GSE110021
RNA-Seq analysis of genes and pathways involved in the TGF-ß-driven transformation of fibroblasts to myofibroblasts
Background: Asthma is the most common chronic lung disease in children and young adults worldwide. Airway remodelling (including increased fibroblasts and myofibroblasts in airway walls due to chronic inflammation) differentiates asthmatic from non-asthmatic airways. The increase in airway fibroblasts and myofibroblasts occurs via epithelial to mesenchymal transition (EMT) where epithelial cells lose their tight junctions and are transdifferentiated to mesenchymal cells, with further increases in myofibroblasts occurring via fibroblast-myofibroblast transition (FMT). Transforming growth factor (TGF)-ß is the central EMT- and FMT-inducing cytokine. In this study, we have used next generation sequencing to delineate the changes in the fibroblast transcriptome induced by TGF-ß treatment in both the short term and after differentiation into myofibroblasts, to gain an understanding of the contribution of TGF-b induced transdifferentiation to the asthmatic phenotype. The data obtained from RNAseq analysis was confirmed by quantitative PCR (qPCR). Results: As expected, we found that genes coding for intermediates in the TGF-ß signalling pathways (SMADs) were differentially expressed after treatment, and genes involved in cytoskeletal pathways (FN1, LAMA, ITGB1) were differentially expressed in myofibroblasts compared to fibroblasts. Importantly, genes that were previously shown to be changed in asthmatic lungs (ADAMTS1, DSP, TIMPs, MMPs) were differentially expressed in myofibroblasts, strongly suggesting that TGF-ß mediated differentiation of fibroblasts to myofibroblasts may underlie important changes in the asthmatic airway. We also identified new signalling pathways (AKT, PTEN) that are changed in myofibroblasts compared to fibroblasts. Conclusion: We have found a significant number of genes that are altered after differentiation of fibroblasts into myofibroblasts by TGF-ß treatment, many of which were expected or predicted. However, we also identified novel genes and pathways that were affected after treatment of fibroblasts with TGF-ß, which suggests additional pathways that that are activated during the transition between fibroblasts and myofibroblasts, and may contribute to the asthma phenotype. Overall design: cDNA profiles from WI-38 fibroblasts treated with or without TGFb and collected on day 1 and day 20 after treatment. Triplicate samples were analysed for each timepoint and treatment (12 samples in total).