Saudi Cultural Missions Theses & Dissertations
Permanent URI for this communityhttps://drepo.sdl.edu.sa/handle/20.500.14154/10
Browse
3 results
Search Results
Item Restricted Bioengineering of transcriptional elements driving MMP13 gene in skeletal development(University of Liverpool, 2024-08) AlSalhi, Sara Ibrahim; Bou-Gharios, GeorgeMmp13 is a primary catabolic factor involved in cartilage degradation through its ability to cleave collagen type II and other cellular components. In addition to being necessary for the formation of various cells, organs, and tissues, Mmp13 expression is regulated transcriptionally by two main elements: the proximal promoter and distal enhancers. This study aims to identify the transcriptional elements that regulate Mmp13 gene expression. Identification of novel Mmp13 enhancers was conducted in silico using the Encyclopaedia of DNA Elements (ENCODE), based on histone modifications (Limb H3K4me1 and Limb H3K27Ac), fibroblast coverage, chondrocyte, and embryonic limb regulatory elements from public ChIP-Seq data, and evolutionarily conserved sequences, in addition to transcription factor profiles of Runx2 and vitamin D. All prospective Mmp13 enhancer sequences were analysed using three different software tools: CIIIDER, TRANSFAC, and JASPAR, for the prediction of transcription factor binding sites. Each enhancer sequence was cloned upstream of the Hsp68 silenced promoter and lacZ gene to create a β-galactosidase reporter construct, which was then used to generate transgenic mice. Embryos were collected at E15.5 and tested for lacZ gene expression and tissue expression analysis. Constructs were also transfected in vitro into pre-osteoblasts (MC3T3- E1), NIH3T3 mouse embryo fibroblasts, human chondrocytes (SW1353), and primary chondrocytes extracted from OA patients. Among the seven tested Mmp13 enhancer regions, strong skeletal element expression was detected in the region from -21.9 to -21.1 kb, which overlaps with Runx2 and Sox9 binding sites. Other enhancers revealed some skeletal element activity but were not as prominent. Expression in various tissues and organs, including skin, was observed in multiple regions. In contrast, sequences aligning with the highest peaks of Runx2 at -29 kb and -32.5 kb did not show significant expression. In vitro, Mmp13-transfected enhancer sequences demonstrated enzyme activity, with the highest responses observed in chondrocyte and human cells at -10 kb and -29 kb regions, along with -21.4 kb that indicate a potential regulatory influence. Comparisons of potential enhancers in mouse embryos highlighted the sequences in the intronic 5', -10 kb, -19.2 kb, and -21.4 kb regions as significant Mmp13 enhancers regulating gene expression but not -29 and -32.5 kb regions. Identifying these specific enhancers could lead to targeted therapeutic strategies to modulate MMP13 activity, potentially slowing or preventing cartilage degradation in degenerative diseases.18 0Item Restricted Intracellular and extracellular immunisation for HIV(The University of Manchester, 2024-11) Alatiq, Abdulrahman; Klapper, Paul; Valley, Pamela; Faqih, LaylaThe prevalence of HIV infection remains a serious challenge for public health. HIV infection results in considerable mortality and morbidity, as well as imposing a substantial financial burden on healthcare systems globally. Despite the availability of effective antiretroviral treatments to control infection, an effective protective vaccine remains elusive. This project aimed to develop a practical, affordable, and effective combined vaccine and treatment by combining intracellular antibodies and extracellular immunisation to prevent or reduce HIV viraemia. Specifically, an approach in which DNA-encoding N49P7 broadly neutralising antibodies against the HIV-1 envelope glycoprotein 120 is delivered to cells using recombinant baculovirus or lipid nanoparticle (LNP) transfection. Extracellular antibodies were anticipated to prevent gp120 from attaching to cells, and intracellular antibodies were intended to inhibit the genesis of virions. The expression of N49P7, either as a human IgG antibody or as human Fab, was attempted. The expression of the N49P7 IgG antibody was not achieved in HEK 293 cells despite employing three delivery models for transfections: recombinant baculovirus, MC3-based LNP, or lipofectamine 2000 LNP. While transfection was successfully achieved (monitored via an eGFP gene included within the plasmid design), functional antibody was not achieved. The most likely explanation for the failure was thought to be the use of dual promoters within the expression cassette. Redesign of the plasmid to create a bicistronic vector including the N49P7 Fab region and signal peptide sequences of murine IgG and IL-2 allowed successful expression of N49P7 Fab through recombinant baculovirus or lipofectamine 2000 reagent transfection of HEK 293 cells. However, the expressed N49P7 Fab region was predominantly accumulated intracellularly. A further redesign was implemented to incorporate homogenous signal peptides H7 and L1 into the N49P7 Fab gene, which significantly enhanced the secretion of the antibody fragment. This design maintained functional intracellular and extracellular antibody activity. Lower cellular cytotoxicity was seen with recombinant baculovirus transfection compared to lipofectamine 2000 LNP mediated transfection although both were equally efficient. The selection of an optimum method for transfection will form part of future investigations progressing to animal model testing. This proof of principle study showed that recombinant baculovirus or lipofectamine 2000-mediated transfection systems allowed efficient N49P7 Fab expression both intracellularly and extracellularly in mammalian cells, suggesting that this approach indicates potential for providing a vaccine against HIV infection in addition to a therapeutic intervention for those who already have HIV infection.12 0Item Restricted Resonance in Dissonance: Noise and the Aesthetics of Auditory Abstraction(Pratt Institute, 2024-06-29) Basowad, Mariam Ali S; Liebergesell, Alex; Echeverria, Maria Gracia“Dissonance,” “discordance,” and “cacophony” are among the countless negative connotations associated with noise in the fields of communication, musicology, and sound studies. In these contexts, noise is often perceived as an “interference” that needs to be mitigated and controlled. Similar biases exist in communications design, influenced partly by a historical preference for clarity, order, and coherence, coupled with the belief that noise diverges from conventional design principles. However, in this thesis, noise is presented as an event, a flux, a multisensory entity, and, most importantly, a form of emancipation from established artistic norms. This thesis aims to explore the vital role of abstract sonic noise in communication, considering how to craft more resonant and immersive experiences and unlock new possibilities for creativity and expression. To achieve this objective, the thesis surveys sound, philosophy, and communication studies, drawing upon the insights of esteemed philosophers and composers, including Jacques Attali, Salomé Voegelin, and Luigi Russolo. Additionally, it investigates historical and contemporary practices of incorporating noise in creative fields, such as music and audiovisual art. Through a synthesis of theoretical and practical experimentation, this thesis demonstrates the effectiveness of noise as a communicative and resonant medium.21 0