/library/oar/handle/123456789/136308 2025-12-26T02:23:06Z 2025-12-26T02:23:06Z /library/oar/handle/123456789/141274 2025-11-13T13:03:38Z 2025-01-01T00:00:00Z

Title: Effect Of VPA and TGF-β on epithelial to mesenchymal transition in lung cancer Abstract: Lung cancer is the one of the most diagnosed cancer in the world, as well as the leading cause of death amongst cancer patients. Epithelial to mesenchymal transition (EMT) is the process by which epithelial cells acquire a mesenchymal phenotype, gaining characteristics such as increased migratory capacity, invasiveness, and resistance to treatment. This process is important in cancer as it enables tumour cells to become more mobile and invasive, leading to metastasis. The main aim of this thesis was to investigate how two molecules; valproic acid (VPA) and transforming growth factor-β (TGF-β) affect epithelial–mesenchymal transition (EMT) in lung cancer cells, specifically in H460 cells, which are classified as large cell lung cancer (LCLC) cells. VPA is a histone deacetylase inhibitor (HDACI), which has been shown to either inhibit or activate EMT based on the specific cell line and concentration. TGF-β plays an important role in cell proliferation, differentiation, and wound healing. Cell proliferation and cytotoxicity of VPA and TGF-β were tested on H460 cells by MTT assays. Moreover, gene analysis of mesenchymal biomarkers including FN1, SNAI1, VIM, and ZEB1 was carried out on cells treated with VPA. Results showed that both these molecules demonstrated a dose-dependent influence on H460 cell viability, as they both exerted a cytotoxic effect at higher concentrations. Gene expression analysis showed that VPA treatment affects EMT markers in a time and dose dependent manner, where mesenchymal markers were downregulated at earlier time-points, and generally upregulated at a higher concentration and later time-points. Further work includes performing mesenchymal-associated gene expression analysis on TGF-β-treated cells. Description: B.Sc. (Hons) Med. Biocem.(Melit.)

2025-01-01T00:00:00Z /library/oar/handle/123456789/141273 2025-11-13T13:01:34Z 2025-01-01T00:00:00Z

Title: Characterizing the dynamics of cell death and surface marker changes in tolerized human monocytes treated with epigenetic modifiers Abstract: Sepsis is a life-threatening condition which can be characterised by dysregulated host immune responses to infection, this disease leads to significant global morbidity and mortality with over 48.9 million cases and 11 million deaths annually. The work done in this dissertation aims to investigate the effects of two epigenetic modifiers, known as squalene and butyrate on human monocytes that are treated with LPS to induce the immune tolerance phenotype. Using PBMCs obtained from healthy donors, monocytes were isolated via MACS and treated with LPS (simulating infection) and varying concentrations of squalene and butyrate to determine the ideal working concentration. Once the working concentrations were established, the cells were treated with LPS, squalene and butyrate and left for 24 and 4-hour periods. FACS analysis was then carried out using 7AAD to assess for cell death and CD14, CD206 and HLA-DR to assess cell surface marker changes. ELISAs were carried out in order to assess the levels of TNF-α. The results showed that both squalene and butyrate exhibited cytoprotective properties, which aligns with the current literature. Cell viability was also retained with both squalene and butyrate showcasing significantly lower levels of cell death when compared to the LPS control. Additionally, Cell surface marker changes were also noted for both treatments, indicating that the cells retained their function as well. Description: B.Sc. (Hons) Med. Biocem.(Melit.)

2025-01-01T00:00:00Z /library/oar/handle/123456789/141272 2025-11-13T12:55:29Z 2025-01-01T00:00:00Z

Title: Impact of low-dose glyphosate and aminomethylphosphonic acid exposure on colorectal cancer sensitivity to 5-fluorouracil Abstract: Colorectal cancer (CRC) may arise from a number of environmental pollutants from various origins including agriculture. Glyphosate is a wide-spectrum herbicide used worldwide. Its negative effects on the human body have been identified when exposure is in high doses and for short spans of time, yet little is known about its chronic low-dose exposure. Its main metabolite by the soil microbiome is aminomethylphosphonic acid (AMPA), the literary knowledge of which regarding its effects in the human body is lacking. In this study, the primary objectives are to identify a dose of glyphosate and AMPA that is sub-lethal yet effective in the CRC cell lines HCT116, DLD-1, and LoVo. The cells will then be exposed to this dose for two weeks and treated with concentrations of 5-fluorouracil (5-FU), in order to compare their viability to untreated cells. Lastly, protein expression analyses will be carried out using Western blotting and Enzyme linked immunosorbent assay (ELISA) to identify pathway mechanisms that may or may not have been activated. A sub-lethal dose of 5 μM glyphosate and AMPA was selected to be used for chronic exposure. Following chronic exposure, overall changes in viability were not significant between treated and untreated cells, however, notable changes were observed in glyphosate-treated cells at 50 μM 5- FU and in AMPA-treated cells at 5 μM 5-FU. Western blots were elusive, showing unwanted variations. ELISA outputs revealed a decrease in activated Epidermal Growth Factor Receptor (EGFR) in treated cells, indicating that EGFR downstream signalling pathways may have not been activated. Further research is needed to validate the reliability of the obtained results and to identify other possible mechanisms that respond to glyphosate and AMPA at low doses through protein expression analysis. Description: B.Sc. (Hons) Med. Biocem.(Melit.)

2025-01-01T00:00:00Z /library/oar/handle/123456789/141271 2025-11-13T12:53:46Z 2025-01-01T00:00:00Z

Title: Metal specificity in superoxide dismutases Abstract: Superoxide dismutases (SODs) are essential metalloenzymes that protect cells from oxidative stress by catalysing the dismutation of superoxide radicals. While iron and manganese SODs are structurally similar, they exhibit strict metal specificity, and the precise determinants of this selectivity remain incompletely understood (Dudev, 2022). This study aimed to investigate the molecular basis of metal specificity in FeSOD isolated from Escherichia coli by sequentially introducing mutations within and near its active site. Building on the established FeSOD[dblDGF] mutant which included five substitutions (Q69G, A141Q, G142D, A68G, Y76F), an additional V70H mutation was introduced to create FeSOD[dblDGF_VH]. The mutant was expressed in E. coli under iron- and manganese-supplemented conditions, purified via immobilized metal affinity chromatography, and characterized through BCA assay, native PAGE with NBT staining, and Fe content analysis. The results demonstrated that FeSOD[dblDGF_VH] displayed higher relative activity when grown in manganese media, indicating a partial shift in metal preference, although overall activity remained lower than wild-type. Iron assays further suggested reduced iron incorporation in manganese-grown samples, supporting the proposed change in metal specificity. These findings highlight the critical role of active site residues in modulating metal selectivity and provide a possible foundation for further research in the V70H mutation. Description: B.Sc. (Hons) Med. Biocem.(Melit.)

2025-01-01T00:00:00Z