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Impact of nanomaterials on mesenchymal stem cells and tissue regeneration
Echalar, Barbora ; Holáň, Vladimír (advisor) ; Hubálek Kalbáčová, Marie (referee) ; Buchtová, Marcela (referee)
Nanomaterials (NMs) are widely used in medicine for their antimicrobial properties. They are part of antibacterial coatings, creams, pharmaceutical vehicles or additives in drugs and other medical products. However, the impact of NMs on human organism is still not completely established. Nanoparticles (NPs) penetrate the cell membrane and enter to intracellular compartments including the nuclei. Different types of NPs could have various side effects on cell functions. These side effects include the damage of stem cells (SCs) or immune cells lead to slower regeneration and impaired wound healing. Therefore, the simultaneous application of NPs during SC therapy could decrease the therapeutic abilities of SCs. One type of SCs tested in clinical therapies nowadays are mesenchymal stem cells (MSCs). Therefore, we studied the impact of metal NPs (i.e. silver, copper oxide, zinc oxide and titanium dioxide) on characteristics and functional properties of mouse MSCs. Additionally, the effect of NPs on the expression of phenotypic markers, metabolic activity, differentiation potential, expression of genes for immunoregulatory molecules and on production of cytokines and growth factors was analyzed. We found that all types of tested NPs had a negative impact on the activity of MSCs and thus could alter tissue...
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Growth factors in cultivated meat production
Janoušek, Jiří ; Rozbeský, Daniel (advisor) ; Mašek, Jan (referee)
The rapid expansion of global meat production poses severe environmental and ethical challenges, necessitating alternatives with reduced impacts. Cultivated meat, produced from animal cells in vitro, emerges as a promising solution. However, common cell culturing techniques using fetal bovine serum (FBS) are not sufficiently scalable for this purpose. The critical role of growth factors in replacing FBS is emphasized, with a focus on summarizing existing literature regarding their importance and potential for economically viable mass adoption. The described growth factor requirements of various cultivated meat production relevant cell types, as well as the potential improvements to the production processes, activity, or stability of the most important growth factors identified, are analyzed. Finally, alternative approaches to the use of growth factors are briefly explored. This work should provide a comprehensive overview of recombinant growth factor production and optimization as it pertains to the area of cultivated meat.
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Mechanisms of Vascularization in Skin Tissue Engineering
Futóová, Terézia ; Brož, Antonín (advisor) ; Šuca, Hubert (referee)
Tissue engineering is a multidisciplinary field dealing with the fabrication of artificial tissue substitutes for regenerative medicine. Current regenerative medicine uses various types of tissue grafts, which have different advantages and disadvantages depending on their origin, such as insufficient amount of replacement tissue when using autologous grafts or immunogenicity of allogeneic or xenogeneic grafts. An alternative could be artificial tissue replacement. Artificial tissue constructs may consist of a non-living matrix and a cellular component. The cellular component may remodel the construct, form a functional part of the construct, or help integrate the construct into the host body. A significant problem in the formation of such replacements is sufficient vascularization. It is essential to keep cells in larger tissue constructs alive. Vascularization can be enhanced by the addition of vascular endothelial cells that can form capillaries independently within the construct. Vascular formation can also be aided by angiogenic growth factors by their direct application to the construct or by their formation, e.g. in stem cells cultured in the construct. Another approach is 3D bioprinting, allowing direct placement of specific cell types, growth factors or biomaterials in the construct. This...
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Smart biomaterials for connective tissues regeneration
Hefka Blahnová, Veronika ; Filová, Eva (advisor) ; Maxová, Hana (referee) ; Motlík, Jan (referee)
Connective tissues are characterized by significant volume of extracellular matrix. Their main role is to provide a mechanical support and protection to other body organs. This thesis is focused on regeneration of bone, cartilage and osteochondral defect. In the experimental part we observed viability and differentiation of human mesenchymal stem cells. In vitro we evaluated the potential of PCL scaffold with addition of growth factors, bone xenograft with biomimetic peptides, collagen I composite with bioceramics and a titanium alloy with nanostructured surface. During following in vivo study, we implanted a cell-free scaffold made of PCL, calcium phosphate and IGF-1, bFGF, TGFβ1 and BMP-2 to osteochondral defect. Unfortunately, addition of growth factors resulted in pathological inflammatory process despite clear beneficial effect in vitro. Likewise, the biomimetic peptide sequences promoted osteogenic differentiation of human mesenchymal stem cells. Addition of certain bioceramics influenced the scaffold morphology in the manner of pore size. However, we did not observe any effect of the surface characteristics on cell behavior. The cells were influenced rather by certain material. On the other hand, surface modification of titanium scaffold by anodic oxidation revealed that the most suitable...
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Changes in placental angiogenesis and their impact on fetal intrauterine growth restriction
Kudějová, Alena ; Švandová, Ivana (advisor) ; Magner, Martin (referee)
Any pathological interference with normal vascular development of placenta may have a critical impact on fetal growth and development. The proliferation and differentiation of several cell types play a very important role in the vascular system of placenta. The main factors taking part in the vascular development of placenta include cell elements (e.g. trofoblast, stromal chorion cells, haemangiogenic progenitors), the extracellular matrix, growth factors and cytokines (e.g. VEGF, PlGF, Ang-1,2 and bFGF). The extrinsic factors may also influence the partial oxygen pressure, nutritients availability, and/or the blood perfusion in placenta. Placental ischaemia leading to the worsening of uteroplacentar perfusion is the most common cause of the intrauterine growth retardation (IUGR). The IUGR development is then the result of insufficient prolongation, branching, and dilatation of capillary loops during the formation of terminal villi. Published studies focusing on growth factors in placentas from physiological pregnancies and pregnancies with IUGR do not give clear results. This BSc. Thesis is a review focused on up-to-date-known data concerning changes in placental angiogenesis and their impact on IUGR. Key words: placenta, angiogenesis, IUGR, pregnancy
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Preparation, characterization and testing of blood derivatives for applications in regenerative medicine
Sovková, Věra ; Amler, Evžen (advisor) ; Kolářová, Hana (referee) ; Vránová, Jana (referee)
Platelet products can be used, thanks to the broad range of bioactive molecules, either as a supplement for cell cultering in vitro alone or for development of cell- or cell-free scaffolds in diverse fields in regenerative medicine. The aim of this study was to prepare several types of platelet products. The concentration of selected molecules were observed. Subsequently, these products were tested with cell cultures in vitro alone or in combination with nanofibres scaffolds prepared by electrospinning or centrifugal spinning. It was found out, that platelets products contains chemokine RANTES and growth factor PDGF in the highest concentrations. It was further discovered the content of pro and antiinflammatory in terleukins and other growth factors. Platelet lysat in concentration 7% is sufficient to replace FBS in keratinocytes and fibroblasts cultures. In the other experiments, platelets in different concentrations were adhered to the scaffolds prepared by electrospinning and centrifugal spinning. Thus prepared scaffolds promote the proliferation and viability of all tested cell types in dose-dependent manner. In the last experiment, the individual components of platelet concentrate were separated and characterized. Their effect to the cell culture were tested. It was examinated the synergic...
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The role of growth factors and cytokines in regulation of mammalian oocyte maturation
Marcollová, Kateřina ; Procházka, Radek (advisor) ; Vávra, Jiří (referee)
Reproduction is key for species survival. Oocytes are blocked in prophase I of meiosis in follicle. Gonadotropin stimulation leads to follicle growth and competent oocyte. First polar body extrusion is a sign of meiotic division resumption. Oocyte is blocked at prophase I by a high cAMP level, activating PKA, that inhibits MPF. CNP binds to NPR2, that stimulates the generation of cGMP in granulosa cells, which diffuses to oocyte where it competes with cAMP on PDE3A. LH and FSH released from pituitary gland initiates signaling cascade restoring meiotic division. Synthesis of AREG, EREG and BTC activates EGFR and MAPK pathways in somatic cells which contribute to reduction of gap junction permeability, lowering NPR2 activity, that leads to reduction of cGMP production and transport. Less cGMP diffuses to oocytes, which leads to PDE3A activation and cAMP hydrolysis. This causes MPF activation and meiotic division resumption. MAPK together with PI3K participate on increasing production of hyaluronic acid, thereby cumulus cells expansion and oocyte ovulation. Oocyte maturation and growth is improved with utilization of in vitro technics and oocyte cultivation with growth factors and cytokines. FLI medium, combining FGF2, LIF and IGF-1, increases oocyte cultivation success and blastocyst quality of farm...
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Preparation, characterization and testing of blood derivatives for applications in regenerative medicine
Sovková, Věra ; Amler, Evžen (advisor) ; Kolářová, Hana (referee) ; Vránová, Jana (referee)
Platelet products can be used, thanks to the broad range of bioactive molecules, either as a supplement for cell cultering in vitro alone or for development of cell- or cell-free scaffolds in diverse fields in regenerative medicine. The aim of this study was to prepare several types of platelet products. The concentration of selected molecules were observed. Subsequently, these products were tested with cell cultures in vitro alone or in combination with nanofibres scaffolds prepared by electrospinning or centrifugal spinning. It was found out, that platelets products contains chemokine RANTES and growth factor PDGF in the highest concentrations. It was further discovered the content of pro and antiinflammatory in terleukins and other growth factors. Platelet lysat in concentration 7% is sufficient to replace FBS in keratinocytes and fibroblasts cultures. In the other experiments, platelets in different concentrations were adhered to the scaffolds prepared by electrospinning and centrifugal spinning. Thus prepared scaffolds promote the proliferation and viability of all tested cell types in dose-dependent manner. In the last experiment, the individual components of platelet concentrate were separated and characterized. Their effect to the cell culture were tested. It was examinated the synergic...
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Growth factors and other bioactive substances for osteogenic differentiation of mesenchymal stem cells
Blahnová, Veronika ; Rampichová, Michala (advisor) ; Vandrovcová, Marta (referee)
The main function of mesenchymal stem cells in the body is to facilitate the restoration and regeneration of damaged tissues. They are known for the ability to differentiate into tissue originating from the mesoderm, which among others includes connective tissues. Due to this feature are MSCs being intensively examined. Different directions of differentiation can be induced by treatment of specific polypeptides, so called growth factors. In the field of tissue engineering are growth factors used to induce and accelerate the healing processes. They may be incorporated into the nanofiber carrier which is inserted into the site of injury. Cells in this area would thus be stimulated by surrounding 3D microenvironment. At the same time the scaffold provides a supply of growth factors which are able to affect metabolism, motility and differentiation of present cells. In order to induce osteogenic differentiation of human MSCs the following bioactive substances were used: TGF-β, bFGF, HGF, IGF-1, VEGF and the BMP-2 and the organic acid taurine. During 21 days lasting experiments, were these molecules added to the medium in various combinations and in the case of taurine also at two different concentrations. Cells were cultured on plastic. The best effect on cellular metabolism of MSCs, evaluated by MTS...
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