Wybrane publikacje

Single-cell, high-throughput analysis of cell docking to vessel wall.

J Cereb Blood Flow Metab. 2019 Nov;39(11):2308-2320. doi: 10.1177/0271678X18805238. Epub 2018 Oct 26.

Andrzejewska A1, Nowakowski A1, Grygorowicz T2, Dabrowska S1, Orzel J3,4, Walczak P5,6,7, Lukomska B1, Janowski M1,5,6.

Author information

  1. NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
  2. Laboratory of Advanced Microscopy Techniques, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
  3. Department of Experimental Pharmacology and Laboratory of Nuclear Magnetic Resonance Imaging, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
  4. Faculty of Electronics, Warsaw University of Technology, Warsaw, Poland.
  5. Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
  6. Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, Johns Hopkins University, Baltimore, MD, USA.
  7. Department of Neurology and Neurosurgery, Faculty of Medical Sciences, University of Warmia and Mazury, Olsztyn, Poland.

Abstract

Therapeutic potential of mesenchymal stem cells (MSCs) has been reported consistently in animal models of stroke, with mechanism mainly through immunomodulation and paracrine activity. Intravenous injection has been a prevailing route for MSCs administration, but cell quantities needed when scaling-up from mouse to human are extremely high putting into question feasibility of that approach. Intra-arterial delivery directly routes the cells to the brain thus lowering the required dose. Cell engineering may additionally improve cell homing, further potentiating the value of intra-arterial route. Therefore, our goal was to create microfluidic platform for screening and fast selection of molecules that enhance the docking of stem cells to vessel wall. We hypothesized that our software will be capable of detecting distinct docking properties of naïve and ITGA4-engineered MSCs. Indeed, the cell flow tracker analysis revealed positive effect of cell engineering on docking frequency of MSCs (42% vs. 9%, engineered vs. control cells, p < 0.001). These observations were then confirmed in an animal model of focal brain injury where cell engineering resulted in improved homing to the brain. To conclude, we developed a platform to study the docking of cells to the vessel wall which is highly relevant for intraarterial cell targeting or studies on neuroinflammation.

KEYWORDS:

ITGA4; Mesenchymal stem cells; docking; mRNA; microfluidic assay; stroke

Human bone marrow mesenchymal stem cell-derived extracellular vesicles attenuate neuroinflammation evoked by focal brain injury in rats

J Neuroinflammation. 2019; 16: 216.

Published online 2019 Nov 13. doi: 10.1186/s12974-019-1602-5

Dabrowska S1, Andrzejewska A1, Strzemecki D2, Muraca M3, Janowski M1, Lukomska B4.

Author information

  1. NeuroRepair Department, Mossakowski Medical Research Centre, PAS, 5 Pawinskiego Street, 02-106, Warsaw, Poland.
  2. Department of Experimental Pharmacology, Mossakowski Medical Research Centre, PAS, 5 Pawinskiego Street, 02-106, Warsaw, Poland.
  3. Department of Women's and Children's Health, University of Padua, Via Giustiniani 3, 35128, Padua, Italy.
  4. NeuroRepair Department, Mossakowski Medical Research Centre, PAS, 5 Pawinskiego Street, 02-106, Warsaw, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

BACKGROUND: Ischemic stroke is the major cause of long-term severe disability and death in aged population. Cell death in the infarcted region of the brain induces immune reaction leading to further progression of tissue damage. Immunomodulatory function of mesenchymal stem cells (MSCs) has been shown in multiple preclinical studies; however, it has not been successfully translated to a routine clinical practice due to logistical, economical, regulatory, and intellectual property obstacles. It has been recently demonstrated that therapeutic effect of intravenously administered MSCs can be recapitulated by extracellular vesicles (EVs) derived from them. However, in contrast to MSCs, EVs were not capable to decrease stroke-induced neuroinflammation. Therefore, the aim of the study was to investigate if intra-arterial delivery of MSC-derived EVs will have stronger impact on focal brain injury-induced neuroinflammation, which mimics ischemic stroke, and how it compares to MSCs.

METHODS: The studies were performed in adult male Wistar rats with focal brain injury induced by injection of 1 μl of 50 nmol ouabain into the right hemisphere. Two days after brain insult, 5 × 105 human bone marrow MSCs (hBM-MSCs) labeled with Molday ION or 1.3 × 109 EVs stained with PKH26 were intra-arterially injected into the right hemisphere under real-time MRI guidance. At days 1, 3, and 7 post-transplantation, the rats were decapitated, the brains were removed, and the presence of donor cells or EVs was analyzed. The cellular immune response in host brain was evaluated immunohistochemically, and humoral factors were measured by multiplex immunoassay.

RESULTS: hBM-MSCs and EVs transplanted intra-arterially were observed in the rat ipsilateral hemisphere, near the ischemic region. Immunohistochemical analysis of brain tissue showed that injection of hBM-MSCs or EVs leads to the decrease of cell activation by ischemic injury, i.e., astrocytes, microglia, and infiltrating leucocytes, including T cytotoxic cells. Furthermore, we observed significant decrease of pro-inflammatory cytokines and chemokines after hBM-MSC or EV infusion comparing with non-treated rats with focal brain injury.

CONCLUSIONS: Intra-arterially injected EVs attenuated neuroinflammation evoked by focal brain injury, which mimics ischemic stroke, and this effect was comparable to intra-arterial hBM-MSC transplantation. Thus, intra-arterial injection of EVs might be an attractive therapeutic approach, which obviates MSC-related obstacles.

KEYWORDS: Extracellular vesicles; Immune response; Intra-arterial transplantation; Ischemic brain injury; Mesenchymal stem cells; Stroke

Neuroinflammation as a target for treatment of stroke using mesenchymal stem cells and extracellular vesicles.

J Neuroinflammation. 2019 Sep 12;16(1):178. doi: 10.1186/s12974-019-1571-8.

Dabrowska S1, Andrzejewska A1, Lukomska B1, Janowski M2.

 

Author information

  1. NeuroRepair Department, Mossakowski Medical Research Centre, PAS, 5 Pawinskiego Street, 02-106, Warsaw, Poland.
  2. Department of Diagnostic Radiology and Nuclear Medicine, University of Maryland, Baltimore, HSF III, 620 W. Baltimore street, Baltimore, MD, 21201, USA. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

Ischemic stroke is the third cause of death in the developed countries and the main reason of severe disability. Brain ischemia leads to the production of damage-associated molecular patterns (DAMPs) by neurons and glial cells which results in astrocyte and microglia activation, pro-inflammatory cytokines and chemokines production, blood-brain barrier (BBB) disruption, infiltration of leukocytes from the peripheral blood into the infarcted area, and further exacerbation of tissue damage. However, some immune cells such as microglia or monocytes are capable to change their phenotype to anti-inflammatory, produce anti-inflammatory cytokines, and protect injured nervous tissue. In this situation, therapies, which will modulate the immune response after brain ischemia, such as transplantation of mesenchymal stem cells (MSCs) are catching interest. Many experimental studies of ischemic stroke revealed that MSCs are able to modulate immune response and act neuroprotective, through stimulation of neurogenesis, oligodendrogenesis, astrogenesis, and angiogenesis. MSCs may also have an ability to replace injured cells, but the release of paracrine factors directly into the environment or via extracellular vesicles (EVs) seems to play the most pronounced role. EVs are membrane structures containing proteins, lipids, and nucleic acids, and they express similar properties as the cells from which they are derived. However, EVs have lower immunogenicity, do not express the risk of vessel blockage, and have the capacity to cross the blood-brain barrier. Experimental studies of ischemic stroke showed that EVs have immunomodulatory and neuroprotective properties; therefore, they can stimulate neurogenesis and angiogenesis. Up to now, 20 clinical trials with MSC transplantation into patients after stroke were performed, from which two concerned on only hemorrhagic stroke and 13 studied only on ischemic stroke. There is no clinical trial with EV injection into patients after brain ischemia so far, but the case with miR-124-enriched EVs administration is planned and probably there will be more clinical studies with EV transplantation in the near future.

KEYWORDS:

Extracellular vesicles; Ischemia; Mesenchymal stem cells; Neuro-inflammation; Stroke

Mitochondrial damage by α-synuclein causes cell death in human dopaminergic neurons

Cell Death Dis. 2019 Nov 14;10(11):865. doi: 10.1038/s41419-019-2091-2.

Ganjam GK1,2,3, Bolte K4, Matschke LA5, Neitemeier S6, Dolga AM6,7, Höllerhage M8, Höglinger GU8, Adamczyk A9, Decher N5, Oertel WH10,11, Culmsee C6,11,12.


Author information

  1. Institute for Pharmacology and Clinical Pharmacy, Biochemical-Pharmacological Center, University of Marburg, Marburg, Germany. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..
  2. Department of Neurology, University of Marburg, Marburg, Germany. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..
  3. Center for Mind, Brain and Behaviour - CMBB, Marburg, Germany. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..
  4. Laboratory for Cell Biology I, Department of Biology, University of Marburg, Marburg, Germany.
  5. Institute of Physiology and Pathophysiology, University of Marburg, Marburg, Germany.
  6. Institute for Pharmacology and Clinical Pharmacy, Biochemical-Pharmacological Center, University of Marburg, Marburg, Germany.
  7. Department of Molecular Pharmacology, Groningen Research Institute of Pharmacy, University of Groningen, Groningen, The Netherlands.
  8. German Center for Neurodegenerative Diseases (DZNE), Munich, Germany.
  9. Department of Cellular Signaling, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
  10. Department of Neurology, University of Marburg, Marburg, Germany.
  11. Center for Mind, Brain and Behaviour - CMBB, Marburg, Germany.
  12. Department of Pediatrics, The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, China.

Abstract

Evolving concepts on Parkinson's disease (PD) pathology suggest that α-synuclein (aSYN) promote dopaminergic neuron dysfunction and death through accumulating in the mitochondria. However, the consequence of mitochondrial aSYN localisation on mitochondrial structure and bioenergetic functions in neuronal cells are poorly understood. Therefore, we investigated deleterious effects of mitochondria-targeted aSYN in differentiated human dopaminergic neurons in comparison with wild-type (WT) aSYN overexpression and corresponding EGFP (enhanced green fluorescent protein)-expressing controls. Mitochondria-targeted aSYN enhanced mitochondrial reactive oxygen species (ROS) formation, reduced ATP levels and showed severely disrupted structure and function of the dendritic neural network, preceding neuronal death. Transmission electron microscopy illustrated distorted cristae and many fragmented mitochondria in response to WT-aSYN overexpression, and a complete loss of cristae structure and massively swollen mitochondria in neurons expressing mitochondria-targeted aSYN. Further, the analysis of mitochondrial bioenergetics in differentiated dopaminergic neurons, expressing WT or mitochondria-targeted aSYN, elicited a pronounced impairment of mitochondrial respiration. In a pharmacological compound screening, we found that the pan-caspase inhibitors QVD and zVAD-FMK, and a specific caspase-1 inhibitor significantly prevented aSYN-induced cell death. In addition, the caspase inhibitor QVD preserved mitochondrial function and neuronal network activity in the human dopaminergic neurons overexpressing aSYN. Overall, our findings indicated therapeutic effects by caspase-1 inhibition despite aSYN-mediated alterations in mitochondrial morphology and function.


Perineuronal Nets and Their Role in Synaptic Homeostasis

Mateusz Bosiacki 1, Magdalena Ga˛ssowska-Dobrowolska 2, Klaudyna Kojder 3,
Marta Fabianska 4, Dariusz Jezewski 5, Izabela Gutowska 6 and Anna Lubkowska 1,
1 Department of Functional Diagnostics and Physical Medicine, Pomeranian Medical University in Szczecin,Zołnierska 54 Str., 71-210 Szczecin, Poland
2 Department of Cellular Signalling, Mossakowski Medical Research Centre, Polish Academy of Sciences,Pawinskiego 5 Str., 02-106Warsaw, Poland
3 Department of Anaesthesiology and Intensive Care, Pomeranian Medical University in Szczecin,71-252 Szczecin, Poland
4 Institute of Philosophy, University of Szczecin, Krakowska 71-79 Str., 71-017 Szczecin, Poland
5 Department of Neurosurgery and Pediatric Neurosurgery, Department of Applied Neurocognitivistics, Pomeranian Medical University in Szczecin, 71-252 Szczecin, Poland
6 Department of Human Nutrition and Metabolomics, Department of Medical Chemistry, Pomeranian Medical University in Szczecin, Broniewskiego 24 Str., 71-252 Szczecin, Poland

Received: 7 July 2019; Accepted: 16 August 2019; Published: 22 August 2019


Abstract: Extracellular matrix (ECM) molecules that are released by neurons and glial cells form perineuronal nets (PNNs) and modulate many neuronal and glial functions. PNNs, whose structure is still not known in detail, surround cell bodies and dendrites, which leaves free space for synapses to come into contact. A reduction in the expression of many neuronal ECM components adversely a ects processes that are associated with synaptic plasticity, learning, and memory. At the same time, increasedECMactivity, e.g., as a result of astrogliosis following brain damage or in neuroinflammation, can also have harmful consequences. The therapeutic use of enzymes to attenuate elevated neuronal ECM expression after injury or in Alzheimer’s disease has proven to be beneficial by promoting axon growth and increasing synaptic plasticity. Yet, severe impairment of ECM function can also lead to neurodegeneration. Thus, it appears that to ensure healthy neuronal function a delicate balance of ECM components must be maintained. In this paper we review the structure of PNNs and their components, such as hyaluronan, proteoglycans, core proteins, chondroitin sulphate proteoglycans, tenascins, and Hapln proteins. We also characterize the role of ECM in the functioning of the blood-brain barrier, neuronal communication, as well as the participation of PNNs in synaptic plasticity and some clinical aspects of perineuronal net impairment. Furthermore, we discuss the participation of PNNs in brain signaling. Understanding the molecular foundations of the ways that PNNs participate in brain signaling and synaptic plasticity, as well as how they change in
physiological and pathological conditions, may help in the development of new therapies for many degenerative and inflammatory diseases of the brain.

Keywords:perineuronalnets (PNNs); extracellularmatrix(ECM); synaptogenesis; neuronal communication

Directed glial differentiation and transdifferentiation for neural tissue regeneration.

Exp Neurol. 2019 Sep;319:112813. doi: 10.1016/j.expneurol.2018.08.010. Epub 2018 Aug 30.

Janowska J1, Gargas J1, Ziemka-Nalecz M1, Zalewska T1, Buzanska L2, Sypecka J3.

Author information

1 Mossakowski Medical Research Centre, Polish Academy of Sciences, NeuroRepair Department, 5, Pawinskiego str., 02-106 Warsaw, Poland.

2 Mossakowski Medical Research Centre, Polish Academy of Sciences, Stem Cell Bioengineering Unit, 5, Pawinskiego str., 02-106 Warsaw, Poland.

3 Mossakowski Medical Research Centre, Polish Academy of Sciences, NeuroRepair Department, 5, Pawinskiego str., 02-106 Warsaw, Poland. Electronic address: Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

Glial cells which are indispensable for the central nervous system development and functioning, are proven to be vulnerable to a harmful influence of pathological cues and tissue misbalance. However, they are also highly sensitive to both in vitro and in vivo modulation of their commitment, differentiation, activity and even the fate-switch by different types of bioactive molecules. Since glial cells (comprising macroglia and microglia) are an abundant and heterogeneous population of neural cells, which are almost uniformly distributed in the brain and the spinal cord parenchyma, they all create a natural endogenous reservoir of cells for potential neurogenerative processes required to be initiated in response to pathophysiological cues present in the local tissue microenvironment. The past decade of intensive investigation on a spontaneous and enforced conversion of glial fate into either alternative glial (for instance from oligodendrocytes to astrocytes) or neuronal phenotypes, has considerably extended our appreciation of glial involvement in restoring the nervous tissue cytoarchitecture and its proper functions. The most effective modulators of reprogramming processes have been identified and tested in a series of pre-clinical experiments. A list of bioactive compounds which are potent in guiding in vivo cell fate conversion and driving cell differentiation includes a selection of transcription factors, microRNAs, small molecules, exosomes, morphogens and trophic factors, which are helpful in boosting the enforced neuro-or gliogenesis and promoting the subsequent cell maturation into desired phenotypes. Herein, an issue of their utility for a directed glial differentiation and transdifferentiation is discussed in the context of elaborating future therapeutic options aimed at restoring the diseased nervous tissue.

KEYWORDS:

Cell fate conversion; Glial cell reprogramming; Neural development transdifferentiation; Neurorepair; Pre-clinical studies; Therapeutic strategies; Tissue restoration

 

The collagen scaffold supports hiPSC-derived NSC growth and restricts hiPSC.

Front Biosci (Schol Ed). 2019 Mar 1;11:105-121.

Zychowicz M1, Pietrucha K2, Podobinska M1, Kowalska-Wlodarczyk M3, Lenart J4, Augustyniak J1, Buzanska L5.

Author information

1 Stem Cell Bioengineering Unit, Mossakowski Medical Research Centre Polish Academy of Sciences, Pawinskiego 5 St, 02-106, Warsaw, Poland.

2 Department of Material and Commodity Sciences and Textile Metrology, Lodz University of Technology, Zeromskiego 116 St, 90-924, Lodz, Poland.

3 Oil and Gas Institute, National Research Institute, 25 A Lubicz St, 31-503 Cracow, Poland.

4 Department of Neurochemistry, Mossakowski Medical Research Centre Polish Academy of Sciences, Pawinskiego 5 St, 02-106, Warsaw, Poland.

5 Stem Cell Bioengineering Unit, Mossakowski Medical Research Centre Polish Academy of Sciences, Pawinskiego 5 St, 02-106, Warsaw, Poland, Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

 

Abstract

The human induced pluripotent stem cells (hiPSC) are one of the promising candidates as patient specific cell source for autologous transplantation or modeling of diseases. The collagen (Col) scaffolds have been shown suitable to create in vitro biomimetic microenvironment for human neural stem cells, but their ability to accommodate stem cells at different stages of neural differentiation has not been verified yet. In this paper we compare lineage related hiPSC during neural differentiation for their ability to colonize Col scaffold. We have also focused on modification of collagen physicochemical properties with improved mechanical and thermal stability, without loss of its biological activity. The hiPSC expressing markers of pluripotency (OCT4, SOX2, NANOG) after neural commitment are NESTIN, GFAP, PDGFR alpha, beta- TUBULIN III, MAP-2, DCX, GalC positive. We have shown, that Col scaffold was not preferable for hiPSC culture, while the neurally committed population after seeding on Col scaffolds revealed good adhesion, viability, proliferation, along with sustaining markers of neuronal and glial differentiation. The Col scaffold-based 3D culture of hiPSC-NSCs may serve as a research tool for further translational studies.

 

Bezafibrate Upregulates Mitochondrial Biogenesis and Influence Neural Differentiation of Human-Induced Pluripotent Stem Cells.

Mol Neurobiol. 2019 Jun;56(6):4346-4363. doi: 10.1007/s12035-018-1368-2. Epub 2018 Oct 13.

Augustyniak J1, Lenart J2, Gaj P3, Kolanowska M4, Jazdzewski K3,4, Stepien PP5,6,7, Buzanska L8.

Author information

1 Stem Cell Bioengineering Unit, Mossakowski Medical Research Centre Polish Academy of Sciences, Warsaw, Poland.

2 Department of Neurochemistry, Mossakowski Medical Research Centre Polish Academy of Sciences, Warsaw, Poland.

3 Laboratory of Human Cancer Genetics, Centre of New Technologies, University of Warsaw, Warsaw, Poland.

4 Genomic Medicine, Medical University of Warsaw, Warsaw, Poland.

5 Institute of Genetics and Biotechnology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

6 Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland.

7 Centre of New Technologies, University of Warsaw, Warsaw, Poland.

8 Stem Cell Bioengineering Unit, Mossakowski Medical Research Centre Polish Academy of Sciences, Warsaw, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

Bezafibrate (BZ) regulates mitochondrial biogenesis by activation of PPAR's receptors and enhancing the level of PGC-1α coactivator. In this report, we investigated the effect of BZ on the expression of genes (1) that are linked to different pathways involved in mitochondrial biogenesis, e.g., regulated by PPAR's receptors or PGC-1α coactivator, and (2) involved in neuronal or astroglial fate, during neural differentiation of hiPSC. The tested cell populations included hiPSC-derived neural stem cells (NSC), early neural progenitors (eNP), and neural progenitors (NP). RNA-seq analysis showed the expression of PPARA, PPARD receptors and excluded PPARG in all tested populations. The expression of PPARGC1A encoding PGC-1α was dependent on the stage of differentiation: NSC, eNP, and NP differed significantly as compared to hiPSC. In addition, BZ-evoked upregulation of PPARGC1A, GFAP, S100B, and DCX genes coexist with downregulation of MAP2 gene only at the eNP stage of differentiation. In the second task, we investigated the cell sensitivity and mitochondrial biogenesis upon BZ treatment. BZ influenced the cell viability, ROS level, mitochondrial membrane potential, and total cell number in concentration- and stage of differentiation-dependent manner. Induction of mitochondrial biogenesis evoked by BZ determined by the changes in the level of SDHA and COX-1 protein, and mtDNA copy number, as well as the expression of NRF1, PPARGC1A, and TFAM genes, was detected only at NP stage for all tested markers. Thus, developmental stage-specific sensitivity to BZ of neurally differentiating hiPSC can be linked to mitochondrial biogenesis, while fate commitment decisions to PGC-1α (encoded by PPARGC1A) pathway.

KEYWORDS:

Bezafibrate; Mitochondrial biogenesis; NSC; PGC-1α; PPAR’s; hiPSC

 

Species-specific models in toxicology: in vitro epithelial barriers.

Environ Toxicol Pharmacol. 2019 Aug;70:103203. doi: 10.1016/j.etap.2019.103203. Epub 2019 Jun 1.
Bertero A1, Augustyniak J2, Buzanska L2, Caloni F3.
Author information
1 Università degli Studi di Milano, Department of Veterinary Medicine (DIMEVET) Milan, Italy.
2 Stem Cell Bioengineering Unit, Mossakowski Medical Research Centre Polish Academy of Sciences, Warsaw, Poland.
3 Università degli Studi di Milano, Department of Veterinary Medicine (DIMEVET) Milan, Italy. Electronic address: Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..
Abstract
Species-specific in vitro epithelial barriers represent interesting predictive tools for risk assessment evaluation in toxicological studies. Moreover, these models could be applied either as stand-alone methods for the study of absorption, bioavailability, excretion, transport, effects of xenobiotics, or through an Integrated Testing Strategy. The aim of this review is to give a comprehensive overview of in vitro species-specific epithelial barrier models from bovine, dog and swine. Bovine mammary epithelial barrier as a fundamental instrument for the evaluation of the toxicant excretion, the blood brain barrier as a useful first approach in toxicological and pharmacological studies, the porcine intestinal barrier, the canine skin barrier, and finally the pulmonary barrier from bovine and swine species are described in this review.
Copyright © 2019 Elsevier B.V. All rights reserved.
KEYWORDS:
Barriers; Species-specific; Toxicology; in vitro

Species-specific models in toxicology: in vitro epithelial barriers.

Environ Toxicol Pharmacol. 2019 Aug;70:103203. doi: 10.1016/j.etap.2019.103203. Epub 2019 Jun 1.

Bertero A1, Augustyniak J2, Buzanska L2, Caloni F3.

Author information

1        Università degli Studi di Milano, Department of Veterinary Medicine (DIMEVET) Milan, Italy.

2        Stem Cell Bioengineering Unit, Mossakowski Medical Research Centre Polish Academy of Sciences, Warsaw, Poland.

3        Università degli Studi di Milano, Department of Veterinary Medicine (DIMEVET) Milan, Italy. Electronic address: Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

Species-specific in vitro epithelial barriers represent interesting predictive tools for risk assessment evaluation in toxicological studies. Moreover, these models could be applied either as stand-alone methods for the study of absorption, bioavailability, excretion, transport, effects of xenobiotics, or through an Integrated Testing Strategy. The aim of this review is to give a comprehensive overview of in vitro species-specific epithelial barrier models from bovine, dog and swine. Bovine mammary epithelial barrier as a fundamental instrument for the evaluation of the toxicant excretion, the blood brain barrier as a useful first approach in toxicological and pharmacological studies, the porcine intestinal barrier, the canine skin barrier, and finally the pulmonary barrier from bovine and swine species are described in this review.

KEYWORDS:

Barriers; Species-specific; Toxicology; in vitro

 

Organoids are promising tools for species-specific in vitro toxicological studies.

J Appl Toxicol. 2019 Jun 5. doi: 10.1002/jat.3815. [Epub ahead of print]

Augustyniak J1, Bertero A2, Coccini T3, Baderna D4, Buzanska L1, Caloni F2

Author information

1 Department of Stem Cell Bioengineering, Mossakowski Medical Research Centre Polish Academy of Sciences, Warsaw, Poland.

2 Department of Veterinary Medicine (DIMEVET), Università degli Studi di Milano, Milan, Italy.

3 Laboratory of Clinical and Experimental Toxicology, Toxicology Unit, ICS Maugeri SpA-SB, IRCCS Pavia, Pavia, Italy.

4 Laboratory of Environmental Chemistry and Toxicology, Department of Environmental Health Sciences, Istituto di Ricerche Farmacologiche Mario Negri IRCCS, Italy.

Abstract

Organoids are three-dimensional self-aggregating structures generated from stem cells (SCs) or progenitor cells in a process that recapitulates molecular and cellular stages of early organ development. The differentiation process leads to the appearance of specialized mature cells and is connected with changes in the organoid internal structure rearrangement and self-organization. The formation of organ-specific structures in vitro with highly ordered architecture is also strongly influenced by the extracellular matrix. These features make organoids as a powerful model for in vitro toxicology. Nowadays this technology is developing very quickly. In this review we present, from a toxicological and species-specific point of view, the state of the art of organoid generation from adult SCs and pluripotent SCs: embryonic SCs or induced pluripotent SCs. The current culture organoid techniques are discussed for their main advantages, disadvantages and limitations. In the second part of the review, we concentrated on the characterization of species-specific organoids generated from tissue-specific SCs of different sources: mammary (bovine), epidermis (canine), intestinal (porcine, bovine, canine, chicken) and liver (feline, canine).

KEYWORDS:

3D models; ESCs; iPSCs; organoids; spheroids; stem cells; toxicology

 

Reference Gene Validation via RT-qPCR for Human iPSC-Derived Neural Stem Cells and Neural Progenitors.

Mol Neurobiol. 2019 Mar 29. doi: 10.1007/s12035-019-1538-x. [Epub ahead of print]

Augustyniak J1, Lenart J2, Lipka G1, Stepien PP3, Buzanska L4.

Author information

1 Department of Stem Cell Bioengineering, Mossakowski Medical Research Centre Polish Academy of Sciences, 5 Pawinskiego Str., 02-106, Warsaw, Poland.

2 Department of Neurochemistry, Mossakowski Medical Research Centre Polish Academy of Sciences, 5 Pawinskiego Str., 02-106, Warsaw, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

3 Institute of Genetics and Biotechnology, Faculty of Biology, University of Warsaw, 5a Pawinskiego Str., 02-106, Warsaw, Poland.

4 Department of Stem Cell Bioengineering, Mossakowski Medical Research Centre Polish Academy of Sciences, 5 Pawinskiego Str., 02-106, Warsaw, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

 

Abstract

Correct selection of the reference gene(s) is the most important step in gene expression analysis. The aims of this study were to identify and evaluate the panel of possible reference genes in neural stem cells (NSC), early neural progenitors (eNP) and neural progenitors (NP) obtained from human-induced pluripotent stem cells (hiPSC). The stability of expression of genes commonly used as the reference in cells during neural differentiation is variable and does not meet the criteria for reference genes. In the present work, we evaluated the stability of expression of 16 candidate reference genes using the four most popular algorithms: the ΔCt method, BestKeeper, geNorm and NormFinder. All data were analysed using the online tool RefFinder to obtain a comprehensive ranking. Our results indicate that NormFinder is the best tool for reference gene selection in early stages of hiPSC neural differentiation. None of the 16 tested genes is suitable as reference gene for all three stages of development. We recommend using different genes (panel of genes) to normalise RT-qPCR data for each of the neural differentiation stages.

KEYWORDS:

Gene expression reference panel; Neural Progenitor; Neural Stem Cells; Relative gene expression; human induced Pluripotent Stem Cell (hiPSC); quantitative real-time Polymerase Chain Reaction (RT-qPCR)

 

A Distinct Advantage to Intraarterial Delivery of 89Zr-Bevacizumab in PET Imaging of Mice With and Without Osmotic Opening of the Blood-Brain Barrier.

A Distinct Advantage to Intraarterial Delivery of 89Zr-Bevacizumab in PET Imaging of Mice With and Without Osmotic Opening of the Blood-Brain Barrier.

J Nucl Med. 2019 May;60(5):617-622. doi: 10.2967/jnumed.118.218792. Epub 2018 Oct 12.

Lesniak WG1, Chu C1,2, Jablonska A1,2, Du Y1, Pomper MG1, Walczak P1,2,3, Janowski M4,2,5.

Author information

1        Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins    University School of Medicine, Baltimore, Maryland.

2        Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland.

3        Department of Neurosurgery, University of Warmia and Mazury, Olsztyn, Poland; and.

4        Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University School of Medicine, Baltimore, Maryland Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

5        NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

Abstract

Glioblastoma multiforme (GBM) is the most aggressive and common type of brain cancer. Five-year survival rates are below 12%, even with the most aggressive trimodal therapies. Poor blood-brain barrier (BBB) permeability of therapeutics is a major obstacle to efficacy. Intravenous administration of bevacizumab is the standard treatment for GBM. It has been recently demonstrated that a single intraarterial infusion of bevacizumab provides superior therapeutic outcomes in patients with recurrent GBM. Further GBM treatment benefits can be achieved through opening of the BBB before intraarterial infusion of bevacizumab. However, a rationale for intraarterial delivery and BBB opening when delivering antibodies is lacking. A method facilitating quantification of intraarterial delivery of bevacizumab is needed for more effective and personalized GBM treatment. Here, we demonstrate such a method using PET imaging of radiolabeled bevacizumab. Methods: Bevacizumab was conjugated with deferoxamine and subsequently radiolabeled with 89Zr. 89Zr-bevacizumab deferoxamine (89Zr-BVDFO) was prepared with a specific radioactivity of 81.4 ± 7.4 MBq/mg (2.2 ± 0.2 μCi/mg). Brain uptake of 89Zr-BVDFO on carotid artery and tail vein infusion with an intact BBB or with BBB opening with mannitol was initially monitored by dynamic PET, followed by whole-body PET/CT at 1 and 24 h after infusion. Th ex vivo biodistribution of 89Zr-BVDFO was also determined. Results: Intraarterial administration of 89Zr-BVDFO resulted in gradual accumulation of radioactivity in the ipsilateral hemisphere, with 9.16 ± 2.13 percentage injected dose/cm3 at the end of infusion. There was negligible signal observed in the contralateral hemisphere. BBB opening with mannitol before intraarterial infusion of 89Zr-BVDFO resulted in faster and higher uptake in the ipsilateral hemisphere (23.58 ± 4.46 percentage injected dose/cm3) and negligible uptake in the contralateral hemisphere. In contrast, intravenous infusion of 89Zr-BVDFO and subsequent BBB opening did not lead to uptake of radiotracer in the brain. The ex vivo biodistribution results validated the PET/CT studies. Conclusion: Our findings demonstrate that intraarterial delivery of bevacizumab into the brain across an osmotically opened BBB is effective, in contrast to the intravenous route.

© 2019 by the Society of Nuclear Medicine and Molecular Imaging.

KEYWORDS:

brain; drug delivery; endovascular; molecular imaging; nuclear medicine

 

Biodistribution of Glial Progenitors in a Three Dimensional-Printed Model of the Piglet Cerebral Ventricular System.

Biodistribution of Glial Progenitors in a Three Dimensional-Printed Model of the Piglet Cerebral Ventricular System.

Stem Cells Dev. 2019 Apr 15;28(8):515-527. doi: 10.1089/scd.2018.0172. Epub 2019 Mar 28.

Srivastava RK1,2, Jablonska A1,2, Chu C1,2, Gregg L3, Bulte JWM1,2, Koehler RC4, Walczak P1,2,5, Janowski M1,2,6.

Author information

1        Division of MR Research, Russell H. Morgan Department of Radiology and Radiological Science, The Johns Hopkins University School of Medicine, Baltimore, Maryland.

2        Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, The Johns Hopkins University School of Medicine, Baltimore, Maryland.

3        Visualization Core Laboratory, Russell H. Morgan Department of Radiology and Radiological Science, The Johns Hopkins University School of Medicine, Baltimore, Maryland.

4        Department of Anesthesiology and Critical Care Medicine, Translational Tissue Engineering Center, The Johns Hopkins University School of Medicine, Baltimore, Maryland.

5        Department of Neurology and Neurosurgery, University of Warmia and Mazury, Olsztyn, Poland.

6        NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

Abstract

White matter damage persists in hypoxic-ischemic newborns even when treated with hypothermia. We have previously shown that intraventricular delivery of human glial progenitors (GPs) at the neonatal stage is capable of replacing abnormal host glia and rescuing the lifespan of dysmyelinated mice. However, such transplantation in the human brain poses significant challenges as related to high-volume ventricles and long cell migration distances. These challenges can only be studied in large animal model systems. In this study, we developed a three dimensional (3D)-printed model of the ventricular system sized to a newborn pig to investigate the parameters that can maximize a global biodistribution of injected GPs within the ventricular system, while minimizing outflow to the subarachnoid space. Bioluminescent imaging and magnetic resonance imaging were used to image the biodistribution of luciferase-transduced GPs in simple fluid containers and a custom-designed, 3D-printed model of the piglet ventricular system. Seven independent variables were investigated. The results demonstrated that a low volume (0.1 mL) of cell suspension is essential to keep cells within the ventricular system. If higher volumes (1 mL) are needed, a very slow infusion speed (0.01 mL/min) is necessary. Real-time magnetic resonance imaging demonstrated that superparamagnetic iron oxide (SPIO) labeling significantly alters the rheological properties of the GP suspension, such that, even at high speeds and high volumes, the outflow to the subarachnoid space is reduced. Several other factors, including GP species (human vs. mouse), type of catheter tip (end hole vs. side hole), catheter length (0.3 vs. 7.62 m), and cell concentration, had less effect on the overall distribution of GPs. We conclude that the use of a 3D-printed phantom model represents a robust, reproducible, and cost-saving alternative to in vivo large animal studies for determining optimal injection parameters.

KEYWORDS:

CSF; MRI; bioluminescence; brain; glial progenitors; iron oxide; ventricle

 

Challenges and Controversies in Human Mesenchymal Stem Cell Therapy.

Challenges and Controversies in Human Mesenchymal Stem Cell Therapy.

Stem Cells Int. 2019 Apr 9;2019:9628536. doi: 10.1155/2019/9628536. eCollection 2019.

Lukomska B1, Stanaszek L1, Zuba-Surma E2, Legosz P3, Sarzynska S3, Drela K1.

Author information

  1. NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
  2. Department of Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland.
  3. Department of Orthopedics and Traumatology, 1st Faculty of Medicine, Medical University of Warsaw, Warsaw, Poland.

Abstract

Stem cell therapy is being intensely investigated within the last years. Expectations are high regarding mesenchymal stem cell (MSC) treatment in translational medicine. However, many aspects concerning MSC therapy should be profoundly defined. Due to a variety of approaches that are investigated, potential effects of stem cell therapy are not transparent. On the other hand, most results of MSC administration in vivo have confirmed their safety and showed promising beneficial outcomes. However, the therapeutic effects of MSC-based treatment are still not spectacular and there is a potential risk related to MSC applications into specific cell niche that should be considered in long-term observations and follow-up outcomes. In this review, we intend to address some problems and critically discuss the complex nature of MSCs in the context of their effective and safe applications in regenerative medicine in different diseases including graft versus host disease (GvHD) and cardiac, neurological, and orthopedic disorders.

 

Concise Review: Mesenchymal Stem Cells: From Roots to Boost.

Concise Review: Mesenchymal Stem Cells: From Roots to Boost.

 Stem Cells. 2019 Apr 12. doi: 10.1002/stem.3016. [Epub ahead of print]

Andrzejewska A1, Lukomska B1, Janowski M1,2,3.

Author information

1        NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

2        Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.

3        Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, Johns Hopkins University, Baltimore, Maryland, USA.

Abstract

It was shown as long as half a century ago that bone marrow is a source of not only hematopoietic stem cells, but also stem cells of mesenchymal tissues. Then the term of "mesenchymal stem cells" (MSCs) has been coined in early 1990s and over a decade later, the criteria for defining MSCs have been released by International Society for Cellular Therapy. The easy derivation from a variety of fetal and adult tissues and not demanding cell culture conditions made MSCs an attractive research object. It was followed by the avalanche of reports from preclinical studies on potentially therapeutic properties of MSCs, such as immunomodulation, trophic support and capability for a spontaneous differentiation into connective tissue cells, and differentiation into majority of cell types upon specific inductive conditions. Although ontogenesis, niche, and heterogeneity of MSCs are still under investigation, there is a rapid boost of attempts in clinical applications of MSCs, especially for a flood of civilization-driven conditions in so quickly aging societies in not only developed countries, but also very populous developing world. The fields of regenerative medicine and oncology are particularly extensively addressed by MSC applications, in part due to paucity of traditional therapeutic options for these highly demanding and costly conditions. There are currently almost 1,000 clinical trials from entire world registered at clinicalTrials.gov and it seems that we are starting to witness the snowball effect with MSCs becoming a powerful global industry; however, spectacular effects of MSCs in clinic still need to be shown. Stem Cells 2019.

© AlphaMed Press 2019.

KEYWORDS:

Clinical; Differentiation; History; Immunomodulation; Mesenchymal stem cells; Paracrine activity

 

Experimental Strategies of Mesenchymal Stem Cell Propagation: Adverse Events and Potential Risk of Functional Changes.

Experimental Strategies of Mesenchymal Stem Cell Propagation: Adverse Events and Potential Risk of Functional Changes.

Stem Cells Int. 2019 Mar 6;2019:7012692. doi: 10.1155/2019/7012692. eCollection 2019.

Drela K1, Stanaszek L1, Nowakowski A1, Kuczynska Z1, Lukomska B1.

Author information

  1. NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

Abstract

Mesenchymal stem cells (MSCs) are attractive candidates for cell-based tissue repair approaches. Hundreds of clinical trials using MSCs have been completed and many others are still being investigated. For most therapeutic applications, MSC propagation in vitro is often required. However, ex vivo culture condition is not fully physiological and may affect biological properties of MSCs including their regenerative potential. Moreover, both cell cryopreservation and labelling procedure prior to infusion may have the negative impact on their expected effect in vivo. The incidence of MSC transformation during in vitro culture should be also taken into consideration before using cells in stem cell therapy. In our review, we focused on different aspects of MSC propagation that might influence their regenerative properties of MSC. We also discussed the influence of different factors that might abolish MSC proliferation and differentiation as well as potential impact of stem cell senescence and aging. Despite of many positive therapeutic effects of MSC therapy, one has to be conscious about potential cell changes that could appear during manufacturing of MSCs.

 

Effect of the HDAC Inhibitor, Sodium Butyrate, on Neurogenesis in a Rat Model of Neonatal Hypoxia-Ischemia: Potential Mechanism of Action.

Effect of the HDAC Inhibitor, Sodium Butyrate, on Neurogenesis in a Rat Model of Neonatal Hypoxia-Ischemia: Potential Mechanism of Action.

 Mol Neurobiol. 2019 Feb 14. doi: 10.1007/s12035-019-1518-1. [Epub ahead of print]

Jaworska J1, Zalewska T1, Sypecka J1, Ziemka-Nalecz M2.

Author information

1        Neurorepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, 5 A. Pawinskiego Street, 02-106, Warsaw, Poland.

2        Neurorepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, 5 A. Pawinskiego Street, 02-106, Warsaw, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

Neonatal hypoxic-ischemic (HI) brain injury likely represents the major cause of long-term neurodevelopmental disabilities in surviving babies. Despite significant investigations, there is not yet any known reliable treatment to reduce brain damage in suffering infants. Our recent studies in an animal model of HI revealed the therapeutic potential of a histone deacetylase inhibitor (HDACi). The neuroprotective action was connected with the stimulation of neurogenesis in the dentate gyrus subgranular zone. In the current study, we investigated whether HDACi-sodium butyrate (SB)-would also lead to neurogenesis in the subventricular zone (SVZ). By using a neonatal rat model of hypoxia-ischemia, we found that SB treatment stimulated neurogenesis in the damaged ipsilateral side, based on increased DCX labeling, and restored the number of neuronal cells in the SVZ ipsilateral to lesioning. The neurogenic effect was associated with inhibition of inflammation, expressed by a transition of microglia to the anti-inflammatory phenotype (M2). In addition, the administration of SB increased the activation of the TrkB receptor and the phosphorylation of the transcription factor-CREB-in the ipsilateral hemisphere. In contrast, SB administration reduced the level of HI-induced p75NTR. Together, these results suggest that BDNF-TrkB signaling plays an important role in SB-induced neurogenesis after HI. These findings provide the basis for clinical approaches targeted at protecting the newborn brain damage, which may prove beneficial for treating neonatal hypoxia-ischemia.

KEYWORDS:

BDNF–TrkB pathway; CREB; ERK kinase; Neonatal hypoxia–ischemia; Neurogenesis

 

 

The Role of Glia in Canine Degenerative Myelopathy: Relevance to Human Amyotrophic Lateral Sclerosis.

The Role of Glia in Canine Degenerative Myelopathy: Relevance to Human Amyotrophic Lateral Sclerosis.

Mol Neurobiol. 2019 Jan 23. doi: 10.1007/s12035-019-1488-3. [Epub ahead of print]

Golubczyk D1, Malysz-Cymborska I1, Kalkowski L1, Janowski M2,3,4, Coates JR5, Wojtkiewicz J6, Maksymowicz W1, Walczak P7,8,9.

Author information

1        Department of Neurosurgery, School of Medicine, University of Warmia and Mazury, Olsztyn, Poland.

2        Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University, Baltimore, MD, USA.

3        Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, Johns Hopkins University, Baltimore, MD, USA.

4        Department of NeuroRepair, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

5        Department of Veterinary Medicine and Surgery, College of Veterinary Medicine, University of Missouri, Columbia, MO, USA.

6        Department of Pathophysiology, School of Medicine, University of Warmia and Mazury, Olsztyn, Poland.

7        Department of Neurosurgery, School of Medicine, University of Warmia and Mazury, Olsztyn, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

8        Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University, Baltimore, MD, USA. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

9        Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, Johns Hopkins University, Baltimore, MD, USA. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive degeneration of motor neurons and grim prognosis. Over the last decade, studies on neurodegenerative diseases pointed on the role of glia in supporting the proper function of neurons. Particularly, oligodendrocytes were shown to be essential through myelin production and supplying axons with energy metabolites via monocarboxylate transporters (MCT). We have used dogs with naturally occurring degenerative myelopathy (DM) which closely resembles features observed in human ALS. We have performed two types of analysis of spinal cord tissue samples: histology and molecular analysis. Histology included samples collected from dogs that succumbed to the DM at different disease stages, which were compared to age-matched controls as well as put in the context of young spinal cords. Molecular analysis was performed on spinal cords with advanced DM and age-matched samples and included real-time PCR analysis of selected gene products related to the function of neurons, oligodendrocytes, myelin, and MCT. Demyelination has been detected in dogs with DM through loss of eriochrome staining and decreased expression of genes related to myelin including MBP, Olig1, and Olig2. The prominent reduction of MCT1 and MCT2 and increased MCT4 expression is indicative of disturbed energy supply to neurons. While Rbfox3 expression was not altered, the ChAT production was negatively affected. DM in dogs reproduces main features of human ALS including loss of motor neurons, dysregulation of energy supply to neurons, and loss of myelin, and as such is an ideal model system for highly translational studies on therapeutic approaches for ALS.

KEYWORDS:

ALS; Degenerative myelopathy; Demyelination; Disease model; Monocarboxylate transporters

 

SOD1/Rag2 Mice with Low Copy Number of SOD1 Gene as a New Long-Living Immunodeficient Model of ALS.

SOD1/Rag2 Mice with Low Copy Number of SOD1 Gene as a New Long-Living Immunodeficient Model of ALS.

Sci Rep. 2019 Jan 28;9(1):799. doi: 10.1038/s41598-018-37235-w.

Majchrzak M1, Drela K1, Andrzejewska A1, Rogujski P1, Figurska S2, Fiedorowicz M3, Walczak P4,5, Janowski M1,4, Lukomska B1, Stanaszek L6.

Author information

1        NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

2        Laboratory for Genetically Modified Animals, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

3        Department of Experimental Pharmacology and Small Animal Magnetic Resonance Imaging Laboratory, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

4        Johns Hopkins University School of Medicine, Institute for Cell Engineering, Division of MR Research, The Russell H. Morgan Department of Radiology and Radiological Science, Baltimore, MD, USA.

5        Department of Neurosurgery, School of Medicine, Collegium Medicum, University of Warmia and Mazury, Olsztyn, 10-719, Poland.

6        NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland. Ten adres pocztowy jest chroniony przed spamowaniem. Aby go zobaczyć, konieczne jest włączenie w przeglądarce obsługi JavaScript..

Abstract

The most recent research concerning amyotrophic lateral sclerosis (ALS) emphasizes the role of glia in disease development. Thus, one can suspect that the effective therapeutic strategy in treatment of ALS would be replacement of defective glia. One of the basic problems with human glial progenitors (hGRPs) replacement strategies is the time needed for the cells to become fully functional in vivo. The lifespan of most popular high copy number SOD1 mutant mice might be too short to acknowledge benefits of transplanted cells. We focused on developing immunodeficient rag2-/- model of ALS with lower number of transgene copies and longer lifespan. The obtained hSOD1/rag2 double mutant mice have been characterized. QPCR analysis revealed that copy number of hSOD1 transgene varied in our colony (4-8 copies). The difference in transgene copy number may be translated to significant impact on the lifespan. The death of long- and short-living hSOD1/rag2 mice is preceded by muscular weakness as early as one month before death. Importantly, based on magnetic resonance imaging we identified that mutant mice demonstrated abnormalities within the medullar motor nuclei. To conclude, we developed long-living double mutant hSOD1/rag2 mice, which could be a promising model for testing therapeutic utility of human stem cells.

 

Exchange-mode glutamine transport across CNS cell membranes

Neuropharmacology

Available online 8 March 2019

Invited review

Exchange-mode glutamine transport across CNS cell membranes

Jan Albrecht, Magdalena Zielińska

 

https://doi.org/10.1016/j.neuropharm.2019.03.003Get rights and content

Abstract

CNS cell membranes possess four transporters capable of exchanging Lglutamine (Gln) for other amino acids: the large neutral amino acid (LNAA) transporters LAT1 and LAT2, the hybrid basic amino acid (L-arginine (Arg), L-leucine (Leu)/LNAA transporter y+LAT2, and the L-alanine/L-serine/L-cysteine transporter 2 (ASCT2). LAT1/LAT2 and y+LAT2 are present in astrocytes, neurons and the blood brain barrier (BBB) - forming cerebral vascular endothelial cells (CVEC), while the location of ASCT2 in the individual cell types is a matter of debate. In the healthy brain, contribution of the exchangers to Gln shuttling from astrocytes to neurons and thus their role in controlling the conversion of Gln to the amino acid neurotransmitters l-glutamate (Glu) and γ-aminobutyric acid (GABA) and Gln flux across the BBB appears negligible as compared to the system A and system N uniporters. Insofar, except for the contribution of LAT1 to the maintenance of Gln homeostasis in the interstitial fluid (ISF), no well-defined CNS-specific function has been established for either of the three transporters in the healthy brain. The Gln-accepting amino acid exchangers appear to gain significance under conditions of excessive brain Gln load (glutaminosis). Excess Gln efflux across the BBB enhances influx into the brain of L-tryptophan (Trp). Excess of Trp is responsible for overloading the brain with neuroactive compounds: serotonin, kynurenic acid, quinolinic acid and/or oxindole, which contribute to neurotransmission imbalance accompanying hyperammonemia. In turn, alterations of y+LAT2-mediated Gln/Arg exchange and Arg uptake in astrocyte, modulate astrocytic nitric oxide synthesis and oxidative/nitrosative stress in ammonia-overexposed brain.

Keywords

Glutamine, Transport, Amino acid exchanger, Astrocyte, Blood-brain barier, Ammonia