Cellular phenotype characterisation in different variants of visceral myopathy

Sardar, Sabah (2024) Cellular phenotype characterisation in different variants of visceral myopathy. PhD thesis, University of Glasgow.

Abstract

Visceral myopathy is a rare and severe gastrointestinal motility disorder, affecting 1 in 100,000 individuals globally. It is a genetically heterogeneous disease where the genetics behind the disorder are not fully understood. Visceral myopathy results in a pseudo-obstruction in the intestine without the presence of a mechanical obstruction. This has been attributed to an impairment in the contractile phenotype of smooth muscle cells. Several genes, and variants within those genes, have been associated with visceral myopathy, all of which cause similar symptoms to develop in individuals with the disease. Currently, variants in the enteric smooth muscle actin gamma 2 (ACTG2 ) gene have been identified as causing approximately 40-50% of cases. However, the rarity of visceral myopathy coupled with misdiagnosis or late diagnosis, has resulted in limited advances in available treatment options.

Whilst the disease impacts on a mechanical phenotype, its mechanobiology has not been fully investigated yet. Therefore, studying the disorder from a mechanobiological and biochemical perspective is beneficial for gaining insights into the disease pathology in order to aid in developing therapies and improving disease diagnosis. With this in mind, patient dermal fibroblasts were used as a model for investigating the morpho-mechanical and biochemical characteristics of visceral myopathy cells. The variants investigated were ACTG2 variants R257C and R38H, as well as novel PIEZO1 biallelic mutations affecting S1814F and P2230L. Initially, Yes-associated protein (YAP) translocation and traction forces exerted by cells were analysed. This was followed by the study of cell morphology, the mechanical clutch model, and migration. The metabolic profiles were then assessed and comparisons were drawn between visceral myopathy caused by ACTG2 and PIEZO1 mutations. PIEZO1 associated VSCM was found to differ completely from ACTG2 -VSCM, at a morpho-mechanical and metabolomic level.

Furthermore, in this work, YAP nuclear localisation, traction forces, and cell migration were identified as potential disease markers which can be exploited for the development of diagnostic and drug screening assays. However, it was traction forces which emerged as the most promising label-free physical biomarker for visceral myopathy. Particularly, when traction forces are measured using the micropillar array platform as it provides a robust method which has the potential to be used for therapeutic purposes.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Colleges/Schools: College of Science and Engineering > School of Engineering
• Supervisor's Name: Vassalli, Professor Massimo, Dalby, Professor Matthew and Salmeron-Sanchez, Professor Manuel
• Date of Award: 2024
• Depositing User: Theses Team
• Unique ID: glathesis:2024-84706
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 18 Nov 2024 15:59
• Last Modified: 18 Nov 2024 15:59
• Thesis DOI: 10.5525/gla.thesis.84706
• URI: https://theses.gla.ac.uk/id/eprint/84706