One of the challenges in clinical translation of cell-replacement therapies is the definition of optimal cell generation and storage/recovery protocols which would permit a rapid preparation of cell-treatment products for patient administration. Besides, the availability of injection devices that are simple to use is critical for potential future dissemination of any spinally targeted cell-replacement therapy into general medical practice. Here, we compared the engraftment properties of established human-induced pluripotent stem cells (hiPSCs)-derived neural precursor cell (NPCs) line once cells were harvested fresh from the cell culture or previously frozen and then grafted into striata or spinal cord of the immunodeficient rat. A newly developed human spinal injection device equipped with a spinal cord pulsation-cancelation magnetic needle was also tested for its safety in an adult immunosuppressed pig. Previously frozen NPCs showed similar post-grafting survival and differentiation profile as was seen for freshly harvested cells. Testing of human injection device showed acceptable safety with no detectable surgical procedure or spinal NPCs injection-related side effects.

• MeSH
• buněčná diferenciace fyziologie   MeSH
• dospělí MeSH
• genetické vektory genetika   MeSH
• indukované pluripotentní kmenové buňky * fyziologie   transplantace   MeSH
• krysa rodu rattus MeSH
• lidé MeSH
• mícha MeSH
• mozek MeSH
• nervové kmenové buňky * fyziologie   transplantace   MeSH
• odběr biologického vzorku metody   MeSH
• odběr tkání a orgánů metody   MeSH
• prasata MeSH
• přeprogramování buněk * genetika   fyziologie   MeSH
• přežívání štěpu fyziologie   MeSH
• spinální injekce * škodlivé účinky   přístrojové vybavení   metody   MeSH
• transplantace kmenových buněk * škodlivé účinky   přístrojové vybavení   metody   MeSH
• virus Sendai MeSH
• výsledek terapie MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• krysa rodu rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Wnt signaling plays an important role in the self-renewal, fate-commitment and survival of the neural stem/progenitor cells (NS/PCs) of the adult central nervous system (CNS). Ischemic stroke impairs the proper functioning of the CNS and, therefore, active Wnt signaling may prevent, ameliorate, or even reverse the negative effects of ischemic brain injury. In this review, we provide the current knowledge of Wnt signaling in the adult CNS, its status in diverse cell types, and the Wnt pathway's impact on the properties of NS/PCs and glial cells in the context of ischemic injury. Finally, we summarize promising strategies that might be considered for stroke therapy, and we outline possible future directions of the field.

• MeSH
• buněčná diferenciace genetika   MeSH
• cílená molekulární terapie metody   trendy   MeSH
• dospělí MeSH
• ischemie mozku genetika   metabolismus   patologie   patofyziologie   MeSH
• lidé MeSH
• mozek cytologie   patologie   fyziologie   MeSH
• nervové kmenové buňky patologie   fyziologie   MeSH
• neurogeneze fyziologie   MeSH
• neuroglie patologie   fyziologie   MeSH
• signální dráha Wnt genetika   fyziologie   MeSH
• tranzitorní ischemická ataka genetika   metabolismus   patologie   terapie   MeSH
• zdraví MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Generation of neurons in the embryonic neocortex is a balanced process of proliferation and differentiation of neuronal progenitor cells. Canonical Wnt signalling is crucial for expansion of radial glial cells in the ventricular zone and for differentiation of intermediate progenitors in the subventricular zone. We detected abundant expression of two transcrtiption factors mediating canonical Wnt signalling, Tcf7L1 and Tcf7L2, in the ventricular zone of the embryonic neocortex. Conditional knock-out analysis showed that Tcf7L2, but not Tcf7L1, is the principal Wnt mediator important for maintenance of progenitor cell identity in the ventricular zone. In the absence of Tcf7L2, the Wnt activity is reduced, ventricular zone markers Pax6 and Sox2 are downregulated and the neuroepithelial structure is severed due to the loss of apical adherens junctions. This results in decreased proliferation of radial glial cells, the reduced number of intermediate progenitors in the subventricular zone and hypoplastic forebrain. Our data show that canonical Wnt signalling, which is essential for determining the neuroepithelial character of the neocortical ventricular zone, is mediated by Tcf7L2.

• MeSH
• buněčná diferenciace genetika   MeSH
• chlorid-hydrogenuhličitanové antiportéry MeSH
• down regulace genetika   MeSH
• embryo savčí MeSH
• hipokampus cytologie   embryologie   MeSH
• mutace genetika   MeSH
• myši transgenní MeSH
• myši MeSH
• neokortex cytologie   embryologie   MeSH
• nervové kmenové buňky fyziologie   MeSH
• neurogeneze fyziologie   MeSH
• neuroglie MeSH
• neurony fyziologie   MeSH
• počet buněk MeSH
• proliferace buněk genetika   MeSH
• protein 2 podobný transkripčnímu faktoru 7 genetika   metabolismus   MeSH
• proteiny T-boxu metabolismus   MeSH
• proteiny Wnt metabolismus   MeSH
• retinální gangliové buňky fyziologie   MeSH
• signální transdukce genetika   MeSH
• transkripční faktory SOXB1 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: The characterisation of dividing brain cells is fundamental for studies ranging from developmental and stem cell biology, to brain cancers. Whilst there is extensive anatomical data on these dividing cells, limited gene transcription data is available due to technical constraints. NEW METHOD: We focally isolated dividing cells whilst conserving RNA, from culture, primary neural tissue and xenografted glioma tumours, using a thymidine analogue that enables gene transcription analysis. RESULTS: 5-ethynyl-2-deoxyuridine labels the replicating DNA of dividing cells. Once labelled, cultured cells and tissues were dissociated, fluorescently tagged with a revised click chemistry technique and the dividing cells isolated using fluorescence-assisted cell sorting. RNA was extracted and analysed using real time PCR. Proliferation and maturation related gene expression in neurogenic tissues was demonstrated in acutely and 3 day old labelled cells, respectively. An elevated expression of marker and pathway genes was demonstrated in the dividing cells of xenografted brain tumours, with the non-dividing cells showing relatively low levels of expression. COMPARISON WITH EXISTING METHOD: BrdU "immune-labelling", the most frequently used protocol for detecting cell proliferation, causes complete denaturation of RNA, precluding gene transcription analysis. This EdU labelling technique, maintained cell integrity during dissociation, minimized copper exposure during labelling and used a cell isolation protocol that avoided cell lysis, thus conserving RNA. CONCLUSIONS: The technique conserves RNA, enabling the definition of cell proliferation-related changes in gene transcription of neural and pathological brain cells in cells harvested immediately after division, or following a period of maturation.

• MeSH
• analýza jednotlivých buněk metody   MeSH
• čichová sliznice fyziologie   MeSH
• deoxyuridin analogy a deriváty   MeSH
• embryonální kmenové buňky fyziologie   MeSH
• gliom patofyziologie   MeSH
• kultivované buňky MeSH
• lidé MeSH
• mozek * fyziologie   patofyziologie   MeSH
• myši inbrední C57BL MeSH
• myši inbrední NOD MeSH
• myši SCID MeSH
• nádory mozku * patofyziologie   MeSH
• nervové kmenové buňky fyziologie   MeSH
• neurogeneze * fyziologie   MeSH
• neurony * fyziologie   MeSH
• RNA metabolismus   MeSH
• stanovení celkové genové exprese metody   MeSH
• syntetická chemie okamžité shody MeSH
• transplantace nádorů MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Identifying the steps involved in striatal development is important both for understanding the striatum in health and disease, and for generating protocols to differentiate striatal neurons for regenerative medicine. The most prominent neuronal subtype in the adult striatum is the medium spiny projection neuron (MSN), which constitutes more than 85% of all striatal neurons and classically expresses DARPP-32. Through a microarray study of genes expressed in the whole ganglionic eminence (WGE: the developing striatum) in the mouse, we identified the gene encoding the transcription factor Forkhead box protein P1 (FoxP1) as the most highly up-regulated gene, thus providing unbiased evidence for the association of FoxP1 with MSN development. We also describe the expression of FoxP1 in the human fetal brain over equivalent gestational stages. FoxP1 expression persisted through into adulthood in the mouse brain, where it co-localised with all striatal DARPP-32 positive projection neurons and a small population of DARPP-32 negative cells. There was no co-localisation of FoxP1 with any interneuron markers. FoxP1 was detectable in primary fetal striatal cells following dissection, culture, and transplantation into the adult lesioned striatum, demonstrating its utility as an MSN marker for transplantation studies. Furthermore, DARPP-32 expression was absent from FoxP1 knock-out mouse WGE differentiated in vitro, suggesting that FoxP1 is important for the development of DARPP-32-positive MSNs. In summary, we show that FoxP1 labels MSN precursors prior to the expression of DARPP-32 during normal development, and in addition suggest that FoxP1 labels a sub-population of MSNs that are not co-labelled by DARPP-32. We demonstrate the utility of FoxP1 to label MSNs in vitro and following neural transplantation, and show that FoxP1 is required for DARPP-32 positive MSN differentiation in vitro.

• MeSH
• buněčná diferenciace fyziologie   MeSH
• corpus striatum * cytologie   embryologie   růst a vývoj   MeSH
• dopaminem a cAMP regulovaný fosfoprotein 32 metabolismus   MeSH
• embryo savčí MeSH
• forkhead transkripční faktory genetika   metabolismus   MeSH
• jaderné proteiny metabolismus   MeSH
• kultivované buňky MeSH
• myši knockoutované MeSH
• myši MeSH
• neparametrická statistika MeSH
• nervové kmenové buňky fyziologie   transplantace   MeSH
• neurony cytologie   metabolismus   MeSH
• novorozená zvířata MeSH
• plod cytologie   MeSH
• proteiny nervové tkáně metabolismus   MeSH
• represorové proteiny genetika   metabolismus   MeSH
• techniky in vitro MeSH
• transportní proteiny metabolismus   MeSH
• vývojová regulace genové exprese fyziologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The nucleolar protein 2 gene encodes a protein specific for the nucleolus. It is assumed that it plays a role in the synthesis of ribosomes and regulation of the cell cycle. Due to its link to cell proliferation, higher expression of Nop2 indicates a worse tumor prognosis. In this work we used Nop2(gt1gaj) gene trap mouse strain. While lethality of homozygous animals suggested a vital role of this gene, heterozygous animals allowed the detection of expression of Nop2 in various tissues, including mouse brain. Histochemistry, immunohistochemistry and immunoelectron microscopy techniques, applied to a mature mouse brain, human brain and on mouse neural stem cells revealed expression of Nop2 in differentiating cells, including astrocytes, as well as in mature neurons. Nop2 was detected in various regions of mouse and human brain, mostly in large pyramidal neurons. In the human, Nop2 was strongly expressed in supragranular and infragranular layers of the somatosensory cortex and in layer III of the cingulate cortex. Also, Nop2 was detected in CA1 and the subiculum of the hippocampus. Subcellular analyses revealed predominant location of Nop2 within the dense fibrillar component of the nucleolus. To test if Nop2 expression correlates to cell proliferation occurring during tissue regeneration, we induced strokes in mice by middle cerebral artery occlusion. Two weeks after stroke, the number of Nop2/nestin double positive cells in the region affected by ischemia and the periventricular zone substantially increased. Our findings suggest a newly discovered role of Nop2 in both mature neurons and in cells possibly involved in the regeneration of nervous tissue.

• MeSH
• astrocyty fyziologie   MeSH
• cévní mozková příhoda patofyziologie   MeSH
• dospělí MeSH
• infarkt arteria cerebri media MeSH
• jaderné proteiny genetika   metabolismus   MeSH
• kultivované buňky MeSH
• lidé MeSH
• modely nemocí na zvířatech MeSH
• mozek fyziologie   patofyziologie   MeSH
• myši inbrední C57BL MeSH
• myši transgenní MeSH
• nervové kmenové buňky fyziologie   MeSH
• nestin metabolismus   MeSH
• neurogeneze fyziologie   MeSH
• neurony fyziologie   MeSH
• proliferace buněk fyziologie   MeSH
• tRNA-methyltransferasy metabolismus   MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Chromatin compaction mediates progenitor to post-mitotic cell transitions and modulates gene expression programs, yet the mechanisms are poorly defined. Snf2h and Snf2l are ATP-dependent chromatin remodelling proteins that assemble, reposition and space nucleosomes, and are robustly expressed in the brain. Here we show that mice conditionally inactivated for Snf2h in neural progenitors have reduced levels of histone H1 and H2A variants that compromise chromatin fluidity and transcriptional programs within the developing cerebellum. Disorganized chromatin limits Purkinje and granule neuron progenitor expansion, resulting in abnormal post-natal foliation, while deregulated transcriptional programs contribute to altered neural maturation, motor dysfunction and death. However, mice survive to young adulthood, in part from Snf2l compensation that restores Engrailed-1 expression. Similarly, Purkinje-specific Snf2h ablation affects chromatin ultrastructure and dendritic arborization, but alters cognitive skills rather than motor control. Our studies reveal that Snf2h controls chromatin organization and histone H1 dynamics for the establishment of gene expression programs underlying cerebellar morphogenesis and neural maturation.

• MeSH
• adenosintrifosfatasy metabolismus   MeSH
• analýza rozptylu MeSH
• bromodeoxyuridin MeSH
• chromatinová imunoprecipitace MeSH
• chromozomální proteiny, nehistonové metabolismus   MeSH
• fluorescence MeSH
• galaktosidy MeSH
• histony metabolismus   MeSH
• homeodoménové proteiny metabolismus   MeSH
• hybridizace in situ MeSH
• imunohistochemie MeSH
• indoly MeSH
• koncové značení zlomů DNA in situ MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• metoda rotující tyčky MeSH
• mikročipová analýza MeSH
• morfogeneze genetika   fyziologie   MeSH
• mozeček embryologie   MeSH
• myši transgenní MeSH
• myši MeSH
• nervové kmenové buňky metabolismus   fyziologie   MeSH
• počítačové zpracování obrazu MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• Purkyňovy buňky metabolismus   MeSH
• restrukturace chromatinu fyziologie   MeSH
• toloniumchlorid MeSH
• transmisní elektronová mikroskopie MeSH
• vývojová regulace genové exprese genetika   fyziologie   MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Currently, there is no effective strategy for the treatment of spinal cord injury (SCI). A combination of biomaterials and stem cell therapy seems to be a promising approach to increase regenerative potential after SCI. We evaluated the use of a cellpolymer construct based on a combination of the conditionally immortalized spinal progenitor cell line SPC-01_GFP3, derived from human fetal spinal cord tissue, with a serotonin-modified poly(2-hydroxyethyl methacrylate) hydrogel (pHEMA-5HT). We compared the effect of treatment with a pHEMA-5HT hydrogel seeded with SPC-01_GFP3 cells, treatment with a pHEMA-5HT only and no treatment on functional outcome and tissue reconstruction in hemisected rats. Prior to transplantation the cell-polymer construct displayed a high potential to support the growth, proliferation and differentiation of SPC-01 cells in vitro. One month after surgery, combined hydrogel-cell treatment reduced astrogliosis and tissue atrophy and increased axonal and blood vessel ingrowth into the implant; however, two months later only the ingrowth of blood vessels remained increased. SPC-01_GFP3 cells survived well in vivo and expressed advanced markers of neuronal differentiation. However, a majority of the transplanted cells migrated out of the lesion and only rarely remained in the hydrogel. No differences among the groups in motor or sensory recovery were observed. Despite the support of the hydrogel as a cell carrier in vitro, and good results in vivo one month postsurgery, there was only a small effect on long term recovery, mainly due to the limited ability of the hydrogels to support the in vivo growth and differentiation of cells within the implant. Further modifications will be necessary to achieve stable long term improvement in functional outcome.

• MeSH
• atrofie etiologie   terapie   MeSH
• buněčná diferenciace MeSH
• cholinesterasy metabolismus   MeSH
• fetální kmenové buňky fyziologie   MeSH
• jizva farmakoterapie   etiologie   MeSH
• krysa rodu rattus MeSH
• lidé MeSH
• lokomoce účinky léků   fyziologie   MeSH
• modely nemocí na zvířatech MeSH
• myelinové proteiny metabolismus   MeSH
• nervové kmenové buňky fyziologie   MeSH
• PEG-DMA hydrogel aplikace a dávkování   MeSH
• poranění míchy farmakoterapie   chirurgie   MeSH
• potkani Wistar MeSH
• proliferace buněk MeSH
• proteiny nervové tkáně metabolismus   MeSH
• serotonin terapeutické užití   MeSH
• transfekce MeSH
• transplantace kmenových buněk * MeSH
• zelené fluorescenční proteiny genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• lidé MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The evidence for the existence of neurogenesis in the adult mammalian brain, including humans is now widely accepted. Despite the fact that adult neural stem cells appear to be very promising, a wide range of their unrevealed properties, abilities but also limitations under physiological and especially pathological conditions still need to be investigated and explained. Huntington's disease (HD) is characterized by successive degeneration of relatively well-defined neuronal population. Moreover, the most affected region, the caudate nucleus, is adjacent to the subependymal zone (SEZ) neurogenic region. Therefore, the possibility to harness the endogenous neural stem cell capacity for repairing, or at least restricting, the fatal neurodegenerative process in HD patients using promoted neurogenesis in the adult SEZ represent the exciting new possibility in clinical management of this disorder. On the other hand, many questions have to be answered before neuronal replacement therapies using endogenous precursors become a reality, particularly in relation to neurodegenerative diseases. Fundamental for all experimental, functional and future clinical studies is detailed morphological description of structures involved in the process of neurogenesis. The objectives of this review are to describe neurogenesis in the adult murine and human brain (with particular emphasis to morphological aspects of this process) and to determine to what extent it is affected in animal models of HD and in the human HD brain. Due to very limited evidence referring to the impact of striatal pathology of HD phenotype on the adult neurogenesis in the SEZ, some results gained from our studies on two rat models of HD, i.e. the neurotoxic lesion and transgenic HD rats, and on human HD brains are discussed.

• MeSH
• buněčná diferenciace fyziologie   MeSH
• degenerace nervu patologie   MeSH
• fenotyp MeSH
• Huntingtonova nemoc patologie   patofyziologie   terapie   MeSH
• lidé MeSH
• nervové kmenové buňky fyziologie   transplantace   MeSH
• neurogeneze fyziologie   MeSH
• neurony patologie   fyziologie   MeSH
• nucleus caudatus patologie   patofyziologie   MeSH
• proliferace buněk MeSH
• savci MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH