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Role of extracellular matrix in vascular smooth muscle cell behaviour and the identification of novel markers of cell phenotype.
Increased synthesis and deposition of extracellular matrix proteins in the blood vessel wall are implicated in vascular disorders such as atherosclerosis, restenosis and hypertension. (Liau and Chan 1989). The increase in extracellular matrix proteins can be mostly attributed to smooth muscle cells (SMCâs) within vascular lesions (Mecham et al 1987). It is presumed that modulation of SMCâs from their normally quiescent, contractile phenotype to a proliferative synthetic phenotype results in increased synthesis of extracellular matrix proteins. Studies have also demonstrated that signals elicited from the extracellular matrix (ECM) may play a role in the regulation of this SMC phenotypic modulation (Yamamoto et al 1993; Hedin et al 1988). The mechanism by which ECM can alter the phenotypic state of SMCâs is not well understood but clearly involves the induction of intracellular signals as a consequence of ECM ligand - cell surface integrin binding. These signals must subsequently exert downstream molecular events altering gene expression and ultimately cell phenotype. The research project presented in this thesis examined the influence various extracellular matrix substrates have on vascular SMC behaviour in vitro. Initial observations demonstrated that SMCâs cultured on different matrix substrates exhibit distinct morphological growth patterns. Functional differences in SMC proliferation and migration rates were also observed in response to seeding on different ECM surfaces.Analysis of the expression levels of known SMC phenotypic protein markers between SMCâs cultured on different matrix substrates did not reveal any significant differences in protein expression. Slight upregulation of Myosin Light Chain Kinase (MLCK)-210 kd isoform was observed in SMCâs cultured on cellular fibronectin, Collagen III and Vitronectin substrates. The Meta-vinculin protein was upregulated in SMCâs cultured on fibronectin coated substrates.In order to identify altered gene expression patterns induced by ECM adhesion, SMC populations cultured on fibronectin coated plastic and SMCâs grown on uncoated plastic
were selected for analysis by the differential display technique. Differential display is a recently developed PCR based technique that allows the identification of differentially expressed genes between related cell populations (Liang and Pardee 1992). Much effort was spent optimising the differential display procedure to overcome such limitations as primer redundancy and high false positive selection rates. However, as a result of the persistence of false positives only one gene, meta-vinculin was confirmed as being differentially expressed between SMC populations cultured on fibronectin coated and uncoated tissue culture plastic surfaces.It was concluded that the failure to identify a significant number of fibronectin modulated genes was probably a result of limitations in the differential display procedure as carried out in this study, and may also possibly be due to the existence of only very minor
differences in phenotype between SMCâs grown on plastic with or without fibronectin precoating
Evaluation of machine learning classifiers to predict compound mechanism of action when transferred across distinct cell-lines.
Accelerating glioblastoma drug discovery: Convergence of patient-derived models, genome editing and phenotypic screening
High-Content Imaging 2023: A Joint Special Collection with the Society for Biomolecular Imaging and Informatics and SLAS
High content phenotypic and pathway profiling to advance drug discovery in diseases of unmet need
Patient derived glioma stem cell spheroid reporter assays for live cell high content analysis
Evolution and Impact of High Content Imaging
Abstract/outline: The field of high content imaging has steadily evolved and expanded substantially across many industry and academic research institutions since it was first described in the early 1990â˛s. High content imaging refers to the automated acquisition and analysis of microscopic images from a variety of biological sample types. Integration of high content imaging microscopes with multiwell plate handling robotics enables high content imaging to be performed at scale and support medium- to high-throughput screening of pharmacological, genetic and diverse environmental perturbations upon complex biological systems ranging from 2D cell cultures to 3D tissue organoids to small model organisms. In this perspective article the authors provide a collective view on the following key discussion points relevant to the evolution of high content imaging:⢠Evolution and impact of high content imaging: An academic perspective⢠Evolution and impact of high content imaging: An industry perspective⢠Evolution of high content image analysis⢠Evolution of high content data analysis pipelines towards multiparametric and phenotypic profiling applications⢠The role of data integration and multiomics⢠The role and evolution of image data repositories and sharing standards⢠Future perspective of high content imaging hardware and softwar
High Content Phenotypic Profiling in Oesophageal Adenocarcinoma Identifies Selectively Active Pharmacological Classes of Drugs for Repurposing and Chemical Starting Points for Novel Drug Discovery
High Content Phenotypic Screening Identifies Serotonin Receptor Modulators with Selective Activity upon Breast Cancer Cell Cycle and Cytokine Signaling Pathways
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