Understanding the contribution of tumour cells and stroma to changes in microRNA expression in malignant pleural mesothelioma

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Malignant pleural mesothelioma (MPM), a relatively rare cancer arising from the mesothelial lining of the pleura, is a highly aggressive cancer with a very poor prognosis, and an urgent need for more effective treatments. MicroRNAs (miRNAs) are short, non-coding RNA sequences that negatively regulate gene expression and their abundance is frequently dysregulated in cancers. Dysregulation of miRNA expression plays an important role in the biology of MPM and candidate miRNAs have been investigated as diagnostic and prognostic biomarkers, or as potential treatment targets for MPM. Dysregulation of miRNA expression is determined through comparisons of miRNA expression profiles of tumour and corresponding normal tissues. Tumours, however, consist of non-tumour stromal cell types that also play important roles in tumour development. The contribution of stromal cell miRNA expression can influence the selection of dysregulated miRNAs, and their gene targets for analysis. Since in tumour miRNA profiling studies, the source of miRNAs may be from tumour cells, stromal cells or a combination of both, this thesis aims to better understand the contribution of tumour cells and stroma to changes in miRNA expression in MPM. This project investigated the relative contribution of miRNAs from tumour and stromal cells in MSTO and H226 cell line-derived xenograft tumours compared to their corresponding cultured tumour cells. RT-qPCR quantification detected high expression of miR-143-3p, miR-214-3p and miR-223-3p in xenograft tumours and confirmed their stromal origin. Additionally, digital droplet PCR (ddPCR) quantification of the species-specific pri-miRNA transcripts of these three miRNAs further confirmed predominant expression by mouse stromal cells. Although there is strong evidence supporting the expression miR-143-3p, miR-214-3p and miR-223-3p predominantly in stromal cells, these miRNAs have all been reported to be downregulated in MPM. This study therefore also explored the functional consequences of overexpressing these miRNAs in MPM cell lines to test their reported tumour suppressor effects but found an absence of such effects. In conclusion, identifying miRNA dysregulation in MPM by comparing tumour and normal tissue expression profiles is ineffective due to the presence of stromal cells that can contribute to the miRNA expression pool at varying levels, hence influencing downstream gene targets. It is therefore of absolute importance to understand the cell source of miRNAs prior to carrying out functional studies to gain a better understanding of the contribution of miRNAs to the roles of tumour and stromal cells in MPM.
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