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Title: Internalisation and cytotoxicity of alkyl-capped silicon quantum dots (SiDQs) in various mammalian cell lines
Author: Phatvej, Wipaporn
ISNI:       0000 0004 5924 3736
Awarding Body: Newcastle University
Current Institution: University of Newcastle upon Tyne
Date of Award: 2015
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The increasing application of nanoparticles in medicine, particularly in imaging and therapy, is heavily reliant on fundamental understanding of their unique physicochemical properties. This thesis investigates the interactions that occur between nanoparticles and human cells, specifically their toxicity and the mechanism of their internalisation. Alkyl-capped silicon quantum dots (SiQDs) were chosen for this investigation because their physical properties have been well-characterised and they have very bright luminescence. These properties facilitate their detection inside cells by fluorescence microscopy and flow cytometry. Toxicity and uptake mechanisms for SiQDs in human cells were investigated. The intestinal cell line CACO-2 was used as a model for ingestion of nanoparticles. Uptake and accumulation of SiQS in CACO-2 cells was demonstrated by epi-fluorescence microscopy and confocal fluorescence microscopy. Cytotoxicity in CACO-2 cells was studied by oxidative stress measurement using a intracellular dye method (H2DCFDA) (ROS assay), cell viability determination (MTT and ATP assays) and DNA damage measurement (Comet assay). Exposure of CACO-2 to SiQDs resulted in low cytotoxicity with regard to cell viability and effects on ATP production. SiQDs did not induce intracellular ROS production or DNA strand breaks. Over time periods up to 14 days, SiQDs showed no evidence of acute or chronic cytotoxicity. Accumulation of SiQDs inside a selection of human cell lines (CACO-2, HeLa, HepG2, and Huh7) was studied in detail using flow cytometry. SiQDs were internalised by all four cell lines. The highest levels of accumulation were seen with HepG2 and HuH7 cells. Some evidence for a role for caveolin 2 in this process in HuH7 was obtained using inhibitors and by gene expression analysis. The reason why HepG2 cells showed the highest accumulation of SiQDs remains unclear as caveolin expression measured by quantitative reverse transcriptase-PCR in these cells is very low. Further work on a possible role for the protein clathrin in the endocytosis process in these cells is needed as the inhibitor used to investigate this was toxic to the cells and firm conclusions could not be obtained. iii In summary, SiQDs are a highly promising alternative to heavy-metal based quantum dots and as a nanoparticle model to study mechanisms of uptake. SiQDs appear non-toxic in CACO-2 cells. However, the internalisation of SiQDs is cell-line dependent and further studies on their toxicity in cells such as HuH7 which show high levels of internalisation and cells relevant to immune responses such as macrophages are needed.
Supervisor: Not available Sponsor: Royal Thai Government ; Ministry of Science (MOST)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available