Publications by authors named "Jürgen Hescheler"

327 Publications

Impact of Poly(dimethylsiloxane) Surface Modification with Conventional and Amino Acid-Conjugated Self-Assembled Monolayers on the Differentiation of Induced Pluripotent Stem Cells into Cardiomyocytes.

ACS Biomater Sci Eng 2021 04 16;7(4):1539-1551. Epub 2021 Mar 16.

Institute of Biomedical Engineering, Boğaziçi University, Kandilli Campus, Rasathane Cd. Uskudar, 34684 Istanbul, Turkey.

Cardiomyocytes, differentiated from induced pluripotent stem cells (iPSCs), have the potential to produce patient- and disease-specific pharmacological and toxicological platforms, in addition to their cardiac cell therapy applications. However, the lack of both a robust and a simple procedure for scalable cell substrate production is one of the major limitations in this area. Mimicking the natural healthy myocardium extracellular matrix (ECM) properties by altering the cell substrate properties, such as stiffness and chemical/biochemical composition, can significantly affect cell substrate interfacial characteristics and potentially influence cellular behavior and differentiation of iPSCs to cardiomyocytes. Here, we propose a systematic and biomimetic approach, based on the preparation of poly(dimethylsiloxane) (PDMS) substrates having the similar stiffness as healthy heart tissue and a well-defined surface chemistry obtained by conventional [(3-aminopropyl)triethoxysilane (APTES) and octadecyltrimethoxysilane (OTS)] and amino acid (histidine and leucine)-conjugated self-assembled monolayers (SAMs). Among a wide range of different concentrations, the 50:1 prepolymer cross-linker ratio of PDMS allowed adaptation of the myocardium stiffness with a Young's modulus of 23.79 ± 0.61 kPa. Compared with conventional SAM modification, amino acid-conjugated SAMs greatly improved iPSC adhesion, viability, and cardiac marker expression by increasing surface biomimetic properties, whereas all SAMs enhanced cell behavior, with respect to native PDMS. Furthermore, leucine-conjugated SAM modification provided the best environment for cardiac differentiation of iPSCs. This optimized approach can be easily adapted for cardiac differentiation of iPSCs , rendering a very promising tool for microfluidics, drug screening, and organ-on-chip platforms.
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http://dx.doi.org/10.1021/acsbiomaterials.0c01434DOI Listing
April 2021

Pharmacological Neuroenhancement: Current Aspects of Categorization, Epidemiology, Pharmacology, Drug Development, Ethics, and Future Perspectives.

Neural Plast 2021 13;2021:8823383. Epub 2021 Jan 13.

Experimental Neuropsychopharmacology, Federal Institute for Drugs and Medical Devices (Bundesinstitut für Arzneimittel und Medizinprodukte, BfArM), Kurt-Georg-Kiesinger-Allee 3, 53175 Bonn, Germany.

Recent pharmacoepidemiologic studies suggest that pharmacological neuroenhancement (pNE) and mood enhancement are globally expanding phenomena with distinctly different regional characteristics. Sociocultural and regulatory aspects, as well as health policies, play a central role in addition to medical care and prescription practices. The users mainly display self-involved motivations related to cognitive enhancement, emotional stability, and adaptivity. Natural stimulants, as well as drugs, represent substance abuse groups. The latter comprise purines, methylxanthines, phenylethylamines, modafinil, nootropics, antidepressants but also benzodiazepines, -adrenoceptor antagonists, and cannabis. Predominant pharmacodynamic target structures of these substances are the noradrenergic/dopaminergic and cholinergic receptor/transporter systems. Further targets comprise adenosine, serotonin, and glutamate receptors. Meta-analyses of randomized-controlled studies in healthy individuals show no or very limited verifiability of positive effects of pNE on attention, vigilance, learning, and memory. Only some members of the substance abuse groups, i.e., phenylethylamines and modafinil, display positive effects on attention and vigilance that are comparable to caffeinated drinks. However, the development of new antidementia drugs will increase the availability and the potential abuse of pNE. Social education, restrictive regulatory measures, and consistent medical prescription practices are essential to restrict the phenomenon of neuroenhancement with its social, medical, and ethical implications. This review provides a comprehensive overview of the highly dynamic field of pharmacological neuroenhancement and elaborates the dramatic challenges for the medical, sociocultural, and ethical fundaments of society.
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http://dx.doi.org/10.1155/2021/8823383DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7817276PMC
January 2021

Inflammatory Pre-Conditioning of Adipose-Derived Stem Cells with Cerebrospinal Fluid from Traumatic Brain Injury Patients Alters the Immunomodulatory Potential of ADSC Secretomes.

J Neurotrauma 2021 Mar 4. Epub 2021 Mar 4.

Department of Neurosurgery, Medical University Graz, Graz, Austria.

Immunomodulation by adipose-tissue-derived stem cells (ADSCs) is of special interest for the alleviation of damaging inflammatory responses in central nervous system injuries. The present study explored the effects of cerebrospinal fluid (CSF) from traumatic brain injury (TBI) patients on this immunomodulatory potential of ADSCs. CSF conditioning of ADSCs increased messenger RNA levels of both pro- and anti-inflammatory genes compared to controls. Exposure of phorbol-12-myristate-13-acetate-differentiated THP1 macrophages to the secretome of CSF-conditioned ADSCs downregulated both proinflammatory (cyclooxygenase-2, tumor necrosis factor alpha) and anti-inflammatory (suppressor of cytokine signaling 3, interleukin-1 receptor antagonist, and transforming growth factor beta) genes in these cells. Interleukin-10 expression was elevated in both naïve and conditioned secretomes. ADSC secretome treatment, further, induced macrophage maturation of THP1 cells and increased the percentage of CD11b, CD14, CD86, and, to a lesser extent, CD206 cells. This, moreover, enhanced the phagocytic activity of CD14 and CD86 cells, though independently of pre-conditioning. Secretome exposure, finally, also induced a reduction in the percentage of CD192 adherent cells in cultures of peripheral blood mononuclear cells (PBMCs) from both healthy subjects and TBI patients. This limited efficacy (of both naïve and pre-conditioned secretomes) suggests that the effects of lymphocyte-monocyte paracrine signaling on the fate of cultured PBMCs are strongest upon adherent cell populations.
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http://dx.doi.org/10.1089/neu.2020.7017DOI Listing
March 2021

Salicylic diamines selectively eliminate residual undifferentiated cells from pluripotent stem cell-derived cardiomyocyte preparations.

Sci Rep 2021 Jan 27;11(1):2391. Epub 2021 Jan 27.

Center for Physiology and Pathophysiology, Institute for Neurophysiology, Medical Faculty, University of Cologne, Cologne, Germany.

Clinical translation of pluripotent stem cell (PSC) derivatives is hindered by the tumorigenic risk from residual undifferentiated cells. Here, we identified salicylic diamines as potent agents exhibiting toxicity to murine and human PSCs but not to cardiomyocytes (CMs) derived from them. Half maximal inhibitory concentrations (IC) of small molecules SM2 and SM6 were, respectively, 9- and 18-fold higher for human than murine PSCs, while the IC of SM8 was comparable for both PSC groups. Treatment of murine embryoid bodies in suspension differentiation cultures with the most effective small molecule SM6 significantly reduced PSC and non-PSC contamination and enriched CM populations that would otherwise be eliminated in genetic selection approaches. All tested salicylic diamines exerted their toxicity by inhibiting the oxygen consumption rate (OCR) in PSCs. No or only minimal and reversible effects on OCR, sarcomeric integrity, DNA stability, apoptosis rate, ROS levels or beating frequency were observed in PSC-CMs, although effects on human PSC-CMs seemed to be more deleterious at higher SM-concentrations. Teratoma formation from SM6-treated murine PSC-CMs was abolished or delayed compared to untreated cells. We conclude that salicylic diamines represent promising compounds for PSC removal and enrichment of CMs without the need for other selection strategies.
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http://dx.doi.org/10.1038/s41598-021-81351-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7841182PMC
January 2021

The influence of melatonin on the heart rhythm - An in vitro simulation with murine embryonic stem cell derived cardiomyocytes.

Biomed Pharmacother 2021 Apr 12;136:111245. Epub 2021 Jan 12.

Institute for Neurophysiology, Faculty of Medicine, University of Cologne, Cologne, Germany; Praxis für Kardiologie und Angiologie, Landsberg am Lech, Germany; Medical Clinic II, University Clinic of Schleswig-Holstein / Campus Luebeck, University of Luebeck, Luebeck, Germany. Electronic address:

Background: In healthy individuals, a major factor influencing the heart rate variability (HRV) is the circadian rhythm. The role of melatonin as an essential component of the circadian rhythm in the adult human organism and the beneficial effects of a treatment with melatonin during the fetal period is well described. Toxic effects of melatonin are discussed less frequently. Since pharmacological studies cannot be carried out on pregnant women, the establishment of an equivalent in vitro model is important. We therefore tested whether melatonin can influence the beat rate variability (BRV) of spontaneously beating cardiomyocytes derived from murine embryonic stem cells (mESCs) and whether melatonin exhibits toxic effects in this in vitro model.

Methods: Microelectrode Arrays recorded extracellular field potentials of spontaneously beating cardiomyocytes. Melatonin was applied in a concentration range from 10 M to 10 M. The analysis of the BRV focused on time domain methods.

Results: In line with clinical observations, melatonin decreased the beating frequency and increased the BRV. The effect of melatonin up to a concentration of 10 M was reversible, whereas the application of higher concentrations induced an irreversible effect.

Conclusion: The study underlines the potential of this in vitro model to help explore the development of circadian rhythms and their modulation by melatonin in the embryonic phase. The results imply that melatonin influences the heart rhythm as early as during the embryonic heart development. Furthermore, the results indicate a potentially toxic effect of melatonin that has not been described in detail before.
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http://dx.doi.org/10.1016/j.biopha.2021.111245DOI Listing
April 2021

Enhanced hippocampal type II theta activity AND altered theta architecture in mice lacking the Ca3.2 T-type voltage-gated calcium channel.

Sci Rep 2021 Jan 13;11(1):1099. Epub 2021 Jan 13.

Experimental Neuropsychopharmacology, Federal Institute for Drugs and Medical Devices (Bundesinstitut für Arzneimittel und Medizinprodukte, BfArM), Kurt-Georg-Kiesinger-Allee 3, 53175, Bonn, Germany.

T-type Ca channels are assumed to contribute to hippocampal theta oscillations. We used implantable video-EEG radiotelemetry and qPCR to unravel the role of Ca3.2 Ca channels in hippocampal theta genesis. Frequency analysis of spontaneous long-term recordings in controls and Ca3.2 mice revealed robust increase in relative power in the theta (4-8 Hz) and theta-alpha (4-12 Hz) ranges, which was most prominent during the inactive stages of the dark cycles. Urethane injection experiments also showed enhanced type II theta activity and altered theta architecture following Ca3.2 ablation. Next, gene candidates from hippocampal transcriptome analysis of control and Ca3.2 mice were evaluated using qPCR. Dynein light chain Tctex-Type 1 (Dynlt1b) was significantly reduced in Ca3.2 mice. Furthermore, a significant reduction of GABA A receptor δ subunits and GABA B1 receptor subunits was observed in the septohippocampal GABAergic system. Our results demonstrate that ablation of Ca3.2 significantly alters type II theta activity and theta architecture. Transcriptional changes in synaptic transporter proteins and GABA receptors might be functionally linked to the electrophysiological phenotype.
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http://dx.doi.org/10.1038/s41598-020-79763-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806756PMC
January 2021

Persistence of intramyocardially transplanted murine induced pluripotent stem cell-derived cardiomyocytes from different developmental stages.

Stem Cell Res Ther 2021 Jan 8;12(1):46. Epub 2021 Jan 8.

Department of Internal Medicine III, University Hospital of Cologne, Cologne, Germany.

Background: Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) are regarded as promising cell type for cardiac cell replacement therapy, but it is not known whether the developmental stage influences their persistence and functional integration in the host tissue, which are crucial for a long-term therapeutic benefit. To investigate this, we first tested the cell adhesion capability of murine iPSC-CM in vitro at three different time points during the differentiation process and then examined cell persistence and quality of electrical integration in the infarcted myocardium in vivo.

Methods: To test cell adhesion capabilities in vitro, iPSC-CM were seeded on fibronectin-coated cell culture dishes and decellularized ventricular extracellular matrix (ECM) scaffolds. After fixed periods of time, stably attached cells were quantified. For in vivo experiments, murine iPSC-CM expressing enhanced green fluorescent protein was injected into infarcted hearts of adult mice. After 6-7 days, viable ventricular tissue slices were prepared to enable action potential (AP) recordings in transplanted iPSC-CM and surrounding host cardiomyocytes. Afterwards, slices were lysed, and genomic DNA was prepared, which was then used for quantitative real-time PCR to evaluate grafted iPSC-CM count.

Results: The in vitro results indicated differences in cell adhesion capabilities between day 14, day 16, and day 18 iPSC-CM with day 14 iPSC-CM showing the largest number of attached cells on ECM scaffolds. After intramyocardial injection, day 14 iPSC-CM showed a significant higher cell count compared to day 16 iPSC-CM. AP measurements revealed no significant difference in the quality of electrical integration and only minor differences in AP properties between d14 and d16 iPSC-CM.

Conclusion: The results of the present study demonstrate that the developmental stage at the time of transplantation is crucial for the persistence of transplanted iPSC-CM. iPSC-CM at day 14 of differentiation showed the highest persistence after transplantation in vivo, which may be explained by a higher capability to adhere to the extracellular matrix.
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http://dx.doi.org/10.1186/s13287-020-02089-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7792075PMC
January 2021

Generation of transgene-free induced pluripotent stem cells from cardiac fibroblasts of goat embryos.

J Stem Cells Regen Med 2020 11;16(2):34-43. Epub 2020 Dec 11.

Institute of Neurophysiology, University of Cologne, Robert-Koch-Strasse 39, 50931 Cologne, Germany.

Induced pluripotent stem cells (iPSCs) hold a great potential for therapeutic regenerative medicine. The aim of this study was to generate induced pluripotent stem cells from goat embryonic cardiac tissue derived fibroblasts. The isolated cardiac fibroblasts from the cardiac tissue of goat embryos were positive for alfa smooth muscle actin, vimentin and discoidin domain receptor2. From these cells, we generated transgene free iPSCs using piggyBac transposons / transposase using five transcription factors (Oct4, Sox2, Klf, Myc and Lin 28). The generated iPSCs were SSEA1, SSEA4 and Oct4 positive. They were cultured on neofeeders using 20% Serum replacement - IMDM with bFGF. They could form cystic and compact embryoid bodies that showed differentiated ectodermal and mesodermal like cells when cultured using 20% FBS-IMDM without bFGF. The iPSCs, generated in the frame of this approach were produced without the use of integrating virus and the reprogramming transgenes were removed at the end of the process. Though there were limitations in the approach used, a substantial sign of reprogramming was obtained.
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http://dx.doi.org/10.46582/jsrm.1602007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7772807PMC
December 2020

Non-Mendelian inheritance during inbreeding of Ca3.2 and Ca2.3 deficient mice.

Sci Rep 2020 10 2;10(1):15993. Epub 2020 Oct 2.

Institute for Neurophysiology, University of Cologne, Robert-Koch-Str. 39, 50931, Cologne, Germany.

The mating of 77 heterozygous pairs (Ca3.2[+|-] x Ca3.2[+|-]) revealed a significant deviation of genotype distribution from Mendelian inheritance in weaned pups. The mating of 14 pairs (Ca3.2[-|-] female x Ca3.2[+|-] male) and 8 pairs (Ca3.2[+|-] female x Ca3.2[-|-] male) confirmed the significant reduction of deficient homozygous Ca3.2[-|-] pups, leading to the conclusion that prenatal lethality may occur, when one or both alleles, encoding the Ca3.2T-type Ca channel, are missing. Also, the mating of 63 heterozygous pairs (Ca2.3[+|-] x Ca2.3[+|-]) revealed a significant deviation of genotype distribution from Mendelian inheritance in weaned pups, but only for heterozygous male mice, leading to the conclusion that compensation may only occur for Ca2.3[-|-] male mice lacking both alleles of the R-type Ca channel. During the mating of heterozygous parents, the number of female mice within the weaned population does not deviate from the expected Mendelian inheritance. During prenatal development, both, T- and R-type Ca currents are higher expressed in some tissues than postnatally. It will be discussed that the function of voltage-gated Ca channels during prenatal development must be investigated in more detail, not least to understand devastative diseases like developmental epileptic encephalopathies (DEE).
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http://dx.doi.org/10.1038/s41598-020-72912-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7532468PMC
October 2020

Correction: Nemade, H.; et al. Cyclooxygenases Inhibitors Efficiently Induce Cardiomyogenesis in Human Pluripotent Stem Cells. 2020, , 554.

Cells 2020 Sep 24;9(10). Epub 2020 Sep 24.

Institute of Neurophysiology, Faculty of Medicine, University of Cologne, Robert-Koch-Str. 39, 50931 Cologne, Germany.

The authors wish to make the following corrections to this paper [...].
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http://dx.doi.org/10.3390/cells9102165DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7599753PMC
September 2020

Stem cells in natural product and medicinal plant drug discovery-An overview of new screening approaches.

Biomed Pharmacother 2020 Nov 10;131:110730. Epub 2020 Sep 10.

Institute of Neurophysiology, University of Cologne, 50931, Cologne, Germany. Electronic address:

Natural products remain a rich source of new drugs, and the search for bioactive molecules from nature continues to play an important role in the development of new medicines. Also, there is increasing use of herbal medicines for the treatment of a plethora of diseases, and demands for more scientific evidence for their efficacy and safety remains a huge challenge. The propensity of stem cells to differentiate into almost every cell type not only holds promise for the delivery of cell-based therapies for currently incurable diseases or a useful tool in studying cell physiology and pathophysiology. Increasingly, stem cells are becoming an important tool in preclinical drug screening and toxicity testing. In this review, we examine the scientific advances made towards the use of pluripotent stem cells as a model for the screening of plant-based medicines. The combination of well-established in vitro electrophysiological and a plethora of toxicogenomic technologies, together with the optimisation of culture methods of herbal plants and pluripotent stem cells can be explored to establish the basis for efficacy, and tissue/organ-based toxicities of many currently used medicinal plants whose efficacies and toxicities remain unknown.
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http://dx.doi.org/10.1016/j.biopha.2020.110730DOI Listing
November 2020

Acquisition of chromosome 1q duplication in parental and genome-edited human-induced pluripotent stem cell-derived neural stem cells results in their higher proliferation rate in vitro and in vivo.

Cell Prolif 2020 Oct 12;53(10):e12892. Epub 2020 Sep 12.

Center for Physiology and Pathophysiology, Institute for Neurophysiology, Medical Faculty, University of Cologne, Cologne, Germany.

Objectives: Genetic engineering of human-induced pluripotent stem cell-derived neural stem cells (hiPSC-NSC) may increase the risk of genomic aberrations. Therefore, we asked whether genetic modification of hiPSC-NSCs exacerbates chromosomal abnormalities that may occur during passaging and whether they may cause any functional perturbations in NSCs in vitro and in vivo.

Materials And Methods: The transgenic cassette was inserted into the AAVS1 locus, and the genetic integrity of zinc-finger nuclease (ZFN)-modified hiPSC-NSCs was assessed by the SNP-based karyotyping. The hiPSC-NSC proliferation was assessed in vitro by the EdU incorporation assay and in vivo by staining of brain slices with Ki-67 antibody at 2 and 8 weeks after transplantation of ZFN-NSCs with and without chromosomal aberration into the striatum of immunodeficient rats.

Results: During early passages, no chromosomal abnormalities were detected in unmodified or ZFN-modified hiPSC-NSCs. However, at higher passages both cell populations acquired duplication of the entire long arm of chromosome 1, dup(1)q. ZNF-NSCs carrying dup(1)q exhibited higher proliferation rate than karyotypically intact cells, which was partly mediated by increased expression of AKT3 located on Chr1q. Compared to karyotypically normal ZNF-NSCs, cells with dup(1)q also exhibited increased proliferation in vivo 2 weeks, but not 2 months, after transplantation.

Conclusions: These results demonstrate that, independently of ZFN-editing, hiPSC-NSCs have a propensity for acquiring dup(1)q and this aberration results in increased proliferation which might compromise downstream hiPSC-NSC applications.
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http://dx.doi.org/10.1111/cpr.12892DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7574866PMC
October 2020

Following the adverse outcome pathway from micronucleus to cancer using H2B-eGFP transgenic healthy stem cells.

Arch Toxicol 2020 09 22;94(9):3265-3280. Epub 2020 Jul 22.

GOBIO GmbH, Institute for Ecology of Waters and Applied Biology, Scheidertalstraße 69a, 65326, Aarbergen, Hesse, Germany.

In vitro assessment of genotoxicity as an early warning tool for carcinogenicity mainly relies on recording cytogenetic damages (micronuclei, nucleoplasmic bridges) in tumour-derived mammalian cell lines like V79 or CHO. The forecasting power of the corresponding standardised test is based on epidemiological evidence between micronuclei frequencies and cancer incidence. As an alternative to destructive staining of nuclear structures a fish stem cell line transgenic for a fusion protein of histone 2B (H2B) and enhanced green fluorescent protein (eGFP) was established. The cells are derived from koi carp brain (KCB) and distinguish from mammalian culturable cells by non-tumour-driven self-renewal. This technology enables the analysis of genotoxic- and malign downstream effects in situ in a combined approach. In proof-of concept-experiments, we used known carcinogens (4-Nitroquinoline 1-oxide, colchicine, diethylstilbestrol, ethyl methanesulfonate) and observed a significant increase in micronuclei (MNi) frequencies in a dose-dependent manner. The concentration ranges for MNi induction were comparable to human/mammalian cells (i.e. VH-16, CHL and HepG2). Cannabidiol caused the same specific cytogenetic damage pattern as observed in human cells, in particular nucleoplasmic bridges. Metabolic activation of aflatoxin B1 and cyclophosphamide could be demonstrated by pre-incubation of the test compounds using either conventional rat derived S9 mix as well as an in vitro generated biotechnological alternative product ewoS9R. The presented high throughput live H2B-eGFP imaging technology using non-transformed stem cells opens new perspectives in the field of in vitro toxicology. The technology offers experimental access to investigate the effects of carcinogens on cell cycle control, gene expression pattern and motility in the course of malign transformation. The new technology enables the definition of Adverse Outcome Pathways leading to malign cell transformation and contributes to the replacement of animal testing. Summary: Complementation of genotoxicity testing by addressing initiating events leading to malign transformation is suggested. A vertebrate cell model showing "healthy" stemness is recommended, in contrast to malign transformed cells used in toxicology/oncocology.
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http://dx.doi.org/10.1007/s00204-020-02821-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415759PMC
September 2020

Radiation Response of Murine Embryonic Stem Cells.

Cells 2020 07 9;9(7). Epub 2020 Jul 9.

Institute of Neurophysiology and Center for Molecular Medicine Cologne (CMMC), University of Cologne, Robert-Koch-Str. 39, 50931 Cologne, Germany.

To understand the mechanisms of disturbed differentiation and development by radiation, murine CGR8 embryonic stem cells (mESCs) were exposed to ionizing radiation and differentiated by forming embryoid bodies (EBs). The colony forming ability test was applied for survival and the MTT test for viability determination after X-irradiation. Cell cycle progression was determined by flow cytometry of propidium iodide-stained cells, and DNA double strand break (DSB) induction and repair by γH2AX immunofluorescence. The radiosensitivity of mESCs was slightly higher compared to the murine osteoblast cell line OCT-1. The viability 72 h after X-irradiation decreased dose-dependently and was higher in the presence of leukemia inhibitory factor (LIF). Cells exposed to 2 or 7 Gy underwent a transient G2 arrest. X-irradiation induced γH2AX foci and they disappeared within 72 h. After 72 h of X-ray exposure, RNA was isolated and analyzed using genome-wide microarrays. The gene expression analysis revealed amongst others a regulation of developmental genes ( and ), downregulation of genes involved in glycolysis and pyruvate metabolism whereas upregulation of genes related to the p53 signaling pathway. X-irradiated mESCs formed EBs and differentiated toward cardiomyocytes but their beating frequencies were lower compared to EBs from unirradiated cells. These results suggest that X-irradiation of mESCs deregulate genes related to the developmental process. The most significant biological processes found to be altered by X-irradiation in mESCs were the development of cardiovascular, nervous, circulatory and renal system. These results may explain the X-irradiation induced-embryonic lethality and malformations observed in animal studies.
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http://dx.doi.org/10.3390/cells9071650DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7408589PMC
July 2020

Zn2+-induced changes in Cav2.3 channel function: An electrophysiological and modeling study.

J Gen Physiol 2020 09;152(9)

University of Cologne, Faculty of Medicine and University Hospital Cologne, Institute for Neurophysiology, Cologne, Germany.

Loosely bound Zn2+ ions are increasingly recognized as potential modulators of synaptic plasticity and neuronal excitability under normal and pathophysiological conditions. Cav2.3 voltage-gated Ca2+ channels are among the most sensitive targets of Zn2+ and are therefore likely to be involved in the neuromodulatory actions of endogenous Zn2+. Although histidine residues on the external side of domain I have been implicated in the effects on Cav2.3 channel gating, the exact mechanisms involved in channel modulation remain incompletely understood. Here, we use a combination of electrophysiological recordings, modification of histidine residues, and computational modeling to analyze Zn2+-induced changes in Cav2.3 channel function. Our most important findings are that multiple high- and low-affinity mechanisms contribute to the net Zn2+ action, that Zn2+ can either inhibit or stimulate Ca2+ influx through Cav2.3 channels depending on resting membrane potential, and that Zn2+ effects may persist for some time even after cessation of the Zn2+ signal. Computer simulations show that (1) most salient features of Cav2.3 channel gating in the absence of trace metals can be reproduced by an obligatory model in which activation of two voltage sensors is necessary to open the pore; and (2) most, but not all, of the effects of Zn2+ can be accounted for by assuming that Zn2+ binding to a first site is associated with an electrostatic modification and mechanical slowing of one of the voltage sensors, whereas Zn2+ binding to a second, lower-affinity site blocks the channel and modifies the opening and closing transitions. While still far from complete, our model provides a first quantitative framework for understanding Zn2+ effects on Cav2.3 channel function and a step toward the application of computational approaches for predicting the complex actions of Zn2+ on neuronal excitability.
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http://dx.doi.org/10.1085/jgp.202012585DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7478874PMC
September 2020

Submicromolar copper (II) ions stimulate transretinal signaling in the isolated retina from wild type but not from Ca2.3-deficient mice.

BMC Ophthalmol 2020 May 6;20(1):182. Epub 2020 May 6.

Institute for Neurophysiology, University of Cologne, Robert-Koch Str. 39, D-50931, Cologne, Germany.

Background: So far, only indirect evidence exists for the pharmacoresistant R-type voltage-gated Ca channel (VGCC) to be involved in transretinal signaling by triggering GABA-release onto ON-bipolar neurons. This release of inhibitory neurotransmitters was deduced from the sensitivity of the b-wave to stimulation by Ni, Zn and Cu. To further confirm the interpretation of these findings, we compared the effects of Cu application and chelation (using kainic acid, KA) on the neural retina from wildtype and Ca2.3-deficient mice. Furthermore, the immediately effect of KA on the ERG b-wave modulation was assessed.

Methods: Transretinal signaling was recorded as an ERG from the superfused murine retina isolated from wildtype and Ca2.3-deficient mice.

Results: In mice, the stimulating effect of 100 nM CuCl is absent in the retinae from Ca2.3-deficient mice, but prominent in Ca2.3-competent mice. Application of up to 3 mM tricine does not affect the murine b-wave in both genotypes, most likely because of chelating amino acids present in the murine nutrient solution. Application of 27 μM KA significantly increased the b-wave amplitude in wild type and Ca2.3 (-|-) mice. This effect can most likely be explained by the stimulation of endogenous KA-receptors described in horizontal, OFF-bipolar, amacrine or ganglion cells, which could not be fully blocked in the present study.

Conclusion: Cu-dependent modulation of transretinal signaling only occurs in the murine retina from Ca2.3 competent mice, supporting the ideas derived from previous work in the bovine retina that R-type Ca channels are involved in shaping transretinal responses during light perception.
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http://dx.doi.org/10.1186/s12886-020-01451-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7201970PMC
May 2020

Epac-mediated relaxation in murine basilar arteries depends on membrane permeability of cyclic nucleotide analogues and endothelial aging.

Gen Physiol Biophys 2020 Mar;39(2):157-168

Institute of Vegetative Physiology, Center of Physiology, University of Cologne, Cologne, Germany.

Cerebral blood supply is finely tuned by regulatory mechanisms depending on vessel caliber the disruption of which contributes to the development of diseases such as vascular dementia, Alzheimer's and Parkinson 's diseases. This study scopes whether cAMP-mimetic-ligands relax young and aged murine cerebral arteries, whether this relates to the activation of PKA or Epac signaling pathways and is changed with advanced age. The hormone Urocortin-1 relaxed submaximally contracted young and old basilar arteries with a similar pD2 and DMAX (~ -8.5 and ~ 90% in both groups). In permeabilized arteries, PKA activation by 6-Bnz-cAMP or Epac activation by 8-pCPT-2'- O-Me-cAMP also induced relaxation with pD2 of -6.3 vs. -5.8 in old for PKA-ligands, and -4.4 and -4.0 in old for Epac-ligands. Furthermore, aging significantly increased submaximal Ca2+-induced force. The effect of 8-pCPT-2'-O-Me-cAMP on intact arteries was attenuated by aging or nitric oxide synthase inhibition. No relaxing effect in both age-groups was observed after treatment with PKAactivator, Sp-6-Phe-cAMPS. In conclusion, our results suggest that in intact basilar arteries relaxation induced by cAMP-mimetics refers only to the activation of Epac and is impaired by smooth muscle and endothelial aging. The study presents an interesting option allowing therapeutic discrimination between both pathways, possibly for the exclusive activation of Epac in brain circulatory system.
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http://dx.doi.org/10.4149/gpb_2019055DOI Listing
March 2020

Mechano-Pharmacological Testing of L-Type Ca Channel Modulators via Human Vascular Celldrum Model.

Cell Physiol Biochem 2020 Apr;54(3):371-383

Institute for Neurophysiology, University Cologne, Cologne, Germany.

Background/aims: This study aimed to establish a precise and well-defined working model, assessing pharmaceutical effects on vascular smooth muscle cell monolayer in-vitro. It describes various analysis techniques to determine the most suitable to measure the biomechanical impact of vasoactive agents by using CellDrum technology.

Methods: The so-called CellDrum technology was applied to analyse the biomechanical properties of confluent human aorta muscle cells (haSMC) in monolayer. The cell generated tensions deviations in the range of a few N/m² are evaluated by the CellDrum technology. This study focuses on the dilative and contractive effects of L-type Ca channel agonists and antagonists, respectively. We analyzed the effects of Bay K8644, nifedipine and verapamil. Three different measurement modes were developed and applied to determine the most appropriate analysis technique for the study purpose. These three operation modes are called, particular time mode" (PTM), "long term mode" (LTM) and "real-time mode" (RTM).

Results: It was possible to quantify the biomechanical response of haSMCs to the addition of vasoactive agents using CellDrum technology. Due to the supplementation of 100nM Bay K8644, the tension increased approximately 10.6% from initial tension maximum, whereas, the treatment with nifedipine and verapamil caused a significant decrease in cellular tension: 10nM nifedipine decreased the biomechanical stress around 6,5% and 50nM verapamil by 2,8%, compared to the initial tension maximum. Additionally, all tested measurement modes provide similar results while focusing on different analysis parameters.

Conclusion: The CellDrum technology allows highly sensitive biomechanical stress measurements of cultured haSMC monolayers. The mechanical stress responses evoked by the application of vasoactive calcium channel modulators were quantified functionally (N/m²). All tested operation modes resulted in equal findings, whereas each mode features operation-related data analysis.
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http://dx.doi.org/10.33594/000000225DOI Listing
April 2020

Cyclooxygenases Inhibitors Efficiently Induce Cardiomyogenesis in Human Pluripotent Stem Cells.

Cells 2020 02 27;9(3). Epub 2020 Feb 27.

Institute of Neurophysiology, Faculty of Medicine, University of Cologne, Robert-Koch-Str. 39, 50931 Cologne, Germany.

Application of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) is limited by the challenges in their efficient differentiation. Recently, the Wingless (Wnt) signaling pathway has emerged as the key regulator of cardiomyogenesis. In this study, we evaluated the effects of cyclooxygenase inhibitors on cardiac differentiation of hPSCs. Cardiac differentiation was performed by adherent monolayer based method using 4 hPSC lines (HES3, H9, IMR90, and ES4SKIN). The efficiency of cardiac differentiation was evaluated by flow cytometry and RT-qPCR. Generated hPSC-CMs were characterised using immunocytochemistry, electrophysiology, electron microscopy, and calcium transient measurements. Our data show that the COX inhibitors Sulindac and Diclofenac in combination with CHIR99021 (GSK-3 inhibitor) efficiently induce cardiac differentiation of hPSCs. In addition, inhibition of COX using siRNAs targeted towards COX-1 and/or COX-2 showed that inhibition of COX-2 alone or COX-1 and COX-2 in combination induce cardiomyogenesis in hPSCs within 12 days. Using IMR90-Wnt reporter line, we showed that inhibition of COX-2 led to downregulation of Wnt signalling activity in hPSCs. In conclusion, this study demonstrates that COX inhibition efficiently induced cardiogenesis via modulation of COX and Wnt pathway and the generated cardiomyocytes express cardiac-specific structural markers as well as exhibit typical calcium transients and action potentials. These cardiomyocytes also responded to cardiotoxicants and can be relevant as an in vitro cardiotoxicity screening model.
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http://dx.doi.org/10.3390/cells9030554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7140528PMC
February 2020

Experimentally Induced Convulsive Seizures Are Modulated in Part by Zinc Ions through the Pharmacoresistant Ca2.3 Calcium Channel.

Cell Physiol Biochem 2020 Feb;54(2):180-194

Institute for Neurophysiology, University of Cologne, Köln, Germany,

Background/aims: Still in 1999 the first hints were published for the pharmacoresistant Ca2.3 calcium channel to be involved in the generation of epileptic seizures, as transcripts of alpha1E (Ca2.3) and alpha1G (Ca3.1) are changed in the brain of genetic absence epilepsy rats from Strasbourg (GAERS). Consecutively, the seizure susceptibility of mice lacking Ca2.3 was analyzed in great detail by using 4-aminopyridine, pentylene-tetrazol, N-methyl-D-aspartate and kainic acid to induce experimentally convulsive seizures. Further, γ-hydroxybutyrolactone was used for the induction of non-convulsive absence seizures. For all substances tested, Ca2.3-competent mice differed from their knockout counterparts in the sense that for convulsive seizures the deletion of the pharmacoresistant channel was beneficial for the outcome during experimentally induced seizures [1]. The antiepileptic drug lamotrigine reduces seizure activity in Ca2.3-competent but increases it in Ca2.3-deficient mice. In vivo, Ca2.3 must be under tight control by endogenous trace metal cations (Zn and Cu). The dyshomeostasis of either of them, especially of Cu, may alter the regulation of Ca2.3 severely and its activity for Ca conductance, and thus may change hippocampal and neocortical signaling to hypo- or hyperexcitation.

Methods: To investigate by telemetric EEG recordings the mechanism of generating hyperexcitation by kainate, mice were tested for their sensitivity of changes in neuronal (intracerebroventricular) concentrations of the trace metal cation Zn. As the blood-brain barrier limits the distribution of bioavailable Zn or Cu into the brain, we administered micromolar Zn ions intracerebroventricularly in the presence of 1 mM histidine as carrier and compared the effects on behavior and EEG activity in both genotypes.

Results: Kainate seizures are more severe in Ca2.3-competent mice than in KO mice and histidine lessens seizure severity in competent but not in Ca2.3-deficient mice. Surprisingly, Zn plus histidine resembles the kainate only control with more seizure severity in Ca2.3-competent than in deficient mice.

Conclusion: Ca2.3 represents one important Zn-sensitive target, which is useful for modulating convulsive seizures.
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http://dx.doi.org/10.33594/000000213DOI Listing
February 2020

Stem cells for regenerative medicine and anti-aging.

Authors:
Jurgen Hescheler

J Stem Cells Regen Med 2019 24;15(2):53. Epub 2019 Dec 24.

Dept. of Neurophysiology, University of Cologne, Germany.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6971379PMC
December 2019

Evaluation of chemotherapy with nanosomal paclitaxel and gene therapy expressing apoptosis-inducing proteins in the management of spontaneous canine mammary neoplasm.

J Stem Cells Regen Med 2019 24;15(2):24-34. Epub 2019 Dec 24.

Director, Institute of Neurophysiology, Universität zu Köln, Robert-Koch-Strasse 39, D-50931, Köln, Germany.

Mammary gland tumours are the second most common neoplasm representing about 40-50% of all neoplasm after skin tumour, but the majority of these tumours occur in intact/ non spaying female dogs. Surgical excision of the benign tumour is the standard treatment of canine mammary tumours. Chemotherapy is the choice of treatment if the tumour is malignant or shows evidence of invasion into lymph or blood vessels, however, they showed different side effects and their success rate is varied. Taxanes are now the most promising anti-cancer drugs with little side effects. Gene therapy expressing apoptosis-inducing proteins have ability to kill cancer cells while sparing normal cells. The present study was conducted for exploring the oncolytic effect of viral gene therapy expressing apoptosis-inducing proteins construct (ns1 +vp3), nanosomal paclitaxel as chemotherapeutic agent and surgical therapy in the management of spontaneous canine mammary tumours. Chemotherapy (nanosomal paclitaxel) (n=10), viral gene construct (ns1 +vp3) (n=10) and surgical therapy (n=10) were used in 30 female dogs of different breeds having different types of spontaneous mammary tumours. Chemotherapeutic drug and viral gene construct (ns1 +vp3) induced apoptosis in canine mammary neoplasms were studied using fluorescent activated cell sorting analysis. However, apoptotic percentage was significantly higher in chemotherapeutic group than viral gene construct therapy. No major side effects were observed in any groups. Matrix metalloproteinase-2 was found as an important prognostic tool in the management of canine mammary tumours. In conclusion, chemotherapy with nanosomal paclitaxel proved better than viral gene construct (ns1 +vp3) in the treatment of canine mammary neoplasm.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6971383PMC
December 2019

Inflammatory Microenvironment of Acute Myocardial Infarction Prevents Regeneration of Heart with Stem Cells Therapy.

Cell Physiol Biochem 2019 ;53(5):887-909

International Stem Cell Academy, Cologne/Düsseldorf, Germany,

Over the past years, the benefits of stem cell therapy approach for treatment of the cardiovascular diseases have been shown through the rebuilding of new cardiomyocytes and blood vessels. while a successful regeneration of the myocardium has been proven on the animal models of acute myocardial injuries resulted from the stem cells transplantation, no significant long-term regenerative with autologous stem cell therapy in patients with acute myocardial infarction have been reported based on recent meta-analyses. It seems that the inflammatory microenvironment of acute myocardial infarction has an inhibitory effect on the stem cells potential for regenerating the injured myocardium. Secretion of critical cytokines with pro-inflammatory properties including tumor necrosis factor-α, interleukin-1β, and interleukin-6 as well as induction of hypoxic condition and finally formation of cytotoxic elements cause the cellular death and hinder the stem cells proliferation and differentiation. Based on the evidence, application of some approaches like co-delivery of mesenchymal stem cells with the other useful cells, using the stem cells derived productions, administration of preconditioned and modified cells, and also using the anti-inflammatory agents besides the cell therapy are hypothesized as the primary developed safe and practical approaches for decreasing destructive effects of the inflammation on the implanted stem/progenitor cells. In this review, we critically discuss the quiddity of the inflammatory microenvironment and its promoted mechanisms as the main elements to hinder the efficacy of stem cell therapy in the cases of acute myocardial infarction. Also, we finally propose some applied solutions to the problem of cardiac regeneration with stem cells therapy.
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http://dx.doi.org/10.33594/000000180DOI Listing
November 2019

Generation of human induced pluripotent stem cell-derived cardiomyocytes in 2D monolayer and scalable 3D suspension bioreactor cultures with reduced batch-to-batch variations.

Theranostics 2019 25;9(24):7222-7238. Epub 2019 Sep 25.

Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Medical Faculty, Cologne, Germany.

Human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) are promising candidates to treat myocardial infarction and other cardiac diseases. Such treatments require pure cardiomyocytes (CMs) in large quantities. : In the present study we describe an improved protocol for production of hiPSC-CMs in which hiPSCs are first converted into mesodermal cells by stimulation of wingless (Wnt) signaling using CHIR99021, which are then further differentiated into CM progenitors by simultaneous inhibition of porcupine and tankyrase pathways using IWP2 and XAV939 under continuous supplementation of ascorbate during the entire differentiation procedure. : The protocol resulted in reproducible generation of >90% cardiac troponin T (TNNT2)-positive cells containing highly organized sarcomeres. In 2D monolayer cultures CM yields amounted to 0.5 million cells per cm growth area, and on average 72 million cells per 100 mL bioreactor suspension culture without continuous perfusion. The differentiation efficiency was hardly affected by the initial seeding density of undifferentiated hiPSCs. Furthermore, batch-to-batch variations were reduced by combinatorial use of ascorbate, IWP2, and XAV939. : Combined inhibition of porcupine and tankyrase sub-pathways of Wnt signaling and continuous ascorbate supplementation, enable robust and efficient production of hiPSC-CMs.
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http://dx.doi.org/10.7150/thno.32058DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6831300PMC
September 2020

Functional implications of Ca 2.3 R-type voltage-gated calcium channels in the murine auditory system - novel vistas from brainstem-evoked response audiometry.

Eur J Neurosci 2020 04 3;51(7):1583-1604. Epub 2019 Nov 3.

Experimental Neuropsychopharmacology, Federal Institute for Drugs and Medical Devices (Bundesinstitut für Arzneimittel und Medizinprodukte, BfArM), Bonn, Germany.

Voltage-gated Ca channels (VGCCs) are considered to play a key role in auditory perception and information processing within the murine inner ear and brainstem. In the past, Ca 1.3 L-type VGCCs gathered most attention as their ablation causes congenital deafness. However, isolated patch-clamp investigation and localization studies repetitively suggested that Ca 2.3 R-type VGCCs are also expressed in the cochlea and further components of the ascending auditory tract, pointing to a potential functional role of Ca 2.3 in hearing physiology. Thus, we performed auditory profiling of Ca 2.3 controls, heterozygous Ca 2.3 mice and Ca 2.3 null mutants (Ca 2.3 ) using brainstem-evoked response audiometry. Interestingly, click-evoked auditory brainstem responses (ABRs) revealed increased hearing thresholds in Ca 2.3 mice from both genders, whereas no alterations were observed in Ca 2.3 mice. Similar observations were made for tone burst-related ABRs in both genders. However, Ca 2.3 ablation seemed to prevent mutant mice from total hearing loss particularly in the higher frequency range (36-42 kHz). Amplitude growth function analysis revealed, i.a., significant reduction in ABR wave W and W amplitude in mutant animals. In addition, alterations in W -W interwave interval were observed in female Ca 2.3 mice whereas absolute latencies remained unchanged. In summary, our results demonstrate that Ca 2.3 VGCCs are mandatory for physiological auditory information processing in the ascending auditory tract.
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http://dx.doi.org/10.1111/ejn.14591DOI Listing
April 2020

The Effect of Antiarrhythmic Drugs on the Beat Rate Variability of Human Embryonic and Human Induced Pluripotent Stem Cell Derived Cardiomyocytes.

Sci Rep 2019 10 1;9(1):14106. Epub 2019 Oct 1.

Institute for Neurophysiology, University of Cologne, Cologne, Germany.

Embryonic stem cell (ESC) derived tissue is a promising tool to be used in different clinical, preclinical and also scientific settings, for example as in vivo biological pacemaker, preclinical drug safety screening tool or ultimately as part of a cell replacement therapy. However, before ESC derived tissue can be used routinely for these purposes in humans, further studies are needed. In this context, the aims of the present study were to examine the effect of antiarrhythmic drugs on human ESC (hESC) und human induced pluripotent stem cell (hiPSC) derived cardiomyocytes by analyzing the beat rate variability (BRV), which can be considered as the in vitro equivalent of the heart rate variability (HRV) in vivo. Short-term recordings of extracellular field potentials of spontaneously beating cardiomyocytes derived from hESCs and hiPSCs were made using Microelectrode Arrays (MEA). The effect of Flecainide, Ivabradine and Metoprolol was tested. The offline analysis of the BRV was mainly focused on time domain methods. Additionally a non-linear analysis method was used. The evaluation of the Poincaré-Plots of the measurements without pharmacological intervention revealed that the vast majority of the scatter plots have a similar, ellipsoid shape. Flecainide and Ivabradine influenced BRV parameters significantly, whereas Metoprolol did not alter the BRV markedly. We detected remarkable similarities between the BRV of hESC and hiPSC derived cardiomyocytes in vitro and the HRV in vivo. The effect of antiarrhythmic drugs on spontaneously beating cardiomyocytes derived from hESC and hiPSC was generally consistent with clinical experiences and also with our previous study based on murine ESC derived cardiomyocytes. In conclusion, our study points out the great potential of hESC and hiPSC derived tissue to be used routinely for many different applications in medicine and science.
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http://dx.doi.org/10.1038/s41598-019-50557-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6773847PMC
October 2019

Ketamine Increases Proliferation of Human iPSC-Derived Neuronal Progenitor Cells via Insulin-Like Growth Factor 2 and Independent of the NMDA Receptor.

Cells 2019 09 24;8(10). Epub 2019 Sep 24.

Bio-Pharmaceutical Chemistry and Molecular Pharmacology, Faculty of Applied Natural Sciences, Technische Hochschule Köln, 51373 Leverkusen, Germany.

The N-methyl-D-aspartate (NMDA) receptor antagonist ketamine offers promising perspectives for the treatment of major depressive disorder. Although ketamine demonstrates rapid and long-lasting effects, even in treatment-resistant patients, to date, the underlying mode of action remains elusive. Thus, the aim of our study was to investigate the molecular mechanism of ketamine at clinically relevant concentrations by establishing an in vitro model based on human induced pluripotent stem cells (iPSCs)-derived neural progenitor cells (NPCs). Notably, ketamine increased the proliferation of NPCs independent of the NMDA receptor, while transcriptome analysis revealed significant upregulation of insulin-like growth factor 2 (IGF2) and p11, a member of the S100 EF-hand protein family, which are both implicated in the pathophysiology of depression, 24 h after ketamine treatment. Ketamine (1 µM) was able to increase cyclic adenosine monophosphate (cAMP) signaling in NPCs within 15 min and cell proliferation, while ketamine-induced IGF2 expression was reduced after PKA inhibition with cAMPS-Rp. Furthermore, 24 h post-administration of ketamine (15 mg/kg) in vivo confirmed phosphorylation of extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) in the subgranular zone (SGZ) of the hippocampus in C57BL/6 mice. In conclusion, ketamine promotes the proliferation of NPCs presumably by involving cAMP-IGF2 signaling.
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http://dx.doi.org/10.3390/cells8101139DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6830315PMC
September 2019

Complex Clearance Mechanisms After Intraventricular Hemorrhage and rt-PA Treatment-a Review on Clinical Trials.

Transl Stroke Res 2020 06 14;11(3):337-344. Epub 2019 Sep 14.

Institute of Neurophysiology, Medical Faculty, University of Cologne, Cologne, Germany.

Intracerebral hemorrhage in combination with intraventricular hemorrhage (IVH) is a severe type of stroke frequently leading to prolonged clinical care, continuous disability, shunt dependency, and high mortality. The molecular mechanisms induced by IVH are complex and not fully understood. Moreover, the treatment options for IVH are limited. Intraventricular recombinant tissue plasminogen activator (rt-PA) dissolves the blood clot in the ventricular system; however, whether the clinical outcome is thereby positively affected is still being debated. The mechanistic cascade induced by intraventricular rt-PA therapy may cure and harm in parallel. Despite the fact that intraventricular blood clots are thereby dissolved, blood derivatives enter the parenchyma and may still adversely affect functional structures of the brain: Smaller blood clots may obstruct the perivascular (Virchow-Robin) space and thereby the glymphatic system with detrimental consequences for cerebrospinal fluid (CSF)/interstitial fluid (ISF) flow. These clots, blood cells but also blood derivatives in the perivascular space, destabilize the blood-brain barrier from the brain parenchyma side, thereby also functionally weakening the neurovascular unit. This may lead to further accommodation of serum proteins in the ISF and particularly in the perivascular space further contributing to the adverse effects on the neuronal microenvironment. Finally, the arterial (Pacchionian) granulations have to cope with ISF containing this "blood, cell, and protein cocktail," resulting in obstruction and insufficient function of the arterial granulations, followed by a malresorptive hydrocephalus. Particularly in light of currently improved knowledge on the physiologic and pathophysiologic clearance of cerebrospinal fluid and interstitial fluid, a critical discussion and reevaluation of our current therapeutic strategies to treat intraventricular hemorrhages are needed to successfully treat patients suffering from this severe type of stroke. In this review, we therefore summarize and discuss recent clinical trials and future directions for the field of IVH with respect to the currently increased understanding of the glymphatic system and the neurovascular unit pathophysiology.
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http://dx.doi.org/10.1007/s12975-019-00735-6DOI Listing
June 2020

Intracerebroventricular administration of histidine reduces kainic acid-induced convulsive seizures in mice.

Exp Brain Res 2019 Oct 18;237(10):2481-2493. Epub 2019 Jul 18.

Institute for Neurophysiology, University of Cologne, Robert-Koch-Str. 39, 50931, Koeln, Germany.

Kainic acid (KA)-induced seizures and other experimental models of epilepsy have been proven to be instrumental in identifying novel targets that could be responsible for human icto- and epileptogenesis. We have previously shown that the ablation of pharmacoresistant voltage-gated Ca channels with Ca2.3 as central ion-conducting pore (R-type Ca channel) reduces the sensitivity towards KA-induced epilepsy in mice. In vivo, Ca2.3 channels are thought to be under tight allosteric control by endogenous loosely bound trace metal cations (Zn and Cu) that suppress channel gating via a high-affinity trace metal-binding site. Metal dyshomeostasis in the brain, which is a common feature of (KA-induced) seizures, could therefore alter the normal function of Ca2.3 channels and may shift hippocampal and neocortical signaling towards hyperexcitation. To investigate the role of loosely bound metal ions for KA-induced hyperexcitation in vivo, we examined the effects of manipulating brain trace metal homeostasis in mice. To this end, we developed a murine system for intracerebroventricular administration of trace metal ions and/or histidine (His), which can bind Zn and Cu and is involved in their transendothelial transport at the blood-brain barrier. Unexpectedly, our preliminary findings indicate that application of His alone but not in the presence of Zn has substantial beneficial effects on the outcome of KA-induced epilepsy in mice. As such, our results emphasize previous findings on the complex, two-sided role of loosely bound metal ions with regard to neuronal excitation and degeneration under pathophysiological conditions.
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http://dx.doi.org/10.1007/s00221-019-05605-zDOI Listing
October 2019

Application of Stem Cell Technologies to Regenerate Injured Myocardium and Improve Cardiac Function.

Cell Physiol Biochem 2019 ;53(1):101-120

Institute of Neurophysiology, University of Cologne, Cologne, Germany.

In the recent decades, cardiovascular diseases emerged as the major leading cause of human mortality. However, current clinical approaches still do not encompass a thorough therapeutic solution for improving heart function of the patients who suffered an extensive myocardial injury. Based on this status quo, stem cells could become a novel option, as a natural source of the new myocardium lineage cells, being capable of paracrine factors secretion, protection or even regeneration of the damaged heart muscle. Efficient stem cell-based therapy of the heart should lead to repair or/and replacement of the degenerated tissue with functional myocardial and endothelial cells. Hereon, various types of pluripotent and multipotent stem cells have been already studied in the pre-clinical and clinical settings, demonstrating their cardiomyogenic and regenerative potential. In this context, as a type of male adult stem/ progenitors, spermatogonial stem cells feature a remarkable ability for a formation of cardiovascular lineages, based on our own observations. Presented data supports the presumption, that spermatogonial stem cells not only have a suitable capacity to generate functional heart cells but can also potentially improve the function of an injured myocardium. In this review article, we first describe the essential molecular and pathophysiological mechanisms involved in the heart tissue injury. Afterwards, based on our ongoing study, we review the impact of the stem cell technologies on the regeneration therapy in cardiovascular and myocardial diseases. Particular emphasis is being put on the usability of spermatogonial stem cells in cardiac therapy.
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http://dx.doi.org/10.33594/000000124DOI Listing
July 2019