Papers by Krystyna Domanska-janik
Cell Transplantation, 2012
Mucopolysaccharidosis type I (MPS-I; Hurler syndrome) is an inborn error of metabolism caused by ... more Mucopolysaccharidosis type I (MPS-I; Hurler syndrome) is an inborn error of metabolism caused by lack of the functional lysosomal glycosaminoglycan (GAG)-degrading enzyme α-L-iduronidase (IDUA). Without treatment, the resulting GAG accumulation causes multisystem dysfunction and death within the first decade. Current treatments include allogeneic hematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy. HSCT ameliorates clinical features and extends life but is not available to all patients, and inadequately corrects the most devastating features of the disease including mental retardation and skeletal deformities. Recent developments suggest that stem cells can be used to deliver needed enzymes to the central nervous system. To test this concept, we transplanted bone marrow-derived normal adult human MultiStem® cells into the cerebral lateral ventricles of immunodeficient MPS-I neonatal mice. Transplanted cells and human-specific DNA were detected in the hippocampal formation, striatum, and other areas of the central nervous system. Brain tissue assays revealed significant long-term decrease in GAG levels in the hippocampus and striatum. Sensorimotor testing 6 months after transplantation demonstrated significantly improved rotarod performance of transplanted mice in comparison to nontransplanted and sham-transplanted control animals. These results suggest that a single injection of MultiStem cells into the cerebral ventricles of neonatal MPS-I mice induces sustained reduction in GAG accumulation within the brain, and modest long-term improvement in sensorimotor function.
Toxicology in Vitro, 2005
Neural stem cell line developed from human umbilical cord blood (HUCB-NSC) [Burajska et al., 2003... more Neural stem cell line developed from human umbilical cord blood (HUCB-NSC) [Burajska et al., 2003. Journal of Neurochemistry 85, 33] is an ethically uncontroversial source of stem cells, able to diVerentiate into neuronal, astrocytic and oligodendroglial lineages. Developmental fate decisions of HUCB-NSC can be experimentally manipulated in vitro by the presence of trophic factors, mitogenes and neuromorphogenes, but can also be inXuenced by neurotoxins. In this report two-dimensional (2-D) and three-dimensional (3-D) HUCB-NSC cultures are introduced as useful models for testing developmental neurotoxicity. For 2-D culture models we established a standardized method for the assessment of the growth rate and cell diVerentiation in 96-well plates. The proliferative capacity of the HUCB-NSC was monitored by the MTT test while their ability to diVerentiate into neural-like cells by immunocytochemistry of -tubulin III and MAP-2 for neurons, GFAP and S-100 for astrocytes and GalC for oligodendrocytes. The 3-D culture of HUCB-NSC is represented by neurospheres. Proliferation and migration of the intermediate precursors from attached neurospheres are shown to be controlled and altered by various growth factors and further modulated by the extracellular matrix component-Wbronectin. Thus, neurospheres derived from the HUCB-NSC line can represent a suitable model of the activation of dormant stem cells residing in their niche, and can be used for neurotoxic studies.
Cell transplantation, Jan 31, 2015
Ischemic stroke results in violent impairment of tissue homeostasis leading to severe perturbatio... more Ischemic stroke results in violent impairment of tissue homeostasis leading to severe perturbation within the neurovascular unit (NVU) during recovery period. The aim of study was to assess the potential of mesenchymal stem cells (MSC) originating from Wharton jelly (WJ) to differentiate into functionally competent cells of endothelial lineage (WJ-EPC). Protective effect of either primary WJ-MSC or induced WJ-EPC was investigated and compared after oxygen-glucose deprivation (OGD) of hippocampal organotypic slices (OHC) in the indirect co-culture model. WJ-EPC, primed in EGM-2 media under 5% O2, acquired cobblestone endothelial-like morphology, formed capillary-like structures and actively took up DiI-Ac-LDL. The both cell types (WJ-MSC and WJ-EPC) were positive for CD73, CD90, CD105, VEGFR-2, VEGF, but only endothelial one expressed vWF and PECAM-1 markers at significant levels. In the presence of either WJ-MSC or WJ-EPC culture at compartment below of OGD-injured slices, substanti...
From Mechanisms to Technologies, 2011
Acta neurobiologiae experimentalis, 2011
Many types of neural progenitors from various sources have been evaluated for therapy of CNS diso... more Many types of neural progenitors from various sources have been evaluated for therapy of CNS disorders. Prerequisite for success in cell therapy is the ability for transplanted cells to reach appropriate target such as stroke lesion. We have established neural stem cell line from human umbilical cord blood neural stem (HUCB-NSC). In the present study we evaluated migratory capabilities of cells (HUCB-NSC) and the presence of various migration-related receptors. Immunocytochemical analysis revealed abundant expression of CXCR4, PDGFR-alpha, PDGFR-beta, c-Met, VEGFR, IGF-1R and PSA-NCAM receptors in non-adherent population of HUCB-NSC cultured in serum free (SF) conditions (SF cells). Biological activity of selected receptors was confirmed by HUCB-NSC in vitro migration towards SDF-1 and IGF-1 ligands. Additionally, rat brain-derived homogenates have been assessed for their chemoattractive activity of HUCB-NSC. Our experiments unveiled that brain tissue was more attracted for HUCB-NSC...
Acta neurobiologiae experimentalis, 2006
Human umbilical cord blood (HUCB) is considered a promising source of neural progenitors capable ... more Human umbilical cord blood (HUCB) is considered a promising source of neural progenitors capable of being used for cellular therapies in neurological disorders. Here we review briefly our work on the elucidation of mechanisms and development of practical standards as regards the selection, maintenance and use of cord blood derivatives for such purposes. Our results join those of other recent studies in suggesting strongly that, the generation of neural-like cells from tissue belonging to a different germ layer (such as a cord blood is) is most probably explained by reference to a discrete subpopulation of embryonic-like stem cells of pluripotent characteristics. Such cells identified in cord blood through their expression of specific genetic and protein markers can be expanded in vitro and directed toward neurally-committed progenitors differentiating further into more mature neuron-like or macroglia-like cell phenotypes. From this HUCB-derived neural progenitor fraction a novel neu...
Molecular and Chemical Neuropathology, 1992
Calcium-activated neutral protease (CANP) in normal and dysmyelinating mutant, paralytic tremor (... more Calcium-activated neutral protease (CANP) in normal and dysmyelinating mutant, paralytic tremor (PT) rabbit myelin and premyelin fractions was studied using immature (4-5 wk) or adult animals. The enzyme was estimated by determination of its catalytic activity as well as by using immunoblot analysis after SDS-PAGE separation. The presence of two forms of CANP--one activated by calcium in the micromolar concentration (mu CANP) range and the other exhibiting low calcium sensitivity in the millimolar concentration range (m-CANP)--was found in the myelin and premyelin fractions. The developmental pattern of the enzyme activity was different for each of these two enzyme isoforms depending on the fraction studied. The higher activity on CANP (both isoforms) found in PT myelin and premyelin could be related to delayed myelination and/or to the higher turnover rate of already formed myelin. These results suggest complex and specific roles for these isoenzymes during myelin formation as is discussed further in this article. Our results confirm the extensive degradation of myelin basic protein (MBP), proteolipid protein (PLP), and, to a lesser extent, the other myelin proteins by endo- and exogenous CANP. This degradation process was significantly elevated in PT rabbit myelin. Moreover as was shown by two-dimensional gel electrophoresis, calcium-controlled proteolysis in nonmutant rabbits affected the net-charge of MBP in a manner similar to that reported for PT myelin, suggesting the possible involvement of CANP in the generation of charge isomers of MBP.
Resuscitation, 1979
Energy metabolism was studied in the cerebral cortex of rats during and after hypoxia induced by ... more Energy metabolism was studied in the cerebral cortex of rats during and after hypoxia induced by breathing a gas mixture of 7% O2 in N2 for 2 h. Cortical energy stores (2ATP + ADP + phosphocreatine) remained unchanged after hypoxic treatment. Lactate rose over four-fold. Pyruvate, glucose and glucose 6-phosphate concentrations also increased significantly. Metabolic activity in the cortex expressed as the utilization of high-energy phosphates 5, 10 and 30 s after decapitation was decreased by 30% after hypoxia and remained lowered for 3 h during recovery. This was accompanied by elevated glucose consumption and lactate production, suggesting that the maintenance of the energy balance after hypoxia was partly due to activation of the glycolytic pathway. During the recovery period, these metabolic abnormalities returned towards control values, but, after 6 h of recovery the high-energy phosphate utilization increased transitorily above the control values.
NeuroReport, 1994
The effect of the platelet activating factor (PAF) antagonist BN52021 on [3H]D-aspartate (D-Asp) ... more The effect of the platelet activating factor (PAF) antagonist BN52021 on [3H]D-aspartate (D-Asp) release was investigated in rat hippocampal slices during and after incubation (20 min) in ischaemia-like conditions. Ischaemia did not influence spontaneous D-Asp outflow whereas K(+)-evoked, calcium-dependent release was markedly enhanced in reoxygenated, post-ischaemic slices. These slices also showed a substantial translocation/activation of protein kinase C (PKC). BN52021 blocked both ischaemia-induced effects. Moreover, the PKC inhibitor H7 attenuated post-ischaemic K(+)-evoked D-Asp release when beta-PDBu, a PKC activator, was used to enhance the response of normoxic slices. Assuming that PKC is activated by ischaemia in a PAF-dependent manner and that this activation proceeds to enhanced glutamate exocytosis, we speculate on the involvement of PAF receptor stimulation in the pathology of cerebral ischaemia.
Journal of Neurochemistry, 2002
Paralytic tremor (pt) is a sex-linked mutation in rabbit that affects myelination of the CNS. Mye... more Paralytic tremor (pt) is a sex-linked mutation in rabbit that affects myelination of the CNS. Myelin in the pt brains represents approximately 30% of the normal levels. Previously we showed that the pt mutation affects primarily proteolipid protein (Plp) gene expression. In the present study we investigated the relative effect of the pt mutation on two distinctive Plp gene products, PLP- and DM-20-specific messenger RNAs. Our results showed that both PLP and DM-20 are affected and that the ratio DM-20/PLP was higher in pt rabbits than in age-matched controls. We sequenced normal rabbit PLP cDNA and characterized pt mutation at the DNA level. Rabbit PLP sequence, deduced from cDNA, differs from the human protein only at Thr198. Sequence analysis of the mutant cDNA revealed a transversion T-->A in exon 2 of the Plp gene. This point mutation, which is placed at the end of the first potential transmembrane domain, results in a substitution of His36 by a glutamine. This transversion abolishes a restriction site that enabled us to screen a large number of animals and observe a perfect correlation between the pt allele and the abnormal phenotype.
Journal of Histochemistry & Cytochemistry, 2011
Cell Transplantation, 2013
Since the brain is naturally inefficient in regenerating functional tissue after injury or diseas... more Since the brain is naturally inefficient in regenerating functional tissue after injury or disease, novel restorative strategies including stem cell transplantation and tissue engineering have to be considered. We have investigated the use of such strategies in order to achieve better functional repair outcomes. One of the fundamental challenges of successful transplantation is the delivery of cells to the injured site while maintaining cell viability. Classical cell delivery methods of intravenous or intraparenchymal injections are plagued by low engraftment and poor survival of transplanted stem cells. Novel implantable devices such as 3D bioactive scaffolds can provide the physical and metabolic support required for successful progenitor cell engraftment, proliferation, and maturation. In this study, we performed in situ analysis of laminin-linked dextran and gelatin macroporous scaffolds. We revealed the protective action of gelatin-laminin (GL) scaffolds seeded with mesenchymal stem cells derived from donated human Wharton's jelly (hUCMSCs) against neuroinflammatory reactions of injured mammalian brain tissue. These bioscaffolds have been implanted into (i) intact and (ii) ischemic rat hippocampal organotypic slices and into the striatum of (iii) normal and (iv) focally injured brains of adult Wistar rats. We found that transplantation of hUCMSCs encapsulated in GL scaffolds had a significant impact on the prevention of glial scar formation (low glial acidic fibrillary protein) and in the reduction of neuroinflammation (low interleukin-6 and the microglial markers ED1 and Iba1) in the recipient tissue. Moreover, implantation of hUCMSCs encapsulated within GL scaffolds induced matrix metalloproteinase-2 and -9 proteolytic activities in the surrounding brain tissue. This facilitated scaffold biodegradation while leaving the remaining grafted hUCMSCs untouched. In conclusion, transplanting GL scaffolds preseeded with hUCMSCs into mammalian brain tissue escaped the host's immune system and protected neural tissue from neuroinflammatory injury. This manuscript is published as part of the International Association of Neurorestoratology (IANR) supplement issue of Cell Transplantation.
Brain Research, 2001
Relatively mild ischemic insult can lead to delayed neuronal cell death in vulnerable brain regio... more Relatively mild ischemic insult can lead to delayed neuronal cell death in vulnerable brain regions. We provide evidence that the protein composition of the postsynaptic densities (PSD) undergoes rapid modification after 15 min postdecapitative as well as 5 min transient global ischemia. We observed a significant increase in cPKC and nPKC protein content in the postischemic PSD. Of the calcium-regulated PKC isoforms, the alpha and beta subtypes increase in PSD over ten times above the control values whereas gamma PKC, an isoform most abundant in the native PSD structure, shows relatively smaller changes under ischemic conditions. For the first time, the PSD membrane translocation of Ca(2+)-independent isoforms delta and epsilon is shown. The yield of the PSD protein preparation from the postischemic cortex was two times higher compared with control. This correlated with an abundant increase in electron density and changes in ultrastructure of PSD isolated from postischemic cortex. Also sections from CA1 gerbils hippocampus after transient ischemia showed persistent enlargement of postsynaptic densities up to 24 h of reperfusion. This was accompanied by elevation of the PSD/cytoskeleton-associated alpha, beta PKC immunoreactivity and other changes in neuronal and glial cell morphology typical of the early postischemic degeneration. Sustained changes in PKC composition and organization of postsynaptic membranes during and after ischemia may cause persistent alteration in synaptic transmission and subsequently contribute to delayed neuronal injury.
Brain Research, 2005
Two different models of brain ischemia were used to examine the evoked changes in the tyrosine ph... more Two different models of brain ischemia were used to examine the evoked changes in the tyrosine phosphorylation of NMDA receptor subunits 2A and 2B (NR2A and NR2B), as well as their interactions with non-receptor tyrosine kinases (NRTKs: FAK, PYK2 Src), and PSD-95 protein. Only short-term 5 min ischemia followed by 3 h reperfusion resulted in the elevated tyrosine phosphorylation of both investigated NMDA receptor subunits, but in contrast to previously published data, more pronounced in the case of NR2B. Concomitantly, an increased association of NR2B with FAK, PYK2, Src and PSD-95 has been observed. This sharp early reaction to brief ischemia was markedly attenuated during prolonged recovery (72 h) with almost complete return to control values. The initial recruitment of tyrosine kinases to NMDA receptor during the first 3 h of reperfusion is generally consistent with an active postischemic remodeling of PSD and may participate in the induction of the postischemic signal transduction pathway in gerbil hippocampus. In contrast, ischemia of longer duration (up to 30 min) caused an immediate decrease in the protein levels as well as tyrosine phosphorylation of both NR2A and NR2B subunits which was accompanied by the marked attenuation of the association with their investigated molecular partners-PSD-95 and NRTKs. This effect may be mimicked in vitro by Ca 2+ -dependent activation of endogenous calpains in purified PSD preparation suggesting irreversible deterioration of the synaptic signaling machinery during irreversible long-term ischemia. D
Brain Research, 1993
Paralytic tremor (pt) is a neurological sex-linked recessive mutation in rabbits which is charact... more Paralytic tremor (pt) is a neurological sex-linked recessive mutation in rabbits which is characterized by a coarse body tremor and limb paresis. Morphological studies showed that this mutation affects CNS myelination. Although the number of oligodendrocytes is not reduced, myelination is slower, irregular and defective. We have made a biochemical and molecular analysis of 4-wk-old mutant and normal rabbits. The amount of myelin in the mutant represents only approximately 25% of the normal level. Radioimmunoassay for myelin basic protein showed a reduction to approximately 40% in pt whole-brain homogenate but the difference was not significant in purified myelin. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of brain homogenates followed by immunoblotting showed that all major myelin proteins are affected by the pt mutation, although to different degrees. While most of the myelin proteins are reduced to approximately 60-80% of the normal level, an important reduction to approximately 30%, was measured for the proteolipid protein (PLP). In purified myelin, the difference in PLP concentration was significant while the other specific proteins were less affected. A similar reduction in myelin-protein gene expression was detected at the mRNA level. Sex-linked transmission, low concentrations of PLP and its specific mRNA in the CNS indicate that the pt mutation primarily affects the expression of the Plp gene.
Archives of Biochemistry and Biophysics, 2013
Increasi ng global birth rate, coupled with the aging population surviving into their eighth deca... more Increasi ng global birth rate, coupled with the aging population surviving into their eighth decade has lead to increased incidence diseases, hitherto designated as rare . Brain related ischemia, at birth, or later in life, during, for exampl e stroke, is increasing in global prevalence. Reactive microglia can contribute to neuronal damage as well as compromising transplantion. One potential treatment strategy is cellular therapy, using mesenchymal stem cells (hMSCs), which possess immunomodulatory and cell repair properties. For effective clinical therapy, mechanisms of action must be understood better. Here multicentre international laboratories assessed this question together investigating application of hMSCs neural involvement , with interest in the role of reactive microglia. Modulation by hMSCs in our in vivo and in vitro study shows they decrease markers of microglial activation (lower ED1 and Iba) and astrogliosi s (lower GFAP) following transplantation in an ouabain-induc ed brain ischemia rat model and in organotypic hippocampal cultures. The anti-inflammatory effect in vitro was demonstrated to be CD200 ligand depende nt with ligand expression shown to be increased by IL-4 stimulation. hMSC transplant reduced rat microglial STAT3 gene expression and reduced activation of Y705 phosphorylated STAT3, but STAT3 in the hMSCs themselv es was elevated upon grafting. Surprisingly, activity was dependent on heterodimerisation with STAT1 activated by IL-4 and Oncostatin M. Our study paves the way to preclinical stages of a clinical trial with hMSC, and suggests a non-canon ical JAK-STAT signaling of unphosphorylated STAT3 in immunomo dulatory effects of hMSCs.
Stem Cells and Development, 2009
The umbilical cord blood-derived neural stem/progenitor cells (HUCB-NSCs) potentially represent a... more The umbilical cord blood-derived neural stem/progenitor cells (HUCB-NSCs) potentially represent a rich source of transplantable material for treatment of a wide range of neurological diseases. Although, recently reported effects of their implementation in animal models of brain pathology are still controversial. As a simplified alternative to in vivo transplantation in this work we have applied a long-term organotypic rat hippocampal slice culture (OHC) as a recipient tissue to study bilateral graft/host cells interactions ex vivo. This type of culture can be considered as a kind of reductionistic model of brain transplantation where direct influence of systemic immunological responses to transplanted human cells would be excluded. The transplantation material derived from a HUCB-NSC line developed and characterized in our laboratory and delivered to the slices either as a single-cell suspension or after formation of typical neurospheres in serum-free medium in vitro (N-HUCBs). Experiments were focused on space-temporal context of cell transplantation in relation to their ability to ingrown, migrate, and differentiate within the slice cytoarchitecture. We gain evidences that these responses are strictly dependent on the engraftment site and that cell movement reflects typical routes used for migratory neuroblasts in vivo. The cells implanted at the second week of slice cultivation ingrown readily and deeply into host cytoarchitecture then matured to the level never observed in our transplantation animal models in vivo. Importantly, transplanted neurospheres, in addition to yield exogenous migratory cells to the host tissue can locally inhibit astrocytosis and promote outgrow of DCX-reactive neuroblasts in the surrounding OHC tissue.
Behavioural brain research, Jan 3, 2015
A stroke in humans may induce focal injury to the brain tissue resulting in various disabilities.... more A stroke in humans may induce focal injury to the brain tissue resulting in various disabilities. Although motor deficits are the most discernible, cognitive impairments seem to be crucial for patients mental well-being. The current lack of effective treatments encourages scientists and clinicians to develop novel approaches. Before applying them in clinic, testing for safety and effectiveness in non-human models is necessary. Such animal model should include significant cognitive impairments resulting from brain lesion. We used ouabain stereotactic injection into the right dorsolateral striatum of male Wistar rats, and enriched environment housing. To confirm the brain injury before cognitive testing, rats were given a beam-walking task to evaluate the level of sensorimotor deficits. To determine the cognitive impairment after focal brain damage, rats underwent a set of selected tasks over an observation period of 30 days. Brain injury induced by ouabain significantly impaired the ...
Acta neurobiologiae experimentalis, 2013
The NG2-positive cells are the oligodendrocyte precursors, which, when terminally differentiated,... more The NG2-positive cells are the oligodendrocyte precursors, which, when terminally differentiated, are capable of myelinating the central nervous system. There is however an ever-growing list of evidences that NG2 cells actually possess an intrinsic neurogenic potential and they are capable of neuronal differentiation in response to environmental stimuli. To address the question, we have established a model of an indirect co-culture system of the freshly isolated rat neonatal NG2 cells and organotypic slices derived from two distinct CNS regions (hippocampus and spinal cord) to mimic the nervous tissue microenviroment. The cell differentiation in microenvironment of OGD-injured hippocampal slices has been studied as well. The molecular analysis of selected trophic factors has been performed to determine the patterns of their expression. Indeed, the comparison of the cell commitment and development in various microenvironments has pointed to significant dissimilarities. First of all, ...
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Papers by Krystyna Domanska-janik