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The role of stem and progenitor cells in regeneration of hematopoietic tissue
Faltusová, Kateřina ; Nečas, Emanuel (advisor) ; Hofer, Michal (referee) ; Filipp, Dominik (referee)
Tissue regeneration is a complex and highly orchestrated process dependent on cells with the potential to restore structures and functions and on controlling factors from the tissue microenvironment. Hematopoietic tissue has a high ability to regenerate, which is attributed to the presence of stem cells, but the regeneration of severely damaged adult tissue is still only partially understood. Hematopoietic tissue provides a unique opportunity to study tissue regeneration due to its well-established steady- state structure and function, easy accessibility, advanced research methods, and well-defined embryonic, fetal, and adult stages of development. Embryonic/fetal liver hematopoiesis and adult hematopoiesis recovering from damage share the needto expand populations of progenitors and stem cells in parallel with increasing production of mature blood cells. We analyzed adult hematopoiesis in mice subjected to a submyeloablative dose (6 Gy) of gamma radiation, in which only a few cells with reconstituting capacity survived. We targeted the period of regeneration characterized by the renewed massive production of mature blood cells and the ongoing expansion of immature hematopoietic cells. Cells from the top of the hematopoietic hierarchy, hematopoietic stem cells, and multipotent progenitors are almost missing...
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The role of mTOR signalling pathway in neural differentiation of stem cells
Šintáková, Kristýna ; Jendelová, Pavla (advisor) ; Dráber, Peter (referee)
Spinal cord injury is a very serious, complex, and life changing injury for which today's medicine still does not have an efficient treatment. It is only possible to mitigate the consequences of this injury and the pathological processes associated with it. Neural stem cell transplantation has immunosuppressive effects in the pathology of spinal cord injury and promotes regeneration. mTOR kinase is a member of the crucial intracellular PI3K/Akt/mTOR signalling pathway, making it a suitable target for therapeutic intervention and immunosuppressants such as rapamycin. mTOR signalling is important for neural stem cells and in the pathology of spinal cord injury. The aim of this study was to investigate the role of the mTOR pathway in differentiation of stem cells into neuronal phenotype. Rapamycin was applied to in vitro culture of neural progenitors. Immunocytochemistry and immunoblotting techniques were used to study the effect of this inhibition on the cell phenotype and on the activity of the mTOR pathway. Using the rat model of spinal cord injury in vivo, immunohistochemistry and immunoblotting techniques were used to evaluate the impact of rapamycin inhibition on the mTOR pathway, autophagy, and cytokine production by cells in the damaged tissue. The results show that the mTOR pathway plays role...
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Relevance of Tumor Infiltrating Lymphocytes as a Prognostic Factors at Patients With Portal Vein Embolisation (PVE) and Patinets With PVE and Administration of Autologous Stem Cells
Brůha, Jan ; Třeška, Vladislav (advisor) ; Gürlich, Robert (referee) ; Šubrt, Zdeněk (referee)
Relevance of tumor infiltrating lymphocytes as a prognostic factors in patients with portal vein embolisation (PVE) and patients with PVE and administration of autologous stem cells Background: low future liver remnant volume (FLRV) is the cause of why 75% of patients with colorectal liver metastases (CLM) are primarily inoperable. Portal vein embolisation (PVE) helps to increase FLRV and so increase the operability. But PVE fails in almost 40 % of patients. Usage of stem cells (SCs) could be the way how to support the effect of PVE. Currently, there are studies of interactions of the immune system and malignancies. We do not know about papers focused on relations of the immune system and CLM in patients treated by PVE. There were not described interactions of ABC transporters and CLM at patients after PVE was performed too. Aims: the aim of this dissertation was to verify the effect of PVE and intraportal administration of SCs on the growth of FLRV and progression of the CLM. Other aims were to evaluate the tumor infiltrating lymphocytes, ABCC10 and ABCC11 transportes in patients treated by surgery for CLM after PVE and their clinical relevances. Methods: intraportal administration of SCs after PVE and their effect was explored in a group of 63 patients (43 patients with PVE alone, 20 in the group PVE with...
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Liver Regeneration with aplication of hematopoetic stem cells after portal vein embolization in pacients with primary inoperative colorectal liver metastases
Fichtl, Jakub ; Třeška, Vladislav (advisor) ; Šnajdauf, Jiří (referee) ; Šubrt, Zdeněk (referee)
Introduction: The reason for the inability of performing the liver resection for colorectal carcinoma metastasis is usually insufficient remnant liver parenchyma after liver resection (future liver remnant volume - FLRV). The current standard method of increasing FLRV is the embolization of the branch of portal vein (portal vein embolization - PVE) on the side of the tumor, and then suspended after hypertrophy of the non-embolised lobe liver resection. Unfortunately, there are some patients who do not increase liver volume despite perfectly executed PVE. Besides that, FLRV occurs during the time necessary for hypertrophy progression of metastatic disease. Therefore, we are trying to find the appropriate way to encourage the growth of remaining liver parenchyma and accelerate hypertrophy of the contralateral liver lobe. From our previous experience (IGA MZ NS 10240), it is possible to be optimistic that there hope is the way of hematopoietic progenitor cells (HPC - adult stem cells) after previous PVE to non-embolised branches of the portal vein. These cells do not only accelerate liver regeneration, but are also able to improve its function (function of the liver) which is especially important for patients after neoadjuvant chemotherapy (steatohepatitis or steatofibrosis), and for patients with...
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Maternal and fetal microchimerism.
Hynková, Marie ; Tlapáková, Tereza (advisor) ; Pačes, Jan (referee)
Microchimerism is defined as the presence of small amount of genetically distinct cells in one individual. It most often arises during pregnancy and breastfeeding. It can also arise from blood transfusion or organ transplantation. During pregnancy there occurs a bidirectional migration of cells through the placenta between the mother and the fetus. Cells which are transferred during pregnancy and breastfeeding can persist in the offspring until adulthood. During breastfeeding, a big number of immune cells is transferred to the offspring via mother's milk. These cells protect the offspring against pathogens and are involved in modulation of its immune system. Fetal cells persist in mother organism even decades after giving birth and can have long-lasting effect on mother's health condition. These fetal cells can help to regenerate mother's damaged tissues, but they can also contribute to the development of serious autoimmune diseases.
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Immunomodulatory mechanisms of stem cells and their use for therapy of ocular disorders
Heřmánková, Barbora
Stem cell-based therapy represents a perspective approach for the treatment of many so far incurable diseases. Mesenchymal stem cells (MSC) are currently the most studied stem cells. They are able to differentiate into different cell types, to produce growth and trophic factors and can suppress the functions of cells of the immune system. During the study of the immunomodulatory properties of MSC, we focused on their effect on B cells. The mechanism of impact of interferon-γ (IFN-γ) on MSC and their effect on the production of interleukin 10 (IL-10) by B cells was analysed. We have demonstrated that MSC-treated with IFN-γ inhibit production of IL-10 by activated B cells via the cyclooxygenase-2 involving pathway. Due to their regenerative and immunomodulatory properties, MSC can be for treatment of many diseases. In this study we focused on the disease and damage of the eye. The limbal stem cells (LSC) are used for the treatment of damaged ocular surface, however their isolation is difficult and they can not be used in all cases of damage. Appropriate candidates in these cases are MSC. Therefore we have decided to compare the therapeutic potential of LSC and MSC isolated from bone marrow or adipose tissue. The study have shown that MSC isolated from bone marrow have a similar regenerative effect on...
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The effect of carbon nanostructures on human cell behavior and the role of fetal bovine serum in cell adhesion
Jannová, Martina
Graphene (G) and nanocrystalline diamond (NCD) are carbon allotropes and promising nanomaterials with an excellent combination of their properties, such as high mechanical strength, electrical and thermal conductivity, possibility of functionalization and very high surface area to volume ratio. For these reasons, G and NCD are employed next to electronics in biomedical applications, including implant coating, drug and gene delivery and biosensing. For a fundamental characterization of cell behavior on G and NCD, we studied osteoblast adhesion and proliferation on differently treated G and NCD. Generally, both G and NCD exhibited better properties for osteoblast cultivation than control tissue culture polystyrene. Better cell adhesion but lower cell proliferation were observed on NCD compared to G. The most surprising finding was that hydrophobic G with nanowrinkled topography enhanced cell proliferation extensively, in comparison to hydrophilic and flat G and both NCDs (hydrophobic and hydrophilic) with slightly higher roughness. Promoted cell proliferation enables faster cell colonization of G and NCD substrates, meaning faster new tissue formation which is beneficial in biomedical applications. Furthermore, it was shown that osteoblast adhesion was promoted in the initial absence of fetal bovine...
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Development of extracellular-matrix scaffolds for CNS repair
Výborný, Karel ; Kubinová, Šárka (advisor) ; Hampl, Aleš (referee) ; Vandrovcová, Marta (referee)
Výborný, Karel. Vývoj materiálů na bázi extracelulární matrix pro léčbu centrálního nervového systému. [Development of extracellular-matrix scaffolds for CNS repair]. Praha, 2020. 105 stran, 3 přílohy. Dizertační práce (Ph.D.). Univerzita Karlova, 2. lékařská fakulta, Ústav experimentální medicíny, AV ČR v.v.i. Abstract Brain and spinal cord injury are a serious traumata and despite intensive research, there is still no effective treatment for patients. One of the studied approaches is to use various biomaterials to repair the damaged neural tissue. The aim of this thesis is to study the regenerative and neurotrophic effects of injectable extracellular matrix (ECM) hydrogels prepared by decellularization from porcine tissue (brain, spinal cord, bladder) and human umbilical cord (UC) in reconstruction of damaged neural tissue of the brain and spinal cord in rats. We characterized ECM hydrogels in terms of mechanical and rheological properties, structure and composition, cell adhesion, migration and proliferation. In a model of spinal cord hemisection, we compared the regenerative effect of ECM hydrogels derived from porcine spinal cord and urinary bladder. We found no benefits of tissue specific ECM prepared from the tissue of neural origin in terms of its neurotrophic properties in vitro or in vivo....
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Use of corneal endothelium and amniotic membrane for transplantation purposes.
Šmeringaiová, Ingrida ; Jirsová, Kateřina (advisor) ; Netuková, Magdaléna (referee) ; Čejková, Jitka (referee)
Part I: Endothelial cells form the posterior layer of the cornea and are important for maintaining its transparency. Dysfunctional endothelium can only be restored by transplantation. The global shortage of donor corneas requires the search for alternative treatments. The preparation of the graft by tissue engineering methods is complicated by low proliferative capacity of endothelium. To date, no endothelium-specific marker has been defined and the existence of endothelial stem cells has not been confirmed yet. We have prepared a protocol for culturing endothelial cells from research-grade tissue - corneoscleral rims obtained after transplantation or corneas excluded from the transplant process. We monitored localization of selected proteins, including stem cell markers, in native tissue and in primary cell cultures. We prepared up to 6.4 cm2 of endothelium from one cornea/rim, which had cellular features comparable to the native endothelium. This approach can increase the amount of endothelium for research or transplantation purposes. Using indirect immunohistochemistry, we showed that none of the previously proposed endothelial molecular markers is specific for these cells. We detected the expression of stem cell markers throughout the endothelial layer. In the porcine cornea model, we monitored...
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Development of Material Based on Hyaluronic Acid s Hydrogels for Myocardial Regeneration
Kovářová, Lenka ; Kubala,, Lukáš (referee) ; Prokš,, Vladimír (referee) ; Pekař, Miloslav (advisor)
The thesis is focused on material development based on hyaluronic acid usable in regenerative medicine, especially for heart tissue regeneration after myocardial infarction. The object of the study is the oxidized form of hyaluronic acid (HA-Ox) and hydroxyphenyl derivative of HA (HA-TA). HA-Ox can be crosslinked with a bifunctional alkoxyamine POA and HA-TA undergoes an enzymatic reaction in the presence of hydrogen peroxide catalysed by horseradish peroxidase leading to gel formation. To describe the materials, chemical and physical properties, gelation kinetics and conditions of crosslinking reactions were studied. Hydrogels were characterized by mechanical and viscoelastic properties, degradability or stability in simulated body fluids. These hydrogels serve as scaffolds for the selected cell type. To promote cell adhesion and viability, an RGD sequence has been bonded to the structure of HA-TA. This resulting material is also compatible with selected applicators. Its viscosity and extrusion force are low enough to allow application with a catheter with a very small internal diameter. The applicability of the material through the supply tube to the hydrogel reservoir of the second SPREADS device showed good homogeneity, cell distribution and viability. Finally, the material was applied in vivo using these devices during a preclinical study.
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