https://www.um.edu.mt/library/oar/handle/123456789/50033 Wed, 08 Apr 2026 14:31:02 GMT 2026-04-08T14:31:02Z https://www.um.edu.mt/library/oar/handle/123456789/50087 Title: Functional analysis of rare polymorphisms within the Aryl Hydrocarbon receptor (AHR) gene in pituitary adenomas. Abstract: The pituitary gland is integral in hormone secretion and regulation. Pituitary adenomas (PA) are the most frequent pituitary neoplasms, however molecular pathogenesis is largely unknown. The AHR is a ligand-activated transcription factor that regulates expression of various genes that mediate cellular response against xenobiotics, and is also involved in other physiological and pathological processes. Several AHR variants, particularly the Arg554Lys (rs2066853) and Val570Ile (rs4986826) have raised interest due to their location in ex on 10 of the AHR gene, which codes for the transactivation domain (TAD). The TAD domain plays a critical role in mediating transactivation activity of dioxin-responsive genes via recruitment of co-activators, hence suggesting that SNPs occurring within this region should interfere with AHR target gene expression. However, their exact functional role has not been established yet due to inconsistent results from different studies. Studies suggest that these mutations increase risk of developing P A, however functional analysis of these SNPs in a pituitary setting has never been carried out. In this research study, the two AHR variants were introduced in the wildtype AHR expression plasmid by site-directed mutagenesis (SDM). The wildtype and mutants AHR were introduced in GH3 cells by magnetofaction and were treated with 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). Functional analysis of transfected GH3 cells treated with TCDD was carried out using luciferase assays and real-time PCR to detect and quantify AHR-transcriptional activity. Cell proliferation of transfected and TCDD treated GH3 cells was measured using the MTT assay. In the absence and presence of low TCDD concentration, over-expression of AHR and AHR mutants did not affect the proliferative capacity of GH3 cells. Gene expression analysis and quantification analysis of AHR-target genes suggested that these AHR mutants might interfere with AHR target gene expression. However further studies are required to elucidate the precise mechanisms. Genotyping of the Arg554Lys in patients with PA gave a MAF of 3% vs 0% in neonatal controls using allele specific PCR. Description: M.SC.PATHOLOGY Sun, 01 Jan 2017 00:00:00 GMT https://www.um.edu.mt/library/oar/handle/123456789/50087 2017-01-01T00:00:00Z https://www.um.edu.mt/library/oar/handle/123456789/40244 Title: Determining the genotype of the Rh blood group system in the Maltese population Abstract: The Rh blood group system is a complex protein based system that consists of more than 50 antigens with more than 200 different RHD alleles and 50 different RHCE alleles. The five most important antigens from this blood group system are the D, C/c and E/e since they are the cause of most alloimmunisations. The Rh blood group system has a vital role in population genetic studies and by DNA typing important medical issues in transfusion practice can be resolved. Blood group genotyping is the method of choice when serological techniques cannot be used. The aim of the study was, to successfully use methods such as polymerase chain reaction (PCR) to determine the frequency of the RH genotype in the Maltese population. Blood donor samples (n=400) and cord blood samples (n=397) were enrolled in this study. An allele-specific polymerase chain reaction (AS-PCR) method was used to determine the presence of RHD, RHCE*E and RHCE*e, while multiplex PCR was used to test for RHCE*C/c. PCR products were then separated on agarose gel and visualised with ethidium bromide. In these samples, the incidence of RHD was approximately 90% and the remaining samples were negative for the RHD allele. The most common allele was RHCE*e with a percentage of 98% in blood donors and 99% in cord bloods. The most common genotype in RhD positive samples was DCcee and in RhD negative it was dccee. The most common haplotype was DCe making DCe/DCe the most common probable genotype. Like in previous studies, this research also concludes that the distribution of the RII genotype varies in different geographical areas. Description: M.SC.BIOMED.SCI. Sun, 01 Jan 2017 00:00:00 GMT https://www.um.edu.mt/library/oar/handle/123456789/40244 2017-01-01T00:00:00Z https://www.um.edu.mt/library/oar/handle/123456789/34426 Title: Optimization of a flow cytometric method to confirm the differentiation of HL-60 cell lines by candidate chemicals as indicated by screening tests Abstract: Submitted to the Faculty of Medicine and Surgery of the University of Malta in partial fulfilment for the degree of Master in Biomedical Science Acute Myeloid Leukemias (AMLs) are a group of heterogenous haematopoietic disorders. They are characterized by the accumulation of myeloid blasts in the blood and bone marrow resulting in haematopoietic insufficiency with or without leucocytosis. This project is a follow up of collaborative research as a part of a COST Consortium called STEM CHEM. The main aim of this collaboration is to work with chemicals and natural products which induce differentiation of the HL-60 leukaemic cell line. The main aim of this study is to identify the lineage and stage of differentiation of treated HL-60 leukemic cells according to shifts in their respective immunophenotype as the cell differentiates using Flow Cytometry. A panel of antibodies was designed to identify the lineage and maturing pattern of treated HL-60 cells. The panel consists of eight different antibodies: CD45 (gating marker); CD64 and CD 14 (monoytic markers); CD34 (blast marker); CD15 and MPO (granulocytic markers); and CD13 and CD11b (maturing markers). The staining method and the panel of antibodies were optimized. A set of candidate chemicals were tested to identify any shifts in the immunophenotype of the HL-60 cells. The panel of antibodies shows clear shifts in the immunophenotype of cells and therefore the chemicals which induced differentiation (shift in the immunophenotype) were clearly identified. The chemicals tested in this study resulted as showing approximately a PV of 70% between the standard screening tests and the gold standard technique flow cytometry. This study concluded that using screening tests to select the best candidate chemicals for differentiation is a feasible option, however, flow cytometry is important to carry out in order to confirm any shifts in the immunophenotypic pattern of cells and thus determine differentiation. Description: M.SC.BIOMED.SCI. Sun, 01 Jan 2017 00:00:00 GMT https://www.um.edu.mt/library/oar/handle/123456789/34426 2017-01-01T00:00:00Z