Browsing Ph.D. Chemistry by Subject "Microscopy"
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Probes of tocopherol biochemistry: fluorophores, imaging agents, and fake antioxidantsThe body has many defence systems against reactive radical species, but none are as crucial in the protection of lipid membranes as vitamin E. As a result of a selection process mediated by the α-tocopherol transfer protein (α-TTP), α-tocopherol is the only form of vitamin E retained in the body. This chaperon protein has been well studied because of its role in vitamin E transport. Furthermore, malfunctions of α-TTP cause vitamin E deficiency leading to ataxia and other neurodegenerative disease. Protection of neuronal tissue is critical and is reflected in the high retention of α-tocopherol in the central nervous system. Neuronal tissues receive α tocopherol from astrocytes, cells that are linked to hepatic tissue and able to express α-TTP, however the exact path of delivery between these cells is still unclear. A technique called fluorescent microscopy allows the tracking of fluorescent molecules in cells to find their location and interactions with other parts of the cell. The focus of this study is the synthesis of a fluorescent tocopherol analogue with a long absorption wavelength, high photostability, and that binds selectively to α-TTP with high affinity. Most health benefits associated with vitamin E consumption are based on its capability to inhibit lipid peroxidation in cell membranes by scavenging reactive oxygen species (ROS). Oxidative damage in membranes puts cells in a “stressful” state, activating signalling events that trigger apoptosis. Vitamin E down-regulates apoptotic functions like inflammation, macrophage activation and cell arrest in a stressed state, returning the cell back to normal functioning. At the same time, vitamin E has a preventive effect for atherosclerosis, Alzheimer’s and cancer. With the deeper understanding of cell signalling processes associated with vitamin E the question arose whether protein interactions or the ROS scavenging is responsible for cell survival. To test this hypothesis, a non-antioxidant but α-TTP binding tocopherol analogue was synthesized and administered into oxidatively stressed, α-TTP deficient cells. If the cells were unable to restore homeostasis and stop apoptosis with the new molecule, this would suggest that the antioxidant function of α-tocopherol is the reason for survival. Cancer is regarded as one of the most detrimental diseases with a high mortality rate. One key aspect in medical research is the increased drug specificity towards targeting cancer. Chemotherapy applies cytotoxic compounds, which weaken the immune system because both malignant and healthy cells are destroyed. The specificity of the anti-cancer drugs are enhanced when encapsulated into liposomes that bear target-directing molecules such as antibodies which recognize cancer cell specific antigens on the cell membrane. The question remains if the encapsulated drug reaches the cancer or not. Magnetic resonance imaging (MRI) and computed tomography (CT) are used to find malignant tissue in the body. CT imaging uses highly charged X-ray particles to scan the patient, possibly having damaging cytotoxic effects. Obtaining MRI results require the use of contrast agents to enhance the quality of images. These agents are based on transition metals, which potentially have chronic toxicity when retained in the body. Alternatively short-lived radiotracers that emit a γ-photon upon positron decay are used through a process called positron emission tomography (PET). Rapid decay times make the use of PET a less toxic alternative, however the decay products might be toxic to the cell. For this reason a vitamin E based PET agent was created, which produces naturally safe decay products based on known metabolites of vitamin E, useful to track liposomal delivery of chemotherapeutic agents. This work describes the non-radioactive synthetic procedures towards a variety of vitamin E PET analogues. The cytotoxicity of the most promising vitamin E PET tracer was evaluated along with its synthetic byproducts.