During early prenatal stages of brain development, serotonin (5-HT)-specific neurons migrate through somal translocation to form the raphe nuclei and subsequently begin to project to their target regions. The rostral cluster of cells, comprising the median and dorsal raphe (DR), innervates anterior regions of the brain, including the prefrontal cortex. Differential analysis of the mouse 5-HT system transcriptome identified enrichment of cell adhesion molecules in 5-HT neurons of the DR. One of these molecules, cadherin-13 (Cdh13) has been shown to play a role in cell migration, axon pathfinding, and synaptogenesis. This study aimed to investigate the contribution of Cdh13 to the development of the murine brain 5-HT system.
Methods: For detection of Cdh13 and components of the 5-HT system at different embryonic developmental stages of the mouse brain, we employed immunofluorescence protocols and imaging techniques, including epifluorescence, confocal and structured illumination microscopy. The consequence of CDH13 loss-of-function mutations on brain 5-HT system development was explored in a mouse model of Cdh13 deficiency.
Results: Our data show that in murine embryonic brain Cdh13 is strongly expressed on 5-HT specific neurons of the DR and in radial glial cells (RGCs), which are critically involved in regulation of neuronal migration. We observed that 5-HT neurons are intertwined with these RGCs, suggesting that these neurons undergo RGC-guided migration. Cdh13 is present at points of intersection between these two cell types. Compared to wildtype controls, Cdh13-deficient mice display increased cell densities in the DR at embryonic stages E13.5, E17.5, and adulthood, and higher serotonergic innervation of the prefrontal cortex at E17.5.
Conclusion: Our findings provide evidence for a role of CDH13 in the development of the serotonergic system in early embryonic stages. Specifically, we indicate that Cdh13 deficiency affects the cell density of the developing DR and the posterior innervation of the prefrontal cortex (PFC), and therefore might be involved in the migration, axonal outgrowth and terminal target finding of DR 5-HT neurons. Dysregulation of CDH13 expression may thus contribute to alterations in this system of neurotransmission, impacting cognitive function, which is frequently impaired in neurodevelopmental disorders including attention-deficit/hyperactivity and autism spectrum disorders.
In vitro models of the human blood-brain barrier (BBB) are highly desirable for drug development. This study aims to analyze a set of ten different BBB culture models based on primary cells, human induced pluripotent stem cells (hiPSCs), and multipotent fetal neural stem cells (fNSCs). We systematically investigated the impact of astrocytes, pericytes, and NSCs on hiPSC-derived BBB endothelial cell function and gene expression. The quadruple culture models, based on these four cell types, achieved BBB characteristics including transendothelial electrical resistance (TEER) up to 2,500 Ω cm\(^{2}\) and distinct upregulation of typical BBB genes. A complex in vivo-like tight junction (TJ) network was detected by freeze-fracture and transmission electron microscopy. Treatment with claudin-specific TJ modulators caused TEER decrease, confirming the relevant role of claudin subtypes for paracellular tightness. Drug permeability tests with reference substances were performed and confirmed the suitability of the models for drug transport studies.