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NG2-glia proliferation and differentiation following CNS injuries
Kirdajová, Denisa ; Anděrová, Miroslava (advisor) ; Procházka, Jan (referee) ; Novák, Ondřej (referee)
NG2-glia proliferation and differentiation following CNS injuries Abstract NG2 glia display wide proliferation and differentiation potential under physiological and pathological conditions. They are very well known as precursors of oligodendrocytes, however, following central nervous system (CNS) injury they play an important role in regeneration. For this reason, we examined these features following different types of brain disorders such as focal cerebral ischemia (FCI), cortical stab wound (SW), and demyelination (DEMY) in young (3-months-old) mice, in which NG2 glia are labeled by tdTomato under the Cspg4 promoter. In the case of FCI, the factor of age was also studied using 18-months-old mice. To address these issues, we employed many techniques on tissue/cellular levels, such as single-cell RT- qPCR, single-cell/bulk RNA-sequencing, immunohistochemistry, and the patch-clamp technique in situ. First, such approach enabled us to distinguish two main populations (NG2 glia, oligodendrocytes), each of them comprising four distinct subpopulations. Next, the expression profiling revealed that a subpopulation of NG2 glia expressing GFAP, a marker of reactive astrocytes, appears transiently after FCI. However, following less severe injury, namely the cortical SW and DEMY, subpopulations mirroring different...
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Vlastnosti a regulace objemově-závislých aniontových kanálů u astrocytů.
Harantová, Lenka ; Anděrová, Miroslava (advisor) ; Moravec, Jan (referee)
Animal cells need to preserve constant volume in the face of osmolarity perturbations to function properly. To regain their original volume after hyposmotically induced swelling, most cell types extrude intracellular electrolytes and organic osmolytes accompanied by osmotically driven water. This process is termed regulatory volume decrease and is ensured by various ion channels and transporters. Recently, much attention has been focused on the ubiquitous volume-regulated anion channels activated by cell swelling. VRACs are moderately outwardly rectifying with intermediary conductance, permeable to inorganic anions and organic osmolytes and sensitive to broad-spectrum anion channels blockers. Functional properties of VRACs in astrocytes are particularly interesting, because many brain pathologies, such as ischemia, traumatic brain injury or hyponatremia, are associated with marked astrocytic swelling and VRACs could thus constitute a possible target for therapy of cerebral edema. Furthermore, VRACs are thought to play a role in cell cycle progression, cell migration, apoptosis and intercellular communication. Despite intensive research, VRACs molecular identity and mechanism of their activation and regulation are still unclear. This work summarizes known facts about VRACs, accentuating their...
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Neurogeneze a gliogeneze v dospělém mozku po ischemickém poškození
Honsa, Pavel ; Anděrová, Miroslava (advisor) ; Vyklický, Ladislav (referee) ; Mazurová, Yvona (referee)
Ischemic brain injury belongs to the most common cause of death in the developed countries. High complexity of this disorder significantly slows and limits the possible treatment. Currently, there is only one treatment available - the application of the thrombolytic, tissue plasminogen activator, while thousands of other drugs failed during clinical testing. Great expectations were seen in the stroke treatment employing neural stem cells obtained from several external sources; nevertheless, low survival rate, limited favorable outcome and enormous ethical issues complicate the application of such therapy. On the other hand, in the adult mammalian brain exist two endogenous processes - neurogenesis and gliogenesis. These processes need to be fully described and understood, in order to employ them as a source of new cells after injury. Therefore, this thesis focuses on the processes of adult neurogenesis and gliogenesis predominantly after ischemia. Adult neurogenesis and gliogenesis are processes, by which neurons or glial cells are generated from stem/progenitor cells. Both these processes are strongly influenced by brain injury; nevertheless, their contribution to regeneration after ischemia in the human brain is negligible. Here, we aimed to describe the role of polydendrocytes in the...
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Astrocyte volume changes during brain ischemic injury
Mikešová, Michaela ; Anděrová, Miroslava (advisor) ; Zemková, Hana (referee)
Brain ischemic injury is a complex of pathophysiological events following transient or permanent reduction of brain blood flow. It results in a disruption of neuronal and astrocytic physiological functions, long-term reduction of brain blood flow leads to the cell death. Number of recent studies is focused on astrocytes, which might play key roles in surviving cells, including neurons, during ischemic injury. Astrocytes provide many important functions, such as maintenance of ionic homeostasis, prevention of excitotoxicity, scavenging free radicals and others and thus astrocytes may dramatically swell during ischemic conditions and contribute notably to the development of cytotoxic edema. This thesis summarizes mechanisms possibly contributing to the astrocytic swelling during brain ischemic injury as well as methods used for studying astrocyte volume changes and their quantification. Since the brain edema dramatically complicates both course and treatment of ischemic injury, knowledge of mechanisms leading to astrocytic swelling and their volume regulation during ischemia/reperfusion might be used for developing new therapeutic approaches for the treatment of cerebral ischemia, mainly for reduction of negative impact of edema.
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Calcium signalling in glial cells in progress of Alzheimer disease
Waloschková, Eliška ; Anděrová, Miroslava (advisor) ; Maršáková, Lenka (referee)
Alzheimer's disease (AD) is a neurodegenerative disorder affecting the entire central nervous system including glial cells. The mechanisms of this disease are not yet entirely clear, although recent studies suggest that among the known hallmarks of AD, such as accumulation of amyloid β and hyperphosphorylated tau, dysregulation of intracellular calcium homeostasis is proposed to be a significant feature both in neurons and glial cells, namely astrocytes and microglia. Glial cells play an important role both in healthy brain and during AD progression. Their major functions, such as supporting neurons or maintaining synapses, are impaired during this disease. Recent findings suggest that aberrant glial calcium signaling activated during AD, could possibly promote the malfunction of these cells and increase their inflammatory response, thus affecting neurons and causing brain damage. It is likely, that the ongoing inflammation and the impaired calcium signaling affect one another, consequently enhancing the progression of AD.
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Induction of neurogenesis and gliogenesis after ischemic CNS injury - the role of Wnt signaling pathway
Koleničová, Denisa ; Anděrová, Miroslava (advisor) ; Nerandžič, Vladimír (referee)
This bachelor thesis is focused on ischemic injury of the central nervous system (CNS), one of the most frequent causes of death and disability in the world, and its possible treatment via the induction of neurogenesis. It consists of three different parts. In the first part, main neurogenic regions of the CNS, the subventricular zone and gyrus dentatus (GD) of the hippocampus, are described at the cellular, as well as molecular level. The second part is mainly dedicated to ischemic injury, to the global and focal brain ischemia. A particular chapter of this part describes changes in neurogenesis and gliogenesis after ischemic injury of the brain, changes in the induction of radial glial cells, proliferation and migration of neural progenitor cells and neuroblasts. In this chapter, we also describe the activation of astrocytes, microglia and NG2 glia (also known as polydendrocytes) after ischemic injury of the CNS. The last, third part of the bachelor thesis, is focused on signaling pathways, which significantly influence neurogenesis: Shh (Sonic hedgehog homolog), Notch and Wnt (Wingles/Integrated) signaling pathways. Special attention is devoted to the Wnt signaling pathway, which is an essential part of molecular mechanisms in nerve cells. Keywords: neurogenesis, gliogenesis, hippocampus, gyrus...
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Induction of neurogenesis and gliogenesis after ischemic injury of CNS
Filipová, Marcela ; Anděrová, Miroslava (advisor) ; Moravec, Jan (referee)
Ischemic injury (stroke) is one of the most common causes of death and disability in humans. Discovery of adult neurogenesis and possibilities to induce neurogenesis by cytokines brought new approaches and hopes in treating the ischemic lesion in future. The aim of this thesis is to describe cellular and molecular mechanisms of neurogenesis, mainly those discovered within last ten years. The first part describes generation of new neurons in the brain under physiological conditions, which is localized in the dentate gyrus of the hippocampus and in the subventricular zone of the lateral ventricles (i.e. in principal neurogenic regions). The second part describes animal models used for studying ischemic injury in rodents and moreover, it focuses on patophysiology of ischemic brain injury, which is accompanied by astrocyte and microglia activation. Further, the ischemia-induced neurogenesis is described in these two major neurogenic regions. Also the important role of NG2 glial cells in central nervous system (CNS) regeneration is pointed out. According to recent findings NG2 glia that are present in all regions of CNS might serve as a potential source of cells for directed differentiation into oligodendrocytes, astrocytes and even neurons during CNS repair/regeneration. In the last part of this work,...
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NMDA receptors in astrocytes: their role in ischemic brain injury
Valný, Martin ; Anděrová, Miroslava (advisor) ; Hock, Miroslav (referee)
Glutamate is the main excitatory neurotransmitter in the mammalian brain, and its transmission is responsible for higher brain functions, such as learning, memory and cognition. Glutamate action is mediated by variety of glutamate receptors, of which N-methyl-D-aspartate (NMDA) receptors are the most remarkable due to their high Ca2+ permeability and complex pharmacology. Despite the widespread expression of NMDA receptors in astroglial cells in different brain regions, they have been studied mostly in neurons. Therefore, the role of astroglial NMDA receptors under physiological conditions as well as in pathological states, such as cerebral ischemia, is not fully understood. The aim of this work was to elucidate the presence, composition and function of these receptors in astrocytes under physiological conditions and after focal cerebral ischemia. For this purpose, we used transgenic (GFAP/EGFP) mice, in which astrocytes express enhanced green fluorescent protein (EGFP) under the control of human promotor for glial fibrillary acidic protein (GFAP) enabling astrocyte isolation and their collection via fluorescence-activated cell sorting. We performed single-cell RT-qPCR analysis of astrocytes isolated from the cortex of adult mice. The analyzed cells were isolated from the uninjured brains of 50...
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Astrocyte volume changes in alpha-syntrophin deficient mice
Mikešová, Michaela ; Anděrová, Miroslava (advisor) ; Vargová, Lýdia (referee)
(EN) The formation of brain oedema, which accompanies ischemic or traumatic brain injuries, originates from a disruption of ionic/neurotransmitter homeostasis that leads to extracellular K+ elevation and neurotransmitter accumulation in the extracellular space. An increased uptake of these osmotically active substances, predominantly provided by astrocytes, is accompanied by intracellular water accumulation via aquaporin-4 (AQP4). Since it has been shown that the removal of perivascular AQP4 via the deletion of α- syntrophin, which is the protein responsible for anchoring AQP4 on the astrocytic membrane (Neely et al. 2001), delays oedema formation and K+ clearance (Amiry-Moghaddam et al. 2003), we aimed to elucidate how the alpha-syntrophin deletion affects astrocyte volume changes in the cortex during pathological states, such as hypoosmotic stress or oxygen- glucose deprivation (OGD), using three-dimensional (3D) confocal morphometry in situ. In order to visualize individual astrocytes that lack alpha-syntrophin, double transgenic mice (GFAP/EGFP/α-Syn-/- ) were generated by crossbreeding GFAP-EGFP mice with α- syntrophin knockout mice. 3D-confocal morphometry revealed that alpha-syntrophin deletion did not alter astrocyte swelling during hypoosmotic stress or their recovery in isotonic solution;...
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