Role of peritoneal mesothelial cells and the inflammatory response in peritoneal fibrosis

Post-operative adhesion is a common complication after abdominal surgery, with high impact on patient wellbeing and healthcare costs. The repair of peritoneum is a complex process involving orderly phases which share some common features to normal wound healing. These include coagulation, infiltration of inflammatory cells, cell proliferation, extracellular matrix (ECM) deposition and remodelling, often with overlap between phases. The unique feature of peritoneal repair is that both small and large peritoneal wounds heal in a similar time. The peritoneum is a monolayer of elongated, flattened, squamous-like peritoneal mesothelial cells (PMC). Local mesothelial cell proliferation, centripetal cell migration from the wound edge, as well as incorporation of free-floating mesothelial cells may all contribute to repair of injured peritoneum. To date, the only well-characterised pathologic mechanism underlying post-operative adhesion formation at the molecular level is the formation of the fibrin layer and regulation of peritoneal fibrinolytic capacity. However, the contributions of collagen deposition and ECM remodelling to the peritoneal repair mechanism are not well understood. This thesis focuses on the role of PMC in the regulation of ECM deposition and remodelling in response to inflammatory stimuli in both in vivo and in vitro models, aiming to identify other key pro-fibrotic factors involved in the development of post-operative adhesion. We first identified that lysyl oxidase (LOX) played a key role in the progression of peritoneal fibrosis by regulating collagen cross-linking and deposition in vivo. The inhibition of LOX enzyme activity prevented the formation of fibrotic tissue by reducing collagen deposition. Meanwhile, dexamethasone (DEX) treatment also minimized the fibrotic response. Furthermore, in vitro studies showed that the induction of collagen deposition factors in PMC, including LOX and pro-collagen I, required both IL-1 and TGF-β signalling pathways. Thus, the combination of IL-1 + TGF-β was adopted in an in vitro model to mimic the inflammatory environment during peritoneal repair. Treatment of PMC with IL-1+TGF-β caused an epithelial-to-mesenchymal transition (EMT). These transformed PMC had enhanced cell motility and were more adherent to fibronectin. Finally, a real-time quantitative PCR-based microarray was used for genomic analysis of ECM-adhesion-related PMC genes in response to IL-1 and TGF-β treatment. The results showed that IL-1 was more involved in regulating ECM degradation by inducing expression of matrix metalloproteinase (MMP) genes, whereas TGF-β mainly affected genes involved in ECM deposition, including collagens and other ECM components. However, both cytokines were shown to regulate some key genes involved in the development of adhesion, including COL16A1, COL7A1, FN1, ITGA5, and TGFB1. Moreover, IL-1 was shown to reduce ITGA4 and ITGB6 expression affecting adherence of PMC to basement membrane, while TGF-β increased MMP14 and MMP16 expression, which could facilitate invasion of EMT-transformed PMC to the site of tissue repair. In summary, this thesis indicates that LOX plays an important role in peritoneal fibrosis. Secondly, a combination of IL-1 and TGF-β1 treatment demonstrates how these factors can act in concert to orchestrate tissue remodelling during peritoneal repair. Finally, genomic analysis of ECM-adhesion genes increases our understanding of aspects of the pathology of post-operative adhesion and identifies novel potential therapeutic targets to prevent adhesion formation.EThOS - Electronic Theses Online ServiceGBUnited Kingdo