Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.713096
Title: The role of NADPH oxidase-2 in the development of cardiac fibrosis
Author: Richards, Daniel Antonio
Awarding Body: King's College London
Current Institution: King's College London (University of London)
Date of Award: 2017
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Abstract:
Background: Cardiac fibrosis, a component of pathological remodelling, often arises secondary to hypertension or myocardial infarction. It leads to systolic and diastolic cardiac dysfunction, since the increased collagen deposition increases myocardial stiffness. Reactive oxygen species (ROS) are implicated in this pathology and NADPH oxidase-2 (NOX2), a major producer of ROS, is elevated in heart failure settings. Previous work showed that global NOX2 knockout (KO) mice had reduced generation of cardiac fibrosis, but the cell-type specific role of NOX2 is yet to be investigated in this context. Aim: This research aimed to determine the role of fibroblast and endothelial cell NOX2 in the development of cardiac fibrosis in mice. In addition, the extent of endothelial-mesenchymal transition (EndoMT) in the development of cardiac fibrosis was to be assessed using a lineage tracing approach. Methods and Results: Tamoxifen-inducible fibroblast-specific NOX2 KO mice were generated by crossing novel NOX2 floxed mice with Collagen 1α2 (Col1α2)-Cre mice. Cardiac NOX2 protein expression was reduced by 60% in these mice. Cardiac fibrosis was induced using chronic angiotensin II (Ang II) infusion (1.1mg/kg/day) and blood pressure was monitored by ambulatory telemetry. Fibroblast NOX2 KO mice had a significantly delayed hypertensive response and less cardiac fibrosis compared to wildtype (WT), as determined by histological analysis of cardiac sections. In contrast, fibroblast NOX2 KO mice were not protected from cardiac fibrosis when subjected to chronic pressure overload by transverse aortic constriction (TAC), nor were they protected from systolic and diastolic dysfunction as determined by serial echocardiography. Similarly, endothelial cell-specific NOX2 KO mice were generated by crossing floxed NOX2 mice with VE-Cadherin (Cdh5)-Cre mice, with a 60% reduction in aortic NOX2 protein levels. These mice were also not protected from developing cardiac fibrosis or cardiac dysfunction when subjected to TAC. The Col1α2 and Cdh5 Cre-recombinase efficiencies were assessed by crossing with a tdTomato “STOP” floxed reporter mouse, showing high tdTomato expression in specific cell types and estimated high efficiency of ≥70% using flow cytometry. The extent of EndoMT was assessed in Cdh5 tdTomato (EndoTom) mice subjected to TAC using a lineage tracing approach. There was no convincing evidence of EndoMT in cardiac or lung tissue, since the percentage of tdTomato+ (Cdh5 lineage traced) cells expressing the fibroblast marker CD140α was not augmented by TAC (determined by flow cytometry). Conclusion: Although fibroblast NOX2 contributes to the development of Ang II-induced cardiac fibrosis it is dispensable for TAC-induced fibrosis. Endothelial cell NOX2 is also dispensable for TAC-induced fibrosis. These results suggest that the cell-specific role of NOX2 in cardiac fibrosis is stimulus dependent. The activation of NOX2 in other cell types may be required for the development of cardiac fibrosis in response to chronic pressure overload. Furthermore, EndoMT appears to be at most a minor contributor to the development of cardiac fibrosis after TAC. Taken together, this work provides new insights into the cell-specific roles of NOX2 in the development of cardiac fibrosis.
Supervisor: Watt, Fiona Mary ; Shah, Ajay Manmohan Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.713096  DOI: Not available
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