Sumeray, Mark Stephen; (1998) An in-vitro model of ischaemia-reperfusion injury in murine myocardium : The role of nitric oxide. Doctoral thesis (Ph.D.), University College London (United Kingdom).

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Abstract

Recent advances in understanding the mouse genome and genetic manipulative techniques have created a unique opportunity to explore the influence of single gene alterations on complex physiological systems. It is essential that physiologists respond to this challenge by careful design of appropriate, miniaturised systems to allow phenotypic changes to be explored. This thesis describes the stages in the establishment of a new model of ischaemia-reperfusion injury in murine myocardium. The first experimental chapter describes the detailed characterisation and validation of the model. Technical aspects of isolated mouse heart perfusion and construction of the Langendorff apparatus are discussed. The end-points of injury (infarct size, contractile recovery, and lactate dehydrogenase release) were co-correlated, and good dose-response relationships demonstrated against duration of global ischaemia. The second series of experiments showed that the model can be used to assess the efficacy of a well-known cardioprotective intervention (ischaemic preconditioning). Short episodes of global ischaemia were used to precondition against a subsequent 30 minute global ischaemic insult. Four periods of five minutes ischaemia were necessary to induce protection (a 42% reduction in infarct size). This was the first demonstration of this phenomenon in the isolated mouse heart using infarct size as the primary end-point. Having established the model and demonstrated its capacity to assess the efficacy of a cardioprotective intervention, the final series of experiments investigated the role of nitric oxide in myocardial ischaemia-reperfusion injury using genetically-altered mice. 'Knockouts' for both constitutive isoforms of nitric oxide synthase (endothelial and neuronal) were used, together with wild-types, to assess the response to a protocol of global ischaemia-reperfusion injury. The results demonstrated, for the first time in murine myocardium, the protective effect of nitric oxide derived from endothelial nitric oxide synthase, and suggested a detrimental role for the neuronal isoform.

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