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Use of cell cultures to test cytotoxicity of natural substances
Blažková, Veronika ; Skoumalová, Petra (referee) ; Márová, Ivana (advisor)
This thesis is aimed on study of cell cultures, primarily on introduction of techniques for cultivation of human cells and optimization of their usage in cytotoxicity tests. The thesis is aimed preferably on natural substances and liposomes that are suitable to be used in cosmetics. Theoretical part contains literature review focused on overview of cell cultures, appropriate cultivation conditions for in vitro testing and summary of cytotoxicity tests. Experimental part contains characteristics of natural extracts with antimicrobial effect in terms of content of polyphenols, flavonoids and antioxidant activity. Further, these extracts were encapsulated into the liposomes. Encapsulation effectivity was determined. Distribution of particles size by dynamic light scattering and finally also stability of prepared particles were also determined. Long-term stability in water and gel environment was tested too. This thesis also contains study of optimization of cell culture of human skin keratinocytes cultivation and use of those cells for testing cytotoxicity of selected natural substances by MTT method.
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Studium dynamiky živých lidských buněk v odezvě na běžné kontaminanty vod s cílem vývoje citlivého specifického analytického postupu
KOŠEK, Jan
The bachelor thesis is devoted to testing the toxicity of chemical substances. In the current chemical and pharmaceutical industry, thousands of new substances are produced annually, which brings about the need to ensure sufficient consumer safety and environmental protection. In vitro testing is becoming an increasingly important research tool in this issue. This work verifies the theoretical foundations and proposes an algorithm developed by Lonhus et al. from the Laboratory of experimental complex systems in Nové Hrady and its possible future applications in in vitro testing. This new approach uses the principles of microrheology and tracks Speed-Up Robust Features (SURF) in a digital image and converts them into vectors of trajectories of intracellular mass particles. Microrheology is a relatively new field of research in life sciences and deals with the study of flow and deformation of materials at the microscopic level. The algorithm separates two main components of the movements inside the cells: direct movement of organelles as larger particles and random Brownian motion. The advantage of this method is its potential of easy use and sensitivity. The method makes it possible to obtain important primary information about the effect of chemical substances on cell cultures.
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Study of the influence of natural drugs on estrogen receptor and confirmation of their effect in cell cultures
Elbelová, Jana ; Carazo Fernández, Alejandro (advisor) ; Pourová, Jana (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Jana Elbelová Supervisor: PharmDr. Alejandro Carazo, Ph.D. Title of the diploma thesis: Study of the influence of natural drugs on estrogen receptor and confirmation of their influence in cell culture Estrogens are one of the main reproductive hormones in women. Their function in the human organism is mediated by the estrogen receptor (ER). Estrogens are involved in important physiological processes such as reproduction, bone metabolism and cardiovascular function. Long term elevated levels of these hormones may also lead to breast cancer development. Phytoestrogens are natural ligands of estrogen receptor widely found in dietary products. These compounds have beneficial properties in treatment of menopausal symptoms thank to their estrogenic activities but are able to function as antiestrogenic agents too. In addition, they may play a role in the development of other illnesses (breast cancer etc.). Therefore, they can be considered as a potential pharmacological tool for the treatment of estrogen-related conditions. Within this master thesis, a total of six compounds, two phytoestrogens and four of their more relevant metabolites, were chosen (genistein, daidzein, S equol, O...
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Experimental model systems to study small DNA viral infection
Bučková, Alžbeta ; Saláková, Martina (advisor) ; Horníková, Lenka (referee)
Merkel cell polyomavirus (MCPyV) and Human papillomavirus 16 (HPV 16) are members of small tumour DNA viruses Polyomaviridae and Papillomaviridae, which represent increasing risk for humans resulting from their oncogenic potential. After the acquisition HPV 16 and MCPyV are able to persist for long term in a form of asymptomatic infection, while the aggressive disease is mostly being cleared by the host immune system. Integration of viral genome into the host DNA causes cell transformation resulting in rare but fatal skin carcinomas and epithelial lesions of anogenital tract, head and oropharynx, that may progress into malignant tumours. Their mechanisms of immune system evasion and complete life cycles are not fully understood to this day which highlights some of the reasons why continuing research in this field is of importance. The aim of this thesis is to review model systems used to study infection of MCPyV and HPV 16 in vitro and in vivo. Key words: Papillomaviruses, polyomaviruses, virus-like particles, pseudoparticles, animal models, cell culture, human papillomavirus 16, Merkel cell polyomavirus, HPV 16, MCPyV
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Adhesion, growth and differentiation of skin cells on nanofibrous polymer membranes
Pajorová, Júlia ; Bačáková, Lucie (advisor) ; Eckhardt, Adam (referee)
Our study contributes to the tissue engineering, mainly to the construction of appropriate scaffolds for regeneration of damaged skin. Simultaneously, it brings valuable insights for basic research in the field of molecular mechanisms of adhesion, proliferation and phenotypic maturation of cells and the control of the cell behavior through the cell extracellular matrix (ECM), represented by synthetic nanofibrous material. Nanofibrous polylactic-co-glycolic acid (PLGA) membranes were prepared by needle-less electrospinning technology. These membranes were further modified with cell adhesion-mediating biomolecules, e.g. collagen, fibronectin and fibrin in order to increase their affinity to colonizing cells. Adhesion, growth and differentiation of keratinocytes (HaCaT) and fibroblasts, i.e. major cell types of epidermis and dermis, were evaluated on these nanofibrous membranes. The results show that the membrane modification using fibrin structures improved adhesion and proliferation of human dermal fibroblasts. The collagen structure on the surface of membranes improved the adhesion and proliferation of human HaCaT keratinocytes. Furthermore, fibrin structure stimulated fibroblasts to produce collagen, which is a major component of ECM in the natural skin dermis. Fibronectin enhanced cell attachment...
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Preparation and characterization of synthetic mRNA coding for pancreatic transcription factors
Loukotová, Šárka ; Hodek, Petr (advisor) ; Jirák, Daniel (referee)
Diabetes mellitus type I is severe autoimmune disease which is caused by destruction of insulin-producing β-cells in pancreas. Diabetic patients are dependent on external usage of insulin during their whole life. Nowadays the only treatment of diabetes type I is transplantation of entire pancreas or isolated Langerhans islets. Due to the fact that this kind of treatment is very demanding and limited availability of suitable donors, the researchers are intensively working on development of new alternative ways how to produce the insulin-producing cells. One of the possible approaches on producing insulin-positive cells is transdifferentiation of pancreatic exocrine cells via transcription factors. In this diploma thesis, the transdifferentiation of exocrine cells AR42J was carried out with in vitro synthesized mRNA encoding transcription factors Pdx1, Ngn3 and MafA. The primary mRNA structure was optimized in order to prepare highly stable mRNA which is correctly translated into the protein. The main stabilizing elements in mRNA structure include 3' and 5' untranslated region derived from highly stable β-globin mRNA. In order to verify the function of synthetic mRNA the immunofluorescence staining of transcription factors has been investigated. Synthetic mRNAs encoding transcription factors Pdx1,...
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Use of cell cultures to test cytotoxicity of natural substances
Blažková, Veronika ; Skoumalová, Petra (referee) ; Márová, Ivana (advisor)
This thesis is aimed on study of cell cultures, primarily on introduction of techniques for cultivation of human cells and optimization of their usage in cytotoxicity tests. The thesis is aimed preferably on natural substances and liposomes that are suitable to be used in cosmetics. Theoretical part contains literature review focused on overview of cell cultures, appropriate cultivation conditions for in vitro testing and summary of cytotoxicity tests. Experimental part contains characteristics of natural extracts with antimicrobial effect in terms of content of polyphenols, flavonoids and antioxidant activity. Further, these extracts were encapsulated into the liposomes. Encapsulation effectivity was determined. Distribution of particles size by dynamic light scattering and finally also stability of prepared particles were also determined. Long-term stability in water and gel environment was tested too. This thesis also contains study of optimization of cell culture of human skin keratinocytes cultivation and use of those cells for testing cytotoxicity of selected natural substances by MTT method.
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