Publications by authors named "Ulrich Martin"

144 Publications

Reprogramming enriches for somatic cell clones with small scale mutations in cancer-associated genes.

Mol Ther 2021 Apr 5. Epub 2021 Apr 5.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic-, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany. Electronic address:

Cellular therapies based on induced pluripotent stem cells (iPSCs) come out of age and an increasing number of clinical trials applying iPSC-based transplants are ongoing or in preparation. Recent studies, however, demonstrated a high number of small scale mutations in iPSCs. Although, the mutational load in iPSCs seems to be largely derived from their parental cells, it is still unknown whether reprogramming may enrich for individual mutations that could lead to loss of functionality and tumor formation from iPSC derivatives. 30 hiPSC lines were analyzed by whole exome sequencing. High accuracy amplicon sequencing showed that all analyzed small scale variants pre-existed in their parental cells and that individual mutations present in small subpopulations of parental cells become enriched among hiPSC clones during reprogramming. Among those, putatively actionable driver mutations affect genes related to cell cycle control, cell death and pluripotency, and may confer a selective advantage during reprogramming. Finally, a shRNA-based experimental approach was applied to provide additional evidence for the individual impact of such genes on the reprogramming efficiency. In conclusion, we show that enriched mutations in curated onco- and tumor suppressor genes may account for an increased tumor risk and impact the clinical value of patient-derived hiPSCs.
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http://dx.doi.org/10.1016/j.ymthe.2021.04.007DOI Listing
April 2021

An early cell shape transition drives evolutionary expansion of the human forebrain.

Cell 2021 Mar 19. Epub 2021 Mar 19.

MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Francis Crick Avenue, Cambridge CB2 0QH, UK. Electronic address:

The human brain has undergone rapid expansion since humans diverged from other great apes, but the mechanism of this human-specific enlargement is still unknown. Here, we use cerebral organoids derived from human, gorilla, and chimpanzee cells to study developmental mechanisms driving evolutionary brain expansion. We find that neuroepithelial differentiation is a protracted process in apes, involving a previously unrecognized transition state characterized by a change in cell shape. Furthermore, we show that human organoids are larger due to a delay in this transition, associated with differences in interkinetic nuclear migration and cell cycle length. Comparative RNA sequencing (RNA-seq) reveals differences in expression dynamics of cell morphogenesis factors, including ZEB2, a known epithelial-mesenchymal transition regulator. We show that ZEB2 promotes neuroepithelial transition, and its manipulation and downstream signaling leads to acquisition of nonhuman ape architecture in the human context and vice versa, establishing an important role for neuroepithelial cell shape in human brain expansion.
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http://dx.doi.org/10.1016/j.cell.2021.02.050DOI Listing
March 2021

Fear extinction learning and anandamide: an fMRI study in healthy humans.

Transl Psychiatry 2021 Mar 15;11(1):161. Epub 2021 Mar 15.

Clinic for Psychiatry/Psychotherapy III, Ulm University, Ulm, Germany.

Anxiety- and trauma-related disorders are severe illnesses with high prevalence. Current treatment options leave room for improvement and the endocannabinoid system (ECS) has become a key target in psychopharmacological research. Rodent models suggest an anxiolytic effect of endocannabinoids and demonstrated that the ECS is involved in the modulation of fear learning and aversive memory consolidation. So far, one prominent target was inhibition of fatty acid amino hydrolase (FAAH), the degrading enzyme of the endocannabinoid anandamide (AEA). Research in humans remains scarce, but genetic studies have found that the single-nucleotide polymorphism (SNP) FAAH C385A (rs324420) is associated with lower catabolic performance of FAAH and increased levels of AEA. Translational research on the ECS in fear learning processes is rare, yet crucial to understand the mechanisms involved. To address this lack of research, we designed a fear conditioning, extinction learning paradigm with 51 healthy, male humans who underwent functional magnetic resonance imaging (fMRI) before analysing baseline and task-related changes of AEA, as well as the FAAH polymorphism (rs324420). The results indicate higher AEA levels in AC-heterozygotes than in CC-individuals (SNP rs324420), but no difference between the groups during extinction learning. However, neural activation of the anterior cingulate cortex and anterior insular cortex during extinction learning correlated positively with AEA baseline levels, and task-related changes in AEA were found particularly during fear extinction, with a modulatory effect on neural activation related to extinction learning. Results indicate a putative role for AEA in fear extinction learning. Pre-treatment with AEA-enhancing drugs could promote extinction learning during psychotherapeutic interventions.
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http://dx.doi.org/10.1038/s41398-020-01177-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7961038PMC
March 2021

High density bioprocessing of human pluripotent stem cells by metabolic control and in silico modeling.

Stem Cells Transl Med 2021 Mar 4. Epub 2021 Mar 4.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Germany.

To harness the full potential of human pluripotent stem cells (hPSCs) we combined instrumented stirred tank bioreactor (STBR) technology with the power of in silico process modeling to overcome substantial, hPSC-specific hurdles toward their mass production. Perfused suspension culture (3D) of matrix-free hPSC aggregates in STBRs was applied to identify and control process-limiting parameters including pH, dissolved oxygen, glucose and lactate levels, and the obviation of osmolality peaks provoked by high density culture. Media supplements promoted single cell-based process inoculation and hydrodynamic aggregate size control. Wet lab-derived process characteristics enabled predictive in silico modeling as a new rational for hPSC cultivation. Consequently, hPSC line-independent maintenance of exponential cell proliferation was achieved. The strategy yielded 70-fold cell expansion in 7 days achieving an unmatched density of 35 × 10 cells/mL equivalent to 5.25 billion hPSC in 150 mL scale while pluripotency, differentiation potential, and karyotype stability was maintained. In parallel, media requirements were reduced by 75% demonstrating the outstanding increase in efficiency. Minimal input to our in silico model accurately predicts all main process parameters; combined with calculation-controlled hPSC aggregation kinetics, linear process upscaling is also enabled and demonstrated for up to 500 mL scale in an independent bioreactor system. Thus, by merging applied stem cell research with recent knowhow from industrial cell fermentation, a new level of hPSC bioprocessing is revealed fueling their automated production for industrial and therapeutic applications.
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http://dx.doi.org/10.1002/sctm.20-0453DOI Listing
March 2021

Production and cryopreservation of definitive endoderm from human pluripotent stem cells under defined and scalable culture conditions.

Nat Protoc 2021 03 12;16(3):1581-1599. Epub 2021 Feb 12.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), REBIRTH-Research Center for Translational and Regenerative Medicine, Hannover Medical School, Hannover, Germany.

The endodermal germ layer gives rise to respiratory epithelium, hepatocytes, pancreatic cells and intestinal lineages, among other cell types. These lineages can be differentiated from human pluripotent stem cells (hPSCs) via a common definitive endoderm (DE) intermediate that is characterized by the co-expression of the cell surface markers CXCR4, c-KIT and EPCAM and the transcription factors SOX17 and FOXA2. Here we provide a detailed protocol for mass production of DE from hPSCs in scalable and easy-to-handle suspension culture using a rotating Erlenmeyer flask or a sophisticated, fully controllable, 150-ml stirred tank bioreactor. This protocol uses two different media formulations that are chemically defined and xeno free and therefore good manufacturing practice ready. Our protocol allows for efficient hPSC-derived DE specification in multicellular aggregates within 3 days and generates up to 1 × 10 DE cells with >92% purity in one differentiation batch when using the bioreactor. The hPSC-derived DE cells that are generated can be cryopreserved for later downstream differentiation into various endodermal lineages. This protocol should facilitate the flexible production of mature DE derivatives for physiologically relevant disease models, high-throughput drug screening, toxicology testing and cellular therapies.
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http://dx.doi.org/10.1038/s41596-020-00470-5DOI Listing
March 2021

Establishment of MHHi001-A-5, a GCaMP6f and RedStar dual reporter human iPSC line for in vitro and in vivo characterization and in situ tracing of iPSC derivatives.

Stem Cell Res 2021 Apr 30;52:102206. Epub 2021 Jan 30.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), REBIRTH Research Center for Translational Regenerative Medicine, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany.

Transgenic hiPSC lines carrying reporter genes represent valuable tools for functional characterization of iPSC derivatives, disease modelling and clinical evaluation of cell therapies. Here, the hiPSC line 'Phoenix' (Haase et al., 2017) was genetically engineered using TALEN-based integration of the calcium sensor GCaMP6f and RedStar reporter into the AAVS1 site. Characterization of undifferentiated cells and functional investigation of hiPSC-derived cardiomyocytes-containing BCTs showed a strong intracellular calcium transient-dependent GCaMP6f and eminent RedStar signal. Therefore, our dual reporter line provides an excellent tool to facilitate monitoring of engraftment, calcium fluctuations and coupling of iPSC derivatives such as cardiomyocytes in vitro and in vivo in animal models.
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http://dx.doi.org/10.1016/j.scr.2021.102206DOI Listing
April 2021

Human heart-forming organoids recapitulate early heart and foregut development.

Nat Biotechnol 2021 Feb 8. Epub 2021 Feb 8.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery (HTTG), REBIRTH-Research Center for Translational Regenerative Medicine, Hannover Medical School, Hannover, Germany.

Organoid models of early tissue development have been produced for the intestine, brain, kidney and other organs, but similar approaches for the heart have been lacking. Here we generate complex, highly structured, three-dimensional heart-forming organoids (HFOs) by embedding human pluripotent stem cell aggregates in Matrigel followed by directed cardiac differentiation via biphasic WNT pathway modulation with small molecules. HFOs are composed of a myocardial layer lined by endocardial-like cells and surrounded by septum-transversum-like anlagen; they further contain spatially and molecularly distinct anterior versus posterior foregut endoderm tissues and a vascular network. The architecture of HFOs closely resembles aspects of early native heart anlagen before heart tube formation, which is known to require an interplay with foregut endoderm development. We apply HFOs to study genetic defects in vitro by demonstrating that NKX2.5-knockout HFOs show a phenotype reminiscent of cardiac malformations previously observed in transgenic mice.
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http://dx.doi.org/10.1038/s41587-021-00815-9DOI Listing
February 2021

Dual Function of iPSC-Derived Pericyte-Like Cells in Vascularization and Fibrosis-Related Cardiac Tissue Remodeling In Vitro.

Int J Mol Sci 2020 Nov 25;21(23). Epub 2020 Nov 25.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany.

Myocardial interstitial fibrosis (MIF) is characterized by excessive extracellular matrix (ECM) deposition, increased myocardial stiffness, functional weakening, and compensatory cardiomyocyte (CM) hypertrophy. Fibroblasts (Fbs) are considered the principal source of ECM, but the contribution of perivascular cells, including pericytes (PCs), has gained attention, since MIF develops primarily around small vessels. The pathogenesis of MIF is difficult to study in humans because of the pleiotropy of mutually influencing pathomechanisms, unpredictable side effects, and the lack of available patient samples. Human pluripotent stem cells (hPSCs) offer the unique opportunity for the de novo formation of bioartificial cardiac tissue (BCT) using a variety of different cardiovascular cell types to model aspects of MIF pathogenesis in vitro. Here, we have optimized a protocol for the derivation of hPSC-derived PC-like cells (iPSC-PCs) and present a BCT in vitro model of MIF that shows their central influence on interstitial collagen deposition and myocardial tissue stiffening. This model was used to study the interplay of different cell types-i.e., hPSC-derived CMs, endothelial cells (ECs), and iPSC-PCs or primary Fbs, respectively. While iPSC-PCs improved the sarcomere structure and supported vascularization in a PC-like fashion, the functional and histological parameters of BCTs revealed EC- and PC-mediated effects on fibrosis-related cardiac tissue remodeling.
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http://dx.doi.org/10.3390/ijms21238947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7728071PMC
November 2020

Fgf10 Signaling-Based Evidence for the Existence of an Embryonic Stage Distinct From the Pseudoglandular Stage During Mouse Lung Development.

Front Cell Dev Biol 2020 22;8:576604. Epub 2020 Oct 22.

Key laboratory of Interventional Pulmonology of Zhejiang Province, Department of Pulmonary and Critical Care Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.

The existence during mouse lung development of an embryonic stage temporally and functionally distinct from the subsequent pseudoglandular stage has been proposed but never demonstrated; while studies in human embryonic lung tissue fail to recapitulate the molecular control of branching found in mice. Lung development in mice starts officially at embryonic day (E) 9.5 when on the ventral side of the primary foregut tube, both the trachea and the two primary lung buds emerge and elongate to form a completely separate structure from the foregut by E10. In the subsequent 6 days, the primary lung buds undergo an intense process of branching to form a ramified tree by E16.5. We used transgenic mice allowing to transiently inhibit endogenous fibroblast growth factor 10 (Fgf10) activity in mutant embryos at E9, E9.5, and E11 upon intraperitoneal exposure to doxycycline and examined the resulting lung phenotype at later developmental stages. We also determined using gene arrays the transcriptomic response of flow cytometry-isolated human alveolar epithelial progenitor cells derived from hESC or hiPSC, grown for 12 or 24 h, in the presence or absence of recombinant FGF10. Following injection at E9, the corresponding mutant lungs at E18.5 appear almost normal in size and shape but close up examination indicate failure of the right lung to undergo lobar septation. At E9.5, the lungs are slightly hypoplastic but display normal differentiation and functionality. However, at E11, the lungs show impaired branching and are no longer functional. Using gene array data, we report only a partial overlap between human and mouse in the genes previously shown to be regulated by Fgf10 at E12.5. This study supports the existence of an embryonic stage of lung development where Fgf10 signaling does not play a function in the branching process but rather in lobar septation. It also posits that functional comparisons between mouse and human organogenesis must account for these distinct stages.
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http://dx.doi.org/10.3389/fcell.2020.576604DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7642470PMC
October 2020

Generation of two hiPSC clones (MHHi019-A, MHHi019-B) from a primary ciliary dyskinesia patient carrying a homozygous deletion in the NME5 gene (c.415delA (p.Ile139Tyrfs*8)).

Stem Cell Res 2020 10 7;48:101988. Epub 2020 Sep 7.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Germany. Electronic address:

Primary ciliary dyskinesia (PCD) is a genetic disorder characterized by defects in motile cilia and is known to occur in about 1 in 20,000 live births (Horani and Ferkol, 2018). Among the many genes associated with PCD, NME5, a gene encoding a protein involved in ciliary function, was recently reported to be involved in PCD (Anderegg et al., 2019; Cho et al., 2020). We have established two human induced pluripotent stem cell clones from a PCD patient carrying a deletion in the NME5 gene (c.415delA (p.Ile139Tyrfs*8)).
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http://dx.doi.org/10.1016/j.scr.2020.101988DOI Listing
October 2020

Generation of two human induced pluripotent stem cell lines (MHHi017-A, MHHi017-B) from a patient with primary ciliary dyskinesia carrying a homozygous mutation (c.7915C > T [p.Arg2639*]) in the DNAH5 gene.

Stem Cell Res 2020 07 20;46:101848. Epub 2020 May 20.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Germany; REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany.

Dynein axonemal heavy chain 5 (DNAH5) is part of a microtubule-associated protein complex found within the cilia of the lung. Mutations in the DNAH5 gene lead to impaired ciliary function and are linked to primary ciliary dyskinesia (PCD), a rare autosomal recessive disorder. We established two human induced pluripotent stem cell (hiPSC) lines generated from a patient with PCD and homozygous mutation in the corresponding DNAH5 gene. These cell lines represent an excellent tool for modeling the ciliary dysfunction in PCD.
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http://dx.doi.org/10.1016/j.scr.2020.101848DOI Listing
July 2020

Generation of two hiPSC lines (MHHi016-A, MHHi016-B) from a primary ciliary dyskinesia patient carrying a homozygous 5 bp duplication (c.248_252dup (p.Gly85Cysfs*11)) in exon 1 of the CCNO gene.

Stem Cell Res 2020 07 18;46:101850. Epub 2020 May 18.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Germany.

Cyclin O (CCNO) is involved in cell cycle regulation and mutations of CCNO are linked to the rare genetic disease primary ciliary dyskinesia (PCD). Mutations in CCNO are associated with reduced cilia number and cilia agenesis on epithelia of the respiratory tract. This article deals with the description of two hiPSC lines generated from a PCD patient carrying a mutation in exon 1 of the CCNO gene. The lines offer a valuable tool for in vitro modeling PCD pathophysiology.
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http://dx.doi.org/10.1016/j.scr.2020.101850DOI Listing
July 2020

and Interspecies Chimera Assay Using Early Pig Embryos.

Cell Reprogram 2020 06 19;22(3):118-133. Epub 2020 May 19.

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut, Neustadt, Germany.

Chimeric pigs harboring organs derived from human stem cells are promising for patient-specific regenerative therapies. Induced pluripotent stem cells (iPSCs) can contribute to all cell types of the fetus, including germline after injection into embryos. However, ethical concerns prohibit testing human iPSCs in chimera assays. Here, we evaluated porcine embryos as hosts for an interspecies chimera assay using iPSCs from either cynomolgus monkeys (cyiPSCs) or mouse (miPSCs). To establish an culture system compatible for cyiPSCs and porcine embryos, we determined blastocyst development in eight different stem cell media. The highest developmental rates of blastocysts were achieved in Knockout Dulbecco's modified Eagle's medium with 20% knockout serum replacement. We found that cyiPSCs injected into porcine embryos survived and were mostly located in the trophectoderm (TE). Instead, when miPSCs were injected into porcine embryos, the cells rapidly proliferated. The behavior of chimeras developed was recapitulated ; cyiPSCs were observed in the TE, but not in the porcine epiblast. However, when miPSCs were injected into derived porcine embryos, mouse cells were found in both, the epiblast and TE. These results demonstrate that porcine embryos could be useful for evaluating the interspecies chimera-forming ability of iPSCs from different species.
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http://dx.doi.org/10.1089/cell.2019.0107DOI Listing
June 2020

Advanced Single-Cell Mapping Reveals that in hESC Cardiomyocytes Contraction Kinetics and Action Potential Are Independent of Myosin Isoform.

Stem Cell Reports 2020 05 16;14(5):788-802. Epub 2020 Apr 16.

Institute of Molecular and Cell Physiology, Hannover Medical School, Carl-Neuberg Straße 1, 30625 Hannover, Germany.

Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) represent an attractive model to investigate CM function and disease mechanisms. One characteristic marker of ventricular specificity of human CMs is expression of the ventricular, slow β-myosin heavy chain (MyHC), as opposed to the atrial, fast α-MyHC. The main aim of this study was to investigate at the single-cell level whether contraction kinetics and electrical activity of hESC-CMs are influenced by the relative expression of α-MyHC versus β-MyHC. For effective assignment of functional parameters to the expression of both MyHC isoforms at protein and mRNA levels in the very same hESC-CMs, we developed a single-cell mapping technique. Surprisingly, α- versus β-MyHC was not related to specific contractile or electrophysiological properties of the same cells. The multiparametric cell-by-cell analysis suggests that in hESC-CMs the expression of genes associated with electrical activity, contraction, calcium handling, and MyHCs is independently regulated.
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http://dx.doi.org/10.1016/j.stemcr.2020.03.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7220955PMC
May 2020

Targeted Integration of Inducible Caspase-9 in Human iPSCs Allows Efficient Clearance of iPSCs and iPSC-Macrophages.

Int J Mol Sci 2020 Apr 3;21(7). Epub 2020 Apr 3.

Institute of Experimental Hematology, REBIRTH, Hannover Medical School, Hannover 30625, Germany.

Induced pluripotent stem cells (iPSCs) offer great promise for the field of regenerative medicine, and iPSC-derived cells have already been applied in clinical practice. However, potential contamination of effector cells with residual pluripotent cells (e.g., teratoma-initiating cells) or effector cell-associated side effects may limit this approach. This also holds true for iPSC-derived hematopoietic cells. Given the therapeutic benefit of macrophages in different disease entities and the feasibility to derive macrophages from human iPSCs, we established human iPSCs harboring the inducible Caspase-9 (iCasp9) suicide safety switch utilizing transcription activator-like effector nuclease (TALEN)-based designer nuclease technology. Mono- or bi-allelic integration of the iCasp9 gene cassette into the AAVS1 locus showed no effect on the pluripotency of human iPSCs and did not interfere with their differentiation towards macrophages. In both, iCasp9-mono and iCasp9-bi-allelic clones, concentrations of 0.1 nM AP20187 were sufficient to induce apoptosis in more than 98% of iPSCs and their progeny-macrophages. Thus, here we provide evidence that the introduction of the iCasp9 suicide gene into the AAVS1 locus enables the effective clearance of human iPSCs and thereof derived macrophages.
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http://dx.doi.org/10.3390/ijms21072481DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7177583PMC
April 2020

Generation of an induced pluripotent stem cell line (MHHi018-A) from a patient with Cystic Fibrosis carrying p.Asn1303Lys (N1303K) mutation.

Stem Cell Res 2020 04 25;44:101744. Epub 2020 Mar 25.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), REBIRTH-Research Center for Translational and Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany.

Cystic Fibrosis (CF) is a genetic disease caused by mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene which encodes for a chloride ion channel regulating the balance of salt and water across secretory epithelia. Here we generated an iPSC line from a CF patient homozygous for the p.Asn1303Lys mutation, a Class II folding defect mutation. This iPSC line provides a useful resource for disease modeling and to investigate the pharmacological response to CFTR modulators in iPSC derived epithelia.
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http://dx.doi.org/10.1016/j.scr.2020.101744DOI Listing
April 2020

Generation of three induced pluripotent stem cell lines (MHHi012-A, MHHi013-A, MHHi014-A) from a family with Loeys-Dietz syndrome carrying a heterozygous p.M253I (c.759G>A) mutation in the TGFBR1 gene.

Stem Cell Res 2020 03 4;43:101707. Epub 2020 Feb 4.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH-Cluster of Excellence, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Germany.

Loeys-Dietz syndrome (LDS) is a rare connective tissue disorder characterized by a genetic predisposition for thoracic aortic aneurysm and dissection. Despite heterozygous loss-of-function mutations in genes for ligand, receptor, or downstream mediators of the transforming growth factor β (TGFβ) pathway, LDS is associated with a signature of high TGFβ signaling. We generated induced pluripotent stem cell (iPSC) lines from three adult LDS-patients (two male, one female) of a family with a heterozygous point mutation in exon 4 of the TGFβ-receptor1 (TGFBR1) gene (p.M253I; c.759G>A). The lines offer a valuable resource for modeling the pathophysiology of genetically mediated aortic disease.
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http://dx.doi.org/10.1016/j.scr.2020.101707DOI Listing
March 2020

The grounding of abstract concepts in the motor and visual system: An fMRI study.

Cortex 2020 03 13;124:1-22. Epub 2019 Nov 13.

Department of Psychiatry, Ulm University, Ulm, Germany. Electronic address:

The grounding of concepts in the sensorimotor brain systems is controversially discussed. Grounded cognition models propose that concepts are represented in modality-specific sensorimotor, but also emotional and introspective brain areas depending on specific experiences during concept acquisition. Accumulating evidence suggests that concrete concepts are closely linked to modality-specific systems, whereas the mere existence of abstract concepts seems to contradict grounded cognition approaches. Here, using functional magnetic resonance imaging, we adopted a theory-driven approach frequently used for investigating concrete concepts to the domain of abstract concepts: We compared brain activation to abstract concepts with a known motor versus visual feature content as determined by a previous property listing study. Carefully matched motor (e.g., fitness) and visual (e.g., beauty) abstract words were presented to 24 participants along with pseudowords while performing a lexical decision task. Furthermore, participants performed two localizer tasks by actually moving their hands (motor localizer) and by looking at real pictures (visual localizer). Processing of motor abstract words specifically activated frontal and parietal motor areas, whereas processing of visual abstract words specifically elicited higher activity in temporo-occipital visual areas, albeit at a more lenient statistical threshold. According to inclusive masking analyses, this differential activity pattern to motor and visual abstract concepts overlapped with brain activations observed during hand movements (pre- and postcentral gyrus) and object perception (fusiform and lingual gyrus). Thus, consistent with the grounded cognition framework, our results suggest that, similar to concrete concepts, abstract concepts related to action and vision are grounded in modality-specific brain systems typically engaged in actual perception and action depending on their conceptual feature content.
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http://dx.doi.org/10.1016/j.cortex.2019.10.014DOI Listing
March 2020

Chemically-Defined, Xeno-Free, Scalable Production of hPSC-Derived Definitive Endoderm Aggregates with Multi-Lineage Differentiation Potential.

Cells 2019 12 4;8(12). Epub 2019 Dec 4.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), REBIRTH - Center for Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany.

For the production and bio-banking of differentiated derivatives from human pluripotent stem cells (hPSCs) in large quantities for drug screening and cellular therapies, well-defined and robust procedures for differentiation and cryopreservation are required. Definitive endoderm (DE) gives rise to respiratory and digestive epithelium, as well as thyroid, thymus, liver, and pancreas. Here, we present a scalable, universal process for the generation of DE from human-induced pluripotent stem cells (hiPSCs) and embryonic stem cells (hESCs). Optimal control during the differentiation process was attained in chemically-defined and xeno-free suspension culture, and high flexibility of the workflow was achieved by the introduction of an efficient cryopreservation step at the end of DE differentiation. DE aggregates were capable of differentiating into hepatic-like, pancreatic, intestinal, and lung progenitor cells. Scale-up of the differentiation process using stirred-tank bioreactors enabled production of large quantities of DE aggregates. This process provides a useful advance for versatile applications of DE lineages, in particular for cell therapies and drug screening.
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http://dx.doi.org/10.3390/cells8121571DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6953099PMC
December 2019

Generation of a NKX2.1 - p63 double transgenic knock-in reporter cell line from human induced pluripotent stem cells (MHHi006-A-4).

Stem Cell Res 2020 01 19;42:101659. Epub 2019 Nov 19.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH-Cluster of Excellence, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Germany. Electronic address:

Tumor protein p63 (p63) encodes for a transcription factor of the p53 family and is a marker for respiratory basal cells. Based on a NKX2.1 knock-in reporter cell line from human induced pluripotent stem cells (hiPSCs) (MHHi06-A-2) we established a NKX2.1/p63 double transgenic knock-in reporter cell line using TALEN technology. The reporter enables the optimization and monitoring of hiPSC differentiation towards NKX2.1/p63 double positive cells as well as enrichment for single or double positive cells.
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http://dx.doi.org/10.1016/j.scr.2019.101659DOI Listing
January 2020

Generation of a CFTR knock-in reporter cell line (MHHi006-A-1) from a human induced pluripotent stem cell line.

Stem Cell Res 2019 10 20;40:101542. Epub 2019 Aug 20.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH-Cluster of Excellence, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), German Center for Lung Research (DZL), Germany. Electronic address:

CFTR encodes for a chloride ion channel expressed primarily in secretory epithelia in the airways, intestine, liver and other tissues. Mutations in the CFTR gene have been identified in people suffering from Cystic Fibrosis. Here, we established a CFTR knock-in reporter cell line from a human iPSC line (MHHi006-A) using TALEN technology. The reporter enables the monitoring and optimization of the differentiation of pluripotent stem cells into CFTR expressing epithelia on a single cell level, as well as the enrichment of CFTR positive cells, which represent an excellent tool for Cystic Fibrosis disease modelling, drug screening and ultimately cellular therapies.
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http://dx.doi.org/10.1016/j.scr.2019.101542DOI Listing
October 2019

Continuous WNT Control Enables Advanced hPSC Cardiac Processing and Prognostic Surface Marker Identification in Chemically Defined Suspension Culture.

Stem Cell Reports 2019 08 25;13(2):366-379. Epub 2019 Jul 25.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic-, Transplantation and Vascular Surgery, REBIRTH-Cluster of Excellence, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany. Electronic address:

Aiming at clinical translation, robust directed differentiation of human pluripotent stem cells (hPSCs), preferentially in chemically defined conditions, is a key requirement. Here, feasibility of suspension culture based hPSC-cardiomyocyte (hPSC-CM) production in low-cost, xeno-free media compatible with good manufacturing practice standards is shown. Applying stirred tank bioreactor systems at increasing dimensions, our advanced protocol enables routine production of about 1 million hPSC-CMs/mL, yielding ∼1.3 × 10 CM in 150 mL and ∼4.0 × 10 CMs in 350-500 mL process scale at >90% lineage purity. Process robustness and efficiency is ensured by uninterrupted chemical WNT pathway control at early stages of differentiation and results in the formation of almost exclusively ventricular-like CMs. Modulated WNT pathway regulation also revealed the previously unappreciated role of ROR1/CD13 as superior surrogate markers for predicting cardiac differentiation efficiency as soon as 72 h of differentiation. This monitoring strategy facilitates process upscaling and controlled mass production of hPSC derivatives.
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http://dx.doi.org/10.1016/j.stemcr.2019.06.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6700605PMC
August 2019

Generation of a NKX2.1 knock-in reporter cell line from human induced pluripotent stem cells (MHHi006-A-2).

Stem Cell Res 2019 08 28;39:101492. Epub 2019 Jun 28.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH-Cluster of Excellence, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Germany.

NK homeobox 1 (NKX2.1; also known as thyroid transcription factor 1, TTF-1) encodes for a transcription factor involved in the development of thyroid, lung and brain. Here, we established a NKX2.1 knock-in reporter cell line from human induced pluripotent stem cells (iPSCs) using TALEN technology. The reporter enables the optimization and monitoring of the differentiation of pluripotent stem cells (PSCs) into NKX2.1 expressing cells on a single cell level, as well as the enrichment of NKX2.1 positive cells.
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http://dx.doi.org/10.1016/j.scr.2019.101492DOI Listing
August 2019

M. Peroneus Quartus Causing Chronic Peroneal Compartment Syndrome in a Runner Treated by Endoscopic Fasciotomy: A Case Report.

J Foot Ankle Surg 2019 Jul 23;58(4):653-656. Epub 2019 May 23.

Head of Department, Department of Sports Medicine, Rennbahnklinik, Muttenz, Basel, Switzerland.

We present the rare case of a 47-year-old male long-distance runner who was referred to our hospital with a longstanding pain in his left calf. Clinical history, as well as ultrasound and magnetic resonance imaging scans, showed an accessory peroneal muscle. This muscle was identified as a peroneus quartus muscle. On ultrasound, controlled intracompartmental pressure measurement, a chronic peroneal compartment syndrome, was diagnosed. We performed an endoscopic-assisted fasciotomy of the peroneal compartment. This resolved the patient's symptoms completely and allowed the runner to return to competition shortly after the surgery.
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http://dx.doi.org/10.1053/j.jfas.2018.12.008DOI Listing
July 2019

High-Throughput Screening for Modulators of CFTR Activity Based on Genetically Engineered Cystic Fibrosis Disease-Specific iPSCs.

Stem Cell Reports 2019 06 9;12(6):1389-1403. Epub 2019 May 9.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany; REBIRTH-Cluster of Excellence, Hannover Medical School, 30625 Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany. Electronic address:

Organotypic culture systems from disease-specific induced pluripotent stem cells (iPSCs) exhibit obvious advantages compared with immortalized cell lines and primary cell cultures, but implementation of iPSC-based high-throughput (HT) assays is still technically challenging. Here, we demonstrate the development and conduction of an organotypic HT Cl/I exchange assay using cystic fibrosis (CF) disease-specific iPSCs. The introduction of a halide-sensitive YFP variant enabled automated quantitative measurement of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) function in iPSC-derived intestinal epithelia. CFTR function was partially rescued by treatment with VX-770 and VX-809, and seamless gene correction of the p.Phe508del mutation resulted in full restoration of CFTR function. The identification of a series of validated primary hits that improve the function of p.Phe508del CFTR from a library of ∼42,500 chemical compounds demonstrates that the advantages of complex iPSC-derived culture systems for disease modeling can also be utilized for drug screening in a true HT format.
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http://dx.doi.org/10.1016/j.stemcr.2019.04.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6565754PMC
June 2019

GMP-compatible manufacturing of three iPS cell lines from human peripheral blood.

Stem Cell Res 2019 03 11;35:101394. Epub 2019 Feb 11.

Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), REBIRTH Cluster of Excellence, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany.

The utilization of human induced pluripotent stem cells (hiPSCs) for disease modeling and drug discovery is already reality, and several first-in-man-applications as cellular therapeutics have been initiated. Implementation of good manufacturing practice (GMP)-compliant protocols for the generation of hiPSC lines is crucial to increase the application safety as well as to fulfil the legal requirements for clinical trials approval. Here we describe the development of a GMP-compatible protocol for the reprogramming of CD34 hematopoietic stem cells from peripheral blood (CD34 PBHSC) into hiPSCs using Sendai virus-based reprogramming vectors. Three GMP-compatible hiPSC (GMP-hiPSC) lines were manufactured and characterized under these conditions.
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http://dx.doi.org/10.1016/j.scr.2019.101394DOI Listing
March 2019

Author Correction: Reprogramming triggers endogenous L1 and Alu retrotransposition in human induced pluripotent stem cells.

Nat Commun 2018 12 19;9(1):5398. Epub 2018 Dec 19.

Division of Medical Biotechnology, Paul-Ehrlich-Institute, D-63225, Langen, Germany.

This Article contains an error in the author affiliations. The correct affiliation for author Ruchi Shukla is 'MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Crewe Road, Edinburgh, EH4 2XU, UK', and is not 'Mater Research Institute - University of Queensland, TRI Building, Woolloongabba QLD 4102, Australia'.
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http://dx.doi.org/10.1038/s41467-018-07917-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6300592PMC
December 2018

Bioreactor-based mass production of human iPSC-derived macrophages enables immunotherapies against bacterial airway infections.

Nat Commun 2018 11 30;9(1):5088. Epub 2018 Nov 30.

JRG Translational Hematology of Congenital Diseases, Hannover Medical School, Hannover, Germany.

The increasing number of severe infections with multi-drug-resistant pathogens worldwide highlights the need for alternative treatment options. Given the pivotal role of phagocytes and especially alveolar macrophages in pulmonary immunity, we introduce a new, cell-based treatment strategy to target bacterial airway infections. Here we show that the mass production of therapeutic phagocytes from induced pluripotent stem cells (iPSC) in industry-compatible, stirred-tank bioreactors is feasible. Bioreactor-derived iPSC-macrophages (iPSC-Mac) represent a highly pure population of CD45CD11bCD14CD163 cells, and share important phenotypic, functional and transcriptional hallmarks with professional phagocytes, however with a distinct transcriptome signature similar to primitive macrophages. Most importantly, bioreactor-derived iPSC-Mac rescue mice from Pseudomonas aeruginosa-mediated acute infections of the lower respiratory tract within 4-8 h post intra-pulmonary transplantation and reduce bacterial load. Generation of specific immune-cells from iPSC-sources in scalable stirred-tank bioreactors can extend the field of immunotherapy towards bacterial infections, and may allow for further innovative cell-based treatment strategies.
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http://dx.doi.org/10.1038/s41467-018-07570-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6269475PMC
November 2018

Corrigendum to "Anti-androgenic therapy with finasteride improves cardiac function, attenuates remodeling and reverts pathologic gene-expression after myocardial infarction in mice" Journal of Molecular and Cellular Cardiology 122 (2018) 114-124.

J Mol Cell Cardiol 2018 Dec 7;125:205. Epub 2018 Nov 7.

Klinik für Kardiologie und Angiologie, Klinik für Herz-, Thorax-, Transplantations- und Gefäßchirurgie, Medizinische Hochschule Hannover, 30625 Hannover, Germany; Abteilung für Herz- und Kreislaufforschung, European Center for Angioscience (ECAS), Medizinische Fakultät Mannheim der Universität Heidelberg, 68167 Mannheim,Germany. Electronic address:

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http://dx.doi.org/10.1016/j.yjmcc.2018.11.003DOI Listing
December 2018