Publications by authors named "Frank Edenhofer"

73 Publications

Functionally distinct POMC-expressing neuron subpopulations in hypothalamus revealed by intersectional targeting.

Nat Neurosci 2021 May 17. Epub 2021 May 17.

Max Planck Institute for Metabolism Research, Department of Neuronal Control of Metabolism, Cologne, Germany.

Pro-opiomelanocortin (POMC)-expressing neurons in the arcuate nucleus of the hypothalamus represent key regulators of metabolic homeostasis. Electrophysiological and single-cell sequencing experiments have revealed a remarkable degree of heterogeneity of these neurons. However, the exact molecular basis and functional consequences of this heterogeneity have not yet been addressed. Here, we have developed new mouse models in which intersectional Cre/Dre-dependent recombination allowed for successful labeling, translational profiling and functional characterization of distinct POMC neurons expressing the leptin receptor (Lepr) and glucagon like peptide 1 receptor (Glp1r). Our experiments reveal that POMC and POMC neurons represent largely nonoverlapping subpopulations with distinct basic electrophysiological properties. They exhibit a specific anatomical distribution within the arcuate nucleus and differentially express receptors for energy-state communicating hormones and neurotransmitters. Finally, we identify a differential ability of these subpopulations to suppress feeding. Collectively, we reveal a notably distinct functional microarchitecture of critical metabolism-regulatory neurons.
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http://dx.doi.org/10.1038/s41593-021-00854-0DOI Listing
May 2021

Age-dependent instability of mature neuronal fate in induced neurons from Alzheimer's patients.

Cell Stem Cell 2021 Apr 21. Epub 2021 Apr 21.

Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, CA, USA. Electronic address:

Sporadic Alzheimer's disease (AD) exclusively affects elderly people. Using direct conversion of AD patient fibroblasts into induced neurons (iNs), we generated an age-equivalent neuronal model. AD patient-derived iNs exhibit strong neuronal transcriptome signatures characterized by downregulation of mature neuronal properties and upregulation of immature and progenitor-like signaling pathways. Mapping iNs to longitudinal neuronal differentiation trajectory data demonstrated that AD iNs reflect a hypo-mature neuronal identity characterized by markers of stress, cell cycle, and de-differentiation. Epigenetic landscape profiling revealed an underlying aberrant neuronal state that shares similarities with malignant transformation and age-dependent epigenetic erosion. To probe for the involvement of aging, we generated rejuvenated iPSC-derived neurons that showed no significant disease-related transcriptome signatures, a feature that is consistent with epigenetic clock and brain ontogenesis mapping, which indicate that fibroblast-derived iNs more closely reflect old adult brain stages. Our findings identify AD-related neuronal changes as age-dependent cellular programs that impair neuronal identity.
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http://dx.doi.org/10.1016/j.stem.2021.04.004DOI Listing
April 2021

CRISPR/Cas9-edited PKP2 knock-out (JMUi001-A-2) and DSG2 knock-out (JMUi001-A-3) iPSC lines as an isogenic human model system for arrhythmogenic cardiomyopathy (ACM).

Stem Cell Res 2021 May 18;53:102256. Epub 2021 Feb 18.

Comprehensive Heart Failure Center (CHFC), Department of Cardiovascular Genetics, University Clinics Würzburg, Würzburg, Germany; Department of Medicine I, University Clinics Würzburg, Würzburg, Germany. Electronic address:

Arrhythmogenic cardiomyopathy (ACM) is characterized by fibro-fatty replacement of the myocardium, heart failure and life-threatening ventricular arrhythmias. Causal mutations were identified in genes encoding for proteins of the desmosomes, predominantly plakophilin-2 (PKP2) and desmoglein-2 (DSG2). We generated gene-edited knock-out iPSC lines for PKP2 (JMUi001-A-2) and DSG2 (JMUi001-A-3) using the CRISPR/Cas9 system in a healthy control iPSC background (JMUi001-A). Stem cell-like morphology, robust expression of pluripotency markers, embryoid body formation and normal karyotypes confirmed the generation of high quality iPSCs to provide a novel isogenic human in vitro model system mimicking ACM when differentiated into cardiomyocytes.
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http://dx.doi.org/10.1016/j.scr.2021.102256DOI Listing
May 2021

Serotonin-specific neurons differentiated from human iPSCs form distinct subtypes with synaptic protein assembly.

J Neural Transm (Vienna) 2021 02 9;128(2):225-241. Epub 2021 Feb 9.

Division of Molecular Psychiatry, Center of Mental Health, University of Würzburg, Margarete-Höppel-Platz 1, 97080, Würzburg, Germany.

Human induced pluripotent stem cells (hiPSCs) have revolutionized the generation of experimental disease models, but the development of protocols for the differentiation of functionally active neuronal subtypes with defined specification is still in its infancy. While dysfunction of the brain serotonin (5-HT) system has been implicated in the etiology of various neuropsychiatric disorders, investigation of functional human 5-HT specific neurons in vitro has been restricted by technical limitations. We describe an efficient generation of functionally active neurons from hiPSCs displaying 5-HT specification by modification of a previously reported protocol. Furthermore, 5-HT specific neurons were characterized using high-end fluorescence imaging including super-resolution microscopy in combination with electrophysiological techniques. Differentiated hiPSCs synthesize 5-HT, express specific markers, such as tryptophan hydroxylase 2 and 5-HT transporter, and exhibit an electrophysiological signature characteristic of serotonergic neurons, with spontaneous rhythmic activities, broad action potentials and large afterhyperpolarization potentials. 5-HT specific neurons form synapses reflected by the expression of pre- and postsynaptic proteins, such as Bassoon and Homer. The distribution pattern of Bassoon, a marker of the active zone along the soma and extensions of neurons, indicates functionality via volume transmission. Among the high percentage of 5-HT specific neurons (~ 42%), a subpopulation of CDH13 + cells presumably designates dorsal raphe neurons. hiPSC-derived 5-HT specific neuronal cell cultures reflect the heterogeneous nature of dorsal and median raphe nuclei and may facilitate examining the association of serotonergic neuron subpopulations with neuropsychiatric disorders.
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http://dx.doi.org/10.1007/s00702-021-02303-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7914246PMC
February 2021

Generation of induced pluripotent stem cell (iPSC) lines carrying a heterozygous (UKWMPi002-A-1) and null mutant knockout (UKWMPi002-A-2) of Cadherin 13 associated with neurodevelopmental disorders using CRISPR/Cas9.

Stem Cell Res 2021 03 11;51:102169. Epub 2021 Jan 11.

Division of Molecular Psychiatry, Center of Mental Health, University Hospital Würzburg, Germany; Laboratory of Psychiatric Neurobiology, Institute of Molecular Medicine, I.M Sechenov First Moscow State Medical University, Moscow, Russia; Department of Psychiatry and Neuropsychology, School for Mental Health and Neuroscience (MHeNs), Maastricht University, Maastricht, The Netherlands. Electronic address:

Fibroblasts isolated from a skin biopsy of a healthy 46-year-old female were infected with Sendai virus containing the Yamanaka factors to produce transgene-free human induced pluripotent stem cells (iPSCs). CRISPR/Cas9 was used to generate isogenic cell lines with a gene dose-dependent deficiency of CDH13, a risk gene associated with neurodevelopmental and psychiatric disorders. Thereby, a heterozygous CDH13 knockout (CDH13) and a CDH13 null mutant (CDH13) iPSC line was obtained. All three lines showed expression of pluripotency-associated markers, the ability to differentiate into cells of the three germ layers in vitro, and a normal female karyotype.
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http://dx.doi.org/10.1016/j.scr.2021.102169DOI Listing
March 2021

Energy Metabolism Disturbances in Cell Models of PARK2 CNV Carriers with ADHD.

J Clin Med 2020 Dec 18;9(12). Epub 2020 Dec 18.

Department of Psychiatry, Psychotherapy and Psychosomatic Medicine, University Hospital, Goethe University, D-60528 Frankfurt, Germany.

The main goal of the present study was the identification of cellular phenotypes in attention-deficit-/hyperactivity disorder (ADHD) patient-derived cellular models from carriers of rare copy number variants (CNVs) in the locus that have been previously associated with ADHD. Human-derived fibroblasts (HDF) were cultured and human-induced pluripotent stem cells (hiPSC) were reprogrammed and differentiated into dopaminergic neuronal cells (mDANs). A series of assays in baseline condition and in different stress paradigms (nutrient deprivation, carbonyl cyanide m-chlorophenyl hydrazine (CCCP)) focusing on mitochondrial function and energy metabolism (ATP production, basal oxygen consumption rates, reactive oxygen species (ROS) abundance) were performed and changes in mitochondrial network morphology evaluated. We found changes in CNV deletion and duplication carriers with ADHD in PARK2 gene and protein expression, ATP production and basal oxygen consumption rates compared to healthy and ADHD wildtype control cell lines, partly differing between HDF and mDANs and to some extent enhanced in stress paradigms. The generation of ROS was not influenced by the genotype. Our preliminary work suggests an energy impairment in HDF and mDAN cells of CNV deletion and duplication carriers with ADHD. The energy impairment could be associated with the role of dysregulation in mitochondrial dynamics.
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http://dx.doi.org/10.3390/jcm9124092DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7766864PMC
December 2020

Development of a bioreactor system for pre-endothelialized cardiac patch generation with enhanced viscoelastic properties by combined collagen I compression and stromal cell culture.

J Tissue Eng Regen Med 2020 12 18;14(12):1749-1762. Epub 2020 Sep 18.

Chair of Tissue Engineering and Regenerative Medicine, Wuerzburg University Hospital, Wuerzburg, Germany.

Treatment of terminal heart failure still poses a significant clinical problem. Cardiac tissue engineering could offer autologous solutions for the replacement of nonfunctional myocardial tissue. So far, soft matrix construction and missing large-scale prevascularization prevented the application of sizeable cardiac repair patches. We developed a novel bioreactor system for semi-automatic compression of a collagen I hydrogel applying 16 times higher pressure than in previous studies. Resistance towards compression stress was investigated for multiple cardiac-related cell types. For scaffold prevascuarization, a tubular cavity was imprinted during the compaction process. Primary cardiac-derived endothelial cells (ECs) were isolated from human left atrial appendages (HLAAs) and characterized by fluorescence-activated cell sorting (FACS) and immunocytology. EC were then seeded into the preformed channel with dermal fibroblasts as interstitial cell component of the fully cellularized patch. After 8 days of constant perfusion culture within the same bioreactor, scaffold dynamic modulus and cell viability were analyzed. Endothelial proliferation and vessel maturation were examined by immunohistochemistry and transmission electron microscopy. Our design allowed for scaffold production and dynamic culture in a one-stop-shop model. Enhanced compression and cell-mediated matrix remodeling induced a significant increase in scaffold stiffness while ensuring excellent cell survival. For the first time, we could isolate HLAA-derived EC with proliferative potential. ECs within the central channel proliferated during flow culture, continuously expressing endothelial markers (CD31) and displaying basal membrane synthesis (collagen IV, ultrastructural analysis). After 7 days of culture, a complete endothelial monolayer could be observed. Covering cells aligned themselves in flow direction and developed mature cell-cell contacts.
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http://dx.doi.org/10.1002/term.3129DOI Listing
December 2020

Combination of In Situ Lcn2 pRNA-RNAi Nanotherapeutics and iNSC Transplantation Ameliorates Experimental SCI in Mice.

Mol Ther 2020 12 5;28(12):2677-2690. Epub 2020 Aug 5.

Department of Clinical Neurosciences and NIHR Biomedical Research Centre, University of Cambridge, CB2 0HA Cambridge, UK. Electronic address:

Spinal cord injury (SCI) is a debilitating neurological condition characterized by different cellular and molecular mechanisms that interplay in exacerbating the progression of the pathology. No fully restorative therapies are yet available, and it is thus becoming recognized that combinatorial approaches aimed at addressing different aspects of SCI will likely results in greater functional outcomes. Here we employed packaging RNA-mediated RNA interference (pRNA-RNAi) nanotherapeutics to downregulate in situ the expression of lipocalin 2 (Lcn2), a known mediator of neuroinflammation and autocrine mediator of reactive astrogliosis, and to create a more amenable niche for the subsequent transplantation of induced neural stem cells (iNSCs). To our knowledge, this is the first approach that takes advantage of the modular and multifunctional pRNA three-way junction platform in the SCI niche, while also exploiting the therapeutic potential of immune-compatible and feasible iNSC transplants. We show the combination of such treatments in a mouse model of contusion thoracic SCI leads to significant improvement of locomotor function, albeit not better than single pRNA-RNAi treatment, and results in synergistic histopathological effects, such as reduction of glial scar volume, diminished pro-inflammatory response, and promotion of neuronal survival. Our results provide evidence for a novel combinatorial approach for treating SCI.
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http://dx.doi.org/10.1016/j.ymthe.2020.08.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7704756PMC
December 2020

Generation of two patient-derived iPSC lines from siblings (LIBUCi001-A and LIBUCi002-A) and a genetically modified iPSC line (JMUi001-A-1) to mimic dilated cardiomyopathy with ataxia (DCMA) caused by a homozygous DNAJC19 mutation.

Stem Cell Res 2020 07 2;46:101856. Epub 2020 Jun 2.

Comprehensive Heart Failure Center (CHFC) and Department of Internal Medicine I, University Hospital Würzburg, Würzburg, Germany; Libin Cardiovascular Institute of Alberta, Department of Cardiac Sciences, University of Calgary, Calgary, Canada. Electronic address:

Dilated cardiomyopathy with ataxia (DCMA) is an autosomal recessive disorder arising from mutations in DNAJC19. Two patient-derived dermal fibroblast cell lines of siblings with the same homozygous splice acceptor site mutation in DNAJC19 (NM_145261.4):c.130-1G>C were reprogrammed into induced pluripotent stem cell (iPSC) lines (LIBUCi001-A and LIBUCi002-A) using non-integrative Sendai virus. Additionally, a third DNAJC19tv (truncation variant) iPSC line (JMUi001-A-1) was generated by CRISPR/Cas9 in healthy control iPSCs (JMUi001-A). All three DCMA iPSC lines present normal karyotypes, high expression of pluripotency markers and the capacity to differentiate into cells of all three germ layers.
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http://dx.doi.org/10.1016/j.scr.2020.101856DOI Listing
July 2020

Transplantation of induced neural stem cells (iNSCs) into chronically demyelinated corpus callosum ameliorates motor deficits.

Acta Neuropathol Commun 2020 06 9;8(1):84. Epub 2020 Jun 9.

Department of Anatomy, Physiology and Genetics, F. Edward Hebert School of Medicine, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Rd, Bethesda, MD, 20814, USA.

Multiple Sclerosis (MS) causes neurologic disability due to inflammation, demyelination, and neurodegeneration. Immunosuppressive treatments can modify the disease course but do not effectively promote remyelination or prevent long term neurodegeneration. As a novel approach to mitigate chronic stage pathology, we tested transplantation of mouse induced neural stem cells (iNSCs) into the chronically demyelinated corpus callosum (CC) in adult mice. Male C57BL/6 mice fed 0.3% cuprizone for 12 weeks exhibited CC atrophy with chronic demyelination, astrogliosis, and microglial activation. Syngeneic iNSCs were transplanted into the CC after ending cuprizone and perfused for neuropathology 2 weeks later. Magnetic resonance imaging (MRI) sequences for magnetization transfer ratio (MTR), diffusion-weighted imaging (T2), and diffusion tensor imaging (DTI) quantified CC pathology in live mice before and after iNSC transplantation. Each MRI technique detected progressive CC pathology. Mice that received iNSCs had normalized DTI radial diffusivity, and reduced astrogliosis post-imaging. A motor skill task that engages the CC is Miss-step wheel running, which demonstrated functional deficits from cuprizone demyelination. Transplantation of iNSCs resulted in marked recovery of running velocity. Neuropathology after wheel running showed that iNSC grafts significantly increased host oligodendrocytes and proliferating oligodendrocyte progenitors, while modulating axon damage. Transplanted iNSCs differentiated along astrocyte and oligodendrocyte lineages, without myelinating, and many remained neural stem cells. Our findings demonstrate the applicability of neuroimaging and functional assessments for pre-clinical interventional trials during chronic demyelination and detect improved function from iNSC transplantation. Directly reprogramming fibroblasts into iNSCs facilitates the future translation towards exogenous autologous cell therapies.
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http://dx.doi.org/10.1186/s40478-020-00960-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7285785PMC
June 2020

A genome-scale map of DNA methylation turnover identifies site-specific dependencies of DNMT and TET activity.

Nat Commun 2020 05 29;11(1):2680. Epub 2020 May 29.

Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.

DNA methylation is considered a stable epigenetic mark, yet methylation patterns can vary during differentiation and in diseases such as cancer. Local levels of DNA methylation result from opposing enzymatic activities, the rates of which remain largely unknown. Here we developed a theoretical and experimental framework enabling us to infer methylation and demethylation rates at 860,404 CpGs in mouse embryonic stem cells. We find that enzymatic rates can vary as much as two orders of magnitude between CpGs with identical steady-state DNA methylation. Unexpectedly, de novo and maintenance methylation activity is reduced at transcription factor binding sites, while methylation turnover is elevated in transcribed gene bodies. Furthermore, we show that TET activity contributes substantially more than passive demethylation to establishing low methylation levels at distal enhancers. Taken together, our work unveils a genome-scale map of methylation kinetics, revealing highly variable and context-specific activity for the DNA methylation machinery.
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http://dx.doi.org/10.1038/s41467-020-16354-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7260214PMC
May 2020

Next-generation disease modeling with direct conversion: a new path to old neurons.

FEBS Lett 2019 12 26;593(23):3316-3337. Epub 2019 Nov 26.

Department of Genomics, Stem Cell Biology & Regenerative Medicine, Institute of Molecular Biology & CMBI, Leopold-Franzens-University Innsbruck, Innsbruck, Austria.

Within just over a decade, human reprogramming-based disease modeling has developed from a rather outlandish idea into an essential part of disease research. While iPSCs are a valuable tool for modeling developmental and monogenetic disorders, their rejuvenated identity poses limitations for modeling age-associated diseases. Direct cell-type conversion of fibroblasts into induced neurons (iNs) circumvents rejuvenation and preserves hallmarks of cellular aging. iNs are thus advantageous for modeling diseases that possess strong age-related and epigenetic contributions and can complement iPSC-based strategies for disease modeling. In this review, we provide an overview of the state of the art of direct iN conversion and describe the key epigenetic, transcriptomic, and metabolic changes that occur in converting fibroblasts. Furthermore, we summarize new insights into this fascinating process, particularly focusing on the rapidly changing criteria used to define and characterize in vitro-born human neurons. Finally, we discuss the unique features that distinguish iNs from other reprogramming-based neuronal cell models and how iNs are relevant to disease modeling.
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http://dx.doi.org/10.1002/1873-3468.13678DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6907729PMC
December 2019

Take the shortcut - direct conversion of somatic cells into induced neural stem cells and their biomedical applications.

FEBS Lett 2019 12 1;593(23):3353-3369. Epub 2019 Dec 1.

Department of Molecular Biology & CMBI, Genomics, Stem Cell Biology & Regenerative Medicine, Leopold-Franzens-University Innsbruck, Austria.

Second-generation reprogramming of somatic cells directly into the cell type of interest avoids induction of pluripotency and subsequent cumbersome differentiation procedures. Several recent studies have reported direct conversion of human somatic cells into stably proliferating induced neural stem cells (iNSCs). Importantly, iNSCs are easier, faster, and more cost-efficient to generate than induced pluripotent stem cells (iPSCs), and also have a higher level of clinical safety. Stably, self-renewing iNSCs can be derived from different cellular sources, such as skin fibroblasts and peripheral blood mononuclear cells, and readily differentiate into neuronal and glial lineages that are indistinguishable from their iPSC-derived counterparts or from NSCs isolated from primary tissues. This review focuses on the derivation and characterization of iNSCs and their biomedical applications. We first outline different approaches to generate iNSCs and then discuss the underlying molecular mechanisms. Finally, we summarize the preclinical validation of iNSCs to highlight that these cells are promising targets for disease modeling, autologous cell therapy, and precision medicine.
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http://dx.doi.org/10.1002/1873-3468.13656DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6916337PMC
December 2019

Generation of two induced pluripotent stem cell lines from skin fibroblasts of sisters carrying a c.1094C>A variation in the SCN10A gene potentially associated with small fiber neuropathy.

Stem Cell Res 2019 03 2;35:101396. Epub 2019 Feb 2.

Department of Neurology, University of Würzburg, Würzburg, Germany. Electronic address:

Induced pluripotent stem cells (iPSC) were derived from human dermal fibroblasts (HDF) of two siblings with small fiber neuropathy (SFN) potentially based on the same variation in SCN10A but exhibiting diverse disease phenotypes. HDF were reprogrammed using a non-integrating mRNA approach and showed robust expression of pluripotency markers. iPSC displayed no chromosomal aberrations and were differentiated into all three germ-layers. These two cell lines with a familial genetic background may provide a useful in vitro tool to investigate the underlying mechanisms leading to different phenotypes caused by the same variation.
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http://dx.doi.org/10.1016/j.scr.2019.101396DOI Listing
March 2019

Identification of Embryonic Neural Plate Border Stem Cells and Their Generation by Direct Reprogramming from Adult Human Blood Cells.

Cell Stem Cell 2019 01 20;24(1):166-182.e13. Epub 2018 Dec 20.

Heidelberg Institute for Stem Cell Technology and Experimental Medicine (HI-STEM gGmbH), Im Neuenheimer Feld 280, 69120 Heidelberg, Germany; Division of Stem Cells and Cancer, German Cancer Research Center (DKFZ), and DKFZ-ZMBH Alliance, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany; German Cancer Consortium (DKTK), 69120 Heidelberg, Germany. Electronic address:

We report the direct reprogramming of both adult human fibroblasts and blood cells into induced neural plate border stem cells (iNBSCs) by ectopic expression of four neural transcription factors. Self-renewing, clonal iNBSCs can be robustly expanded in defined media while retaining multilineage differentiation potential. They generate functional cell types of neural crest and CNS lineages and could be used to model a human pain syndrome via gene editing of SCN9A in iNBSCs. NBSCs can also be derived from human pluripotent stem cells and share functional and molecular features with NBSCs isolated from embryonic day 8.5 (E8.5) mouse neural folds. Single-cell RNA sequencing identified the anterior hindbrain as the origin of mouse NBSCs, with human iNBSCs sharing a similar regional identity. In summary, we identify embryonic NBSCs and report their generation by direct reprogramming in human, which may facilitate insights into neural development and provide a neural stem cell source for applications in regenerative medicine.
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http://dx.doi.org/10.1016/j.stem.2018.11.015DOI Listing
January 2019

Need for high-resolution Genetic Analysis in iPSC: Results and Lessons from the ForIPS Consortium.

Sci Rep 2018 11 21;8(1):17201. Epub 2018 Nov 21.

Institute of Human Genetics, University Hospital Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Schwabachanlage 10, 91054, Erlangen, Germany.

Genetic integrity of induced pluripotent stem cells (iPSCs) is essential for their validity as disease models and for potential therapeutic use. We describe the comprehensive analysis in the ForIPS consortium: an iPSC collection from donors with neurological diseases and healthy controls. Characterization included pluripotency confirmation, fingerprinting, conventional and molecular karyotyping in all lines. In the majority, somatic copy number variants (CNVs) were identified. A subset with available matched donor DNA was selected for comparative exome sequencing. We identified single nucleotide variants (SNVs) at different allelic frequencies in each clone with high variability in mutational load. Low frequencies of variants in parental fibroblasts highlight the importance of germline samples. Somatic variant number was independent from reprogramming, cell type and passage. Comparison with disease genes and prediction scores suggest biological relevance for some variants. We show that high-throughput sequencing has value beyond SNV detection and the requirement to individually evaluate each clone.
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http://dx.doi.org/10.1038/s41598-018-35506-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6249203PMC
November 2018

Generation of Cardiomyocytes From Vascular Adventitia-Resident Stem Cells.

Circ Res 2018 08;123(6):686-699

From the Institute of Anatomy and Cell Biology II (S.R.M., P.W., J.B., O.S., N.W., L.R., K.L., G.E., C.K.K., F.E., V.P., S.K., S.E.).

Rationale: Regeneration of lost cardiomyocytes is a fundamental unresolved problem leading to heart failure. Despite several strategies developed from intensive studies performed in the past decades, endogenous regeneration of heart tissue is still limited and presents a big challenge that needs to be overcome to serve as a successful therapeutic option for myocardial infarction.

Objective: One of the essential prerequisites for cardiac regeneration is the identification of endogenous cardiomyocyte progenitors and their niche that can be targeted by new therapeutic approaches. In this context, we hypothesized that the vascular wall, which was shown to harbor different types of stem and progenitor cells, might serve as a source for cardiac progenitors.

Methods And Results: We describe generation of spontaneously beating mouse aortic wall-derived cardiomyocytes without any genetic manipulation. Using aortic wall-derived cells (AoCs) of WT (wild type), αMHC (α-myosin heavy chain), and Flk1 (fetal liver kinase 1)-reporter mice and magnetic bead-associated cell sorting sorting of Flk1 AoCs from GFP (green fluorescent protein) mice, we identified Flk1CD (cluster of differentiation) 34Sca-1 (stem cell antigen-1)-CD44 AoCs as the population that gives rise to aortic wall-derived cardiomyocytes. This AoC subpopulation delivered also endothelial cells and macrophages with a particular accumulation within the aortic wall-derived cardiomyocyte containing colonies. In vivo, cardiomyocyte differentiation capacity was studied by implantation of fluorescently labeled AoCs into chick embryonic heart. These cells acquired cardiomyocyte-like phenotype as shown by αSRA (α-sarcomeric actinin) expression. Furthermore, coronary adventitial Flk1 and CD34 cells proliferated, migrated into the myocardium after mouse myocardial infarction, and expressed Isl-1 (insulin gene enhancer protein-1) indicative of cardiovascular progenitor potential.

Conclusions: Our data suggest Flk1CD34 vascular adventitia-resident stem cells, including those of coronary adventitia, as a novel endogenous source for generating cardiomyocytes. This process is essentially supported by endothelial cells and macrophages. In summary, the therapeutic manipulation of coronary adventitia-resident cardiac stem and their supportive cells may open new avenues for promoting cardiac regeneration and repair after myocardial infarction and for preventing heart failure.
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http://dx.doi.org/10.1161/CIRCRESAHA.117.312526DOI Listing
August 2018

Metabolic substrate shift in human induced pluripotent stem cells during cardiac differentiation: Functional assessment using in vitro radionuclide uptake assay.

Int J Cardiol 2018 Oct 21;269:229-234. Epub 2018 Jun 21.

Comprehensive Heart Failure Center, University Hospital of Würzburg, Würzburg, Germany; Department of Nuclear Medicine, University Hospital of Würzburg, Würzburg, Germany; Department of Biomedical Imaging, National Cerebral and Cardiovascular Research Center, Suita, Japan; Division of Medical Technology and Science, Department of Medical Physics and Engineering, Course of Health Science, Osaka University Graduate School of Medicine, Suita, Japan. Electronic address:

Background: Recent developments in cellular reprogramming technology enable the production of virtually unlimited numbers of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM). Although hiPSC-CM share various characteristic hallmarks with endogenous cardiomyocytes, it remains a question as to what extent metabolic characteristics are equivalent to mature mammalian cardiomyocytes. Here we set out to functionally characterize the metabolic status of hiPSC-CM in vitro by employing a radionuclide tracer uptake assay.

Material And Methods: Cardiac differentiation of hiPSC was induced using a combination of well-orchestrated extrinsic stimuli such as WNT activation (by CHIR99021) and BMP signalling followed by WNT inhibition and lactate based cardiomyocyte enrichment. For characterization of metabolic substrates, dual tracer uptake studies were performed with F‑2‑fluoro‑2‑deoxy‑d‑glucose (F-FDG) and I‑β‑methyl‑iodophenyl‑pentadecanoic acid (I-BMIPP) as transport markers of glucose and fatty acids, respectively.

Results: After cardiac differentiation of hiPSCs, in vitro tracer uptake assays confirmed metabolic substrate shift from glucose to fatty acids that was comparable to those observed in native isolated human cardiomyocytes. Immunostaining further confirmed expression of fatty acid transport and binding proteins on hiPSC-CM.

Conclusions: During in vitro cardiac maturation, we observed a metabolic shift to fatty acids, which are known as a main energy source of mammalian hearts, suggesting hi-PSC-CM as a potential functional phenotype to investigate alteration of cardiac metabolism in cardiac diseases. Results also highlight the use of available clinical nuclear medicine tracers as functional assays in stem cell research for improved generation of autologous differentiated cells for numerous biomedical applications.
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http://dx.doi.org/10.1016/j.ijcard.2018.06.089DOI Listing
October 2018

Generation of the human induced pluripotent stem cell line (UKWNLi001-A) from skin fibroblasts of a woman with Fabry disease carrying the X-chromosomal heterozygous c.708 G > C (W236C) missense mutation in exon 5 of the alpha-galactosidase-A gene.

Stem Cell Res 2018 08 10;31:222-226. Epub 2018 Aug 10.

Department of Neurology, University of Würzburg, Würzburg, Germany. Electronic address:

Human dermal fibroblasts (HDF) were isolated from the skin punch biopsy of a 25-year-old woman with Fabry disease (FD), carrying a heterozygous c.708 G > C missense mutation in the alpha-galactosidase A gene. HDF were reprogrammed to induced pluripotent stem cells (iPSC) using synthetic mRNA, preventing the alteration of the genome and retaining the original genotype. FD-W236C-iPSC (UKWNLi001-A) showed typical human embryonic stem cell (hESC)-like morphology, expressed all analyzed pluripotency-associated markers, could be differentiated into cells from all three germ layers, and demonstrated a normal female karyotype. We provide a novel patient-specific cell line, allowing further insights into the pathophysiology of FD. Resource table.
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http://dx.doi.org/10.1016/j.scr.2018.08.009DOI Listing
August 2018

Mitochondrial Aging Defects Emerge in Directly Reprogrammed Human Neurons due to Their Metabolic Profile.

Cell Rep 2018 05;23(9):2550-2558

Laboratory of Genetics, The Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, CA 92037, USA. Electronic address:

Mitochondria are a major target for aging and are instrumental in the age-dependent deterioration of the human brain, but studying mitochondria in aging human neurons has been challenging. Direct fibroblast-to-induced neuron (iN) conversion yields functional neurons that retain important signs of aging, in contrast to iPSC differentiation. Here, we analyzed mitochondrial features in iNs from individuals of different ages. iNs from old donors display decreased oxidative phosphorylation (OXPHOS)-related gene expression, impaired axonal mitochondrial morphologies, lower mitochondrial membrane potentials, reduced energy production, and increased oxidized proteins levels. In contrast, the fibroblasts from which iNs were generated show only mild age-dependent changes, consistent with a metabolic shift from glycolysis-dependent fibroblasts to OXPHOS-dependent iNs. Indeed, OXPHOS-induced old fibroblasts show increased mitochondrial aging features similar to iNs. Our data indicate that iNs are a valuable tool for studying mitochondrial aging and support a bioenergetic explanation for the high susceptibility of the brain to aging.
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http://dx.doi.org/10.1016/j.celrep.2018.04.105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6478017PMC
May 2018

Macrophage-Derived Extracellular Succinate Licenses Neural Stem Cells to Suppress Chronic Neuroinflammation.

Cell Stem Cell 2018 03 22;22(3):355-368.e13. Epub 2018 Feb 22.

Department of Clinical Neurosciences and NIHR Biomedical Research Centre, University of Cambridge, Cambridge, UK. Electronic address:

Neural stem cell (NSC) transplantation can influence immune responses and suppress inflammation in the CNS. Metabolites, such as succinate, modulate the phenotype and function of immune cells, but whether and how NSCs are also activated by such immunometabolites to control immunoreactivity and inflammatory responses is unclear. Here, we show that transplanted somatic and directly induced NSCs ameliorate chronic CNS inflammation by reducing succinate levels in the cerebrospinal fluid, thereby decreasing mononuclear phagocyte (MP) infiltration and secondary CNS damage. Inflammatory MPs release succinate, which activates succinate receptor 1 (SUCNR1)/GPR91 on NSCs, leading them to secrete prostaglandin E2 and scavenge extracellular succinate with consequential anti-inflammatory effects. Thus, our work reveals an unexpected role for the succinate-SUCNR1 axis in somatic and directly induced NSCs, which controls the response of stem cells to inflammatory metabolic signals released by type 1 MPs in the chronically inflamed brain.
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http://dx.doi.org/10.1016/j.stem.2018.01.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5842147PMC
March 2018

Generation of a human induced pluripotent stem cell (iPSC) line from a 51-year-old female with attention-deficit/hyperactivity disorder (ADHD) carrying a duplication of SLC2A3.

Stem Cell Res 2018 04 13;28:136-140. Epub 2018 Feb 13.

Division of Molecular Psychiatry, Center of Mental Health, University of Würzburg, Würzburg, Germany; Laboratory of Psychiatric Neurobiology, Institute of Molecular Medicine, Sechenov First Moscow State Medical University, Moscow, Russia; Department of Translational Neuroscience, School for Mental Health and Neuroscience (MHeNS), Maastricht University, Maastricht, The Netherlands. Electronic address:

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.
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http://dx.doi.org/10.1016/j.scr.2018.02.005DOI Listing
April 2018

BMP/SMAD Pathway Promotes Neurogenesis of Midbrain Dopaminergic Neurons and in Human Induced Pluripotent and Neural Stem Cells.

J Neurosci 2018 02 10;38(7):1662-1676. Epub 2018 Jan 10.

Department of Physiology and Cell Biology, Zlotowski Center for Neuroscience, Faculty of Health Sciences, Ben-Gurion University of the Negev, 84105 Be'er Sheva, Israel,

The embryonic formation of midbrain dopaminergic (mDA) neurons provides critical guidelines for the differentiation of mDA neurons from stem cells, which are currently being developed for Parkinson's disease cell replacement therapy. Bone morphogenetic protein (BMP)/SMAD inhibition is routinely used during early steps of stem cell differentiation protocols, including for the generation of mDA neurons. However, the function of the BMP/SMAD pathway for specification of mammalian mDA neurons is virtually unknown. Here, we report that BMP5/7-deficient mice (; ) lack mDA neurons due to reduced neurogenesis in the mDA progenitor domain. As molecular mechanisms accounting for these alterations in ; mutants, we have identified expression changes of the BMP/SMAD target genes MSX1/2 (msh homeobox 1/2) and SHH (sonic hedgehog). Conditionally inactivating SMAD1 in neural stem cells of mice () hampered the differentiation of progenitor cells into mDA neurons by preventing cell cycle exit, especially of THSOX6 (tyrosine hydroxylase, SRY-box 6) and THGIRK2 (potassium voltage-gated channel subfamily-J member-6) substantia nigra neurons. BMP5/7 robustly increased the differentiation of human induced pluripotent stem cells and induced neural stem cells to mDA neurons by up to threefold. In conclusion, we have identified BMP/SMAD signaling as a novel critical pathway orchestrating essential steps of mammalian mDA neurogenesis that balances progenitor proliferation and differentiation. Moreover, we demonstrate the potential of BMPs to improve the generation of stem-cell-derived mDA neurons , highlighting the importance of sequential BMP/SMAD inhibition and activation in this process. We identify bone morphogenetic protein (BMP)/SMAD signaling as a novel essential pathway regulating the development of mammalian midbrain dopaminergic (mDA) neurons and provide insights into the molecular mechanisms of this process. BMP5/7 regulate MSX1/2 (msh homeobox 1/2) and SHH (sonic hedgehog) expression to direct mDA neurogenesis. Moreover, the BMP signaling component SMAD1 controls the differentiation of mDA progenitors, particularly to substantia nigra neurons, by directing their cell cycle exit. Importantly, BMP5/7 increase robustly the differentiation of human induced pluripotent and induced neural stem cells to mDA neurons. BMP/SMAD are routinely inhibited in initial stages of stem cell differentiation protocols currently being developed for Parkinson's disease cell replacement therapies. Therefore, our findings on opposing roles of the BMP/SMAD pathway during mDA neurogenesis might improve these procedures significantly.
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http://dx.doi.org/10.1523/JNEUROSCI.1540-17.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5815451PMC
February 2018

A novel conditional Aire allele enables cell-specific ablation of the immune tolerance regulator Aire.

Eur J Immunol 2018 03 27;48(3):546-548. Epub 2017 Dec 27.

Laboratory of Immunobiology, Institute of Molecular Genetics of the Czech Academy of Sciences, Prague, Czech Republic.

Medullary thymic epithelial cell (mTEC)-restricted expression of autoimmune regulator (Aire) is essential for establishment of immune tolerance. Recently, Aire was also shown to be expressed in cells of hematopietic and reproductive lineages. Thus, the generation of Aire mouse strain enables the investigation of the cell-specific function of Aire.
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http://dx.doi.org/10.1002/eji.201747267DOI Listing
March 2018

Cadherin-13 Deficiency Increases Dorsal Raphe 5-HT Neuron Density and Prefrontal Cortex Innervation in the Mouse Brain.

Front Cell Neurosci 2017 26;11:307. Epub 2017 Sep 26.

Division of Molecular Psychiatry, Center of Mental Health, University of Würzburg, Würzburg, Germany.

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. 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 loss-of-function mutations on brain 5-HT system development was explored in a mouse model of Cdh13 deficiency. 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. 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.
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http://dx.doi.org/10.3389/fncel.2017.00307DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5623013PMC
September 2017

Scalable stirred suspension culture for the generation of billions of human induced pluripotent stem cells using single-use bioreactors.

J Tissue Eng Regen Med 2018 02 2;12(2):e1076-e1087. Epub 2017 Oct 2.

Stem Cell and Regenerative Medicine Group, Institute of Anatomy and Cell Biology II, University of Würzburg, Würzburg, Germany.

The production of human induced pluripotent stem cells (hiPSCs) in quantities that are relevant for cell-based therapies and cell-loaded implants through standard adherent culture is hardly achievable and lacks process scalability. A promising approach to overcoming these hurdles is the culture of hiPSCs in suspension. In this study, stirred suspension culture vessels were investigated for their suitability in the expansion of two hiPSC lines inoculated as a single cell suspension, with a free scalability between volumes of 50 and 2400 ml. The simple and robust two-step process reported here first generates hiPSC aggregates of 324 ± 71 μm diameter in 7 days in 125 ml spinner flasks (100 ml volume). These are subsequently dissociated into a single cell suspension for inoculation in 3000 ml bioreactors (1000 ml volume), finally yielding hiPSC aggregates of 198 ± 58 μm after 7 additional days. In both spinner flasks and bioreactors, hiPSCs can be cultured as aggregates for more than 40 days in suspension, maintain an undifferentiated state as confirmed by the expression of pluripotency markers TRA-1-60, TRA-1-81, SSEA-4, OCT4, and SOX2, can differentiate into cells of all three germ layers, and can be directed to differentiate into specific lineages such as cardiomyocytes. Up to a 16-fold increase in hiPSC quantity at the 100 ml volume was achieved, corresponding to a fold increase per day of 2.28; at the 1000 ml scale, an additional 10-fold increase was achieved. Taken together, 16 × 10 hiPSCs were expanded into 2 × 10 hiPSCs in 14 days for a fold increase per day of 8.93. This quantity of hiPSCs readily meets the requirements of cell-based therapies and brings their clinical potential closer to fruition.
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http://dx.doi.org/10.1002/term.2435DOI Listing
February 2018

Establishment of a Human Blood-Brain Barrier Co-culture Model Mimicking the Neurovascular Unit Using Induced Pluri- and Multipotent Stem Cells.

Stem Cell Reports 2017 04 23;8(4):894-906. Epub 2017 Mar 23.

University Hospital Würzburg, Chair Tissue Engineering and Regenerative Medicine, 97070 Würzburg, Germany; Translational Center Würzburg "Regenerative Therapies for Oncology and Musculoskeletal Diseases", Branch of Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 97070 Würzburg, Germany. Electronic address:

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 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.
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http://dx.doi.org/10.1016/j.stemcr.2017.02.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5390136PMC
April 2017

Abrogation of Gap Junctional Communication in ES Cells Results in a Disruption of Primitive Endoderm Formation in Embryoid Bodies.

Stem Cells 2017 04 20;35(4):859-871. Epub 2016 Dec 20.

LIMES Institute, University of Bonn, Bonn, Germany.

Gap junctional intercellular communication (GJIC) has been suggested to be involved in early embryonic development but the actual functional role remained elusive. Connexin (Cx) 43 and Cx45 are co-expressed in embryonic stem (ES) cells, form gap junctions and are considered to exhibit adhesive function and/or to contribute to the establishment of defined communication compartments. Here, we describe the generation of Cx43/Cx45-double deficient mouse ES cells to achieve almost complete breakdown of GJIC. Cre-loxP induced deletion of both, Cx43 and Cx45, results in a block of differentiation in embryoid bodies (EBs) without affecting pluripotency marker expression and proliferation in ES cells. We demonstrate that GJIC-incompetent ES cells fail to form primitive endoderm in EB cultures, representing the inductive key step of further differentiation events. Lentiviral overexpression of either Cx43 or Cx45 in Cx43/45 mutants rescued the observed phenotype, confirming the specificity and indicating a partially redundant function of both connexins. Upon differentiation GJIC-incompetent ES cells exhibit a strikingly altered subcellular localization pattern of the transcription factor NFATc3. Control EBs exhibit significantly more activated NFATc3 in cellular nuclei than mutant EBs suggesting that Cx-mediated communication is needed for synchronized NFAT activation to induce orchestrated primitive endoderm formation. Moreover, pharmacological inhibition of NFATc3 activation by Cyclosporin A, a well-described inhibitor of calcineurin, phenocopies the loss of GJIC in control cells. Stem Cells 2017;35:859-871.
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http://dx.doi.org/10.1002/stem.2545DOI Listing
April 2017

The vascular adventitia: An endogenous, omnipresent source of stem cells in the body.

Pharmacol Ther 2017 03 3;171:13-29. Epub 2016 Aug 3.

Institute of Anatomy and Cell Biology, University of Würzburg, Germany. Electronic address:

Until a decade ago it was believed that the wall of adult blood vessels exclusively contains terminally differentiated cell types. A paradigm shift was unavoidable since studies from different groups convincingly showed the presence of vascular wall-resident stem and progenitor cells (VW-SCs) which were identified to particularly reside in the sub-endothelial space and the so-called adventitial "vasculogenic zone". Data published during the last decade uncloaked the fact that VW-SCs have the capacity to differentiate into both vascular and non-vascular cell types. Up to date, little is known about the full capacity of VW-SCs, the exact composition of their endogenous niche and the mechanisms that govern their self-renewal, activation and differentiation. The aim of this review is to provide an overview about the current knowledge on VW-SCs and to highlight the impact of this endogenous niche on health and disease. In addition, we will discuss strategies how these adult stem cells could be manipulated in order to activate and expand them, ideally within their niche at sites of tissue damage and subsequently differentiate them into a desired cell type, e.g. an endothelial cell, a macrophage or a muscle cell. This would pave the way towards new pharmacological strategies for endogenous tissue repair and regeneration.
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http://dx.doi.org/10.1016/j.pharmthera.2016.07.017DOI Listing
March 2017

Low-density lipoprotein receptor-related protein 1 is a novel modulator of radial glia stem cell proliferation, survival, and differentiation.

Glia 2016 08 3;64(8):1363-80. Epub 2016 Jun 3.

Department of Cell Morphology and Molecular Neurobiology, Faculty of Biology and Biotechnology, Ruhr-University, Bochum, D-44780, Germany.

The LDL family of receptors and its member low-density lipoprotein receptor-related protein 1 (LRP1) have classically been associated with a modulation of lipoprotein metabolism. Current studies, however, indicate diverse functions for this receptor in various aspects of cellular activities, including cell proliferation, migration, differentiation, and survival. LRP1 is essential for normal neuronal function in the adult CNS, whereas the role of LRP1 in development remained unclear. Previously, we have observed an upregulation of LewisX (LeX) glycosylated LRP1 in the stem cells of the developing cortex and demonstrated its importance for oligodendrocyte differentiation. In the current study, we show that LeX-glycosylated LRP1 is also expressed in the stem cell compartment of the developing spinal cord and has broader functions in the developing CNS. We have investigated the basic properties of LRP1 conditional knockout on the neural stem/progenitor cells (NSPCs) from the cortex and the spinal cord, created by means of Cre-loxp-mediated recombination in vitro. The functional status of LRP1-deficient cells has been studied using proliferation, differentiation, and apoptosis assays. LRP1 deficient NSPCs from both CNS regions demonstrated altered differentiation profiles. Their differentiation capacity toward oligodendrocyte progenitor cells (OPCs), mature oligodendrocytes and neurons was reduced. In contrast, astrocyte differentiation was promoted. Moreover, LRP1 deletion had a negative effect on NSPCs proliferation and survival. Our observations suggest that LRP1 facilitates NSPCs differentiation via interaction with apolipoprotein E (ApoE). Upon ApoE4 stimulation wild type NSPCs generated more oligodendrocytes, but LRP1 knockout cells showed no response. The effect of ApoE seems to be independent of cholesterol uptake, but is rather mediated by downstream MAPK and Akt activation. GLIA 2016 GLIA 2016;64:1363-1380.
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http://dx.doi.org/10.1002/glia.23009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5033964PMC
August 2016