Garner, IM; (2016) DNA methylation in lung fibroblasts and its role in pulmonary fibrosis. Doctoral thesis , UCL (University College London).

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Text 271015 Final version of thesis IAN GARNER 2015 corrections2.pdf - Accepted Version Download (17MB) | Preview

Abstract

Altered methylation and subsequent changes in gene expression have been implicated in several fibroses including lung however, the full extent and role of altered DNA methylation in fibrotic lung fibroblasts is unknown. Emerging evidence also suggests gender-specific methylation differences are common in disease and could elucidate why diseases characterised by pulmonary fibrosis including idiopathic pulmonary fibrosis (IPF) and systemic sclerosis (SSc) have a sex-biased prevalence. Using a genome-wide array-based approach, this thesis investigates differentially methylated and expressed genes in fibrotic compared to control lung fibroblasts, gender-specific methylation and expression differences and the effects of modulating DNA methylation using a DNA methyltransferase (DNMT) inhibitor, 5-Aza-2’deoxycytidine (5-Aza). Data show primary human IPF and SSc lung fibroblasts have multiple genes with altered DNA methylation and expression compared to control lung fibroblasts. Multiple biological processes were enriched in these genes, many of which are relevant to fibrosis including, transcriptional regulation, extracellular matrix (ECM) organisation, Wnt signalling and apoptosis. Using siRNA knockdown and collagen gel contraction assays, novel genes including Tenascin-XB (TNXB), which encodes the ECM glycoprotein Tenasicn-X (TNX), were identified as having potential functional significance in the pathogenesis of pulmonary fibrosis. Furthermore, multiple genes including TNXB had altered methylation and expression in IPF compared to SSc lung fibroblasts and may distinguish IPF from other diseases associated with pulmonary fibrosis. Multiple genes were identified with gender-specific differences in methylation and expression in lung fibroblasts. Interestingly, multiple genes with altered methylation in IPF males compared to control males were not the same genes with altered methylation in IPF females compared to control females, which may in part explain why IPF predominates in males. The final chapter of my thesis shows 5-Aza treatment alters the methylation and expression of multiple genes in primary human lung fibroblasts. Strong correlation between changes in methylation and changes in expression were identified suggesting DNA methylation can directly regulate the expression of multiple genes in lung fibroblasts.

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