ISCHAEMIC ACUTE RENAL FAILURE AND AGE: MODULATION BY ANTI-INFLAMMATORY EMBRYONIC STEM CELL-DERIVED MACROPHAGES

Renal ischaemia-reperfusion injury follows the restoration of blood flow to kidneys starved of blood following a prolonged lowering of blood pressure caused, for example, by severe infection or trauma. The kidney is infiltrated with white blood cells such as macrophages and many kidney cells die resulting in acute renal failure and the requirement for dialysis. Elderly individuals are more vulnerable to developing acute renal failure as a complication of other illnesses and have a worse outcome compared to younger individuals. This proposal will use state-of-the-art embryonic stem cell technology together with genetic and pharmacological manipulation to generate significant numbers of macrophages that express the anti-inflammatory enzyme hemeoxygenase-1. The nature of these embryonic stem cell-derived macrophages will be scrutinised in cell culture and they will then be administered to mice with ischaemia-reperfusion injury as ?cell therapy? with the prediction that they will significantly protect kidney function. The effect of macrophage cell therapy will be specifically examined in old mice that develop worse renal failure than young mice following a period of diminished blood flow to the kidney. This work will lay the foundation for further studies utilising the potential anti-inflammatory and reparative properties of embryonic stem cell-derived ?designer? macrophages.

Renal ischaemia-reperfusion injury (IRI) plays a key role in the development of acute renal failure. Ischaemic acute renal failure remains a serious clinical disorder with significant morbidity and mortality and has an increased incidence and worse outcome in the elderly. Our recent work using a murine model of renal IRI has generated 2 key observations. First, old mice (52 weeks of age) exhibit worse renal failure and increased structural injury following IRI compared to 8 week old adult mice with significantly reduced upregulation of the anti-inflammatory renoprotective enzyme hemeoxygenase-1 (HO-1) evident in old mice. Second, the intravenous administration of macrophages genetically modified to overexpress HO-1 significantly attenuates the development of acute renal failure following renal IRI. We now plan to extend this work by using macrophages generated in vitro from embryonic stem (ES) cells (?designer macrophages?) that have been genetically or pharmacologically manipulated to overexpress HO-1. We will generate ES cell-derived macrophages in vitro and characterise their phenotype using various assays including the cytokine response to inflammatory stimuli, phagocytosis etc. The functional and histological effect of administering modified and control ES cell-derived macrophages to both young and old mice with renal IRI will be determined. Macrophage ?cell therapy? has great potential to alleviate human disease affecting the kidney and ES-derived macrophages are more attractive than monocytes-derived macrophages for several reasons. ES-derived macrophages represent a potentially ?off the shelf? resource that can be rapidly administered to patients without the requirement for human primary monocytes/macrophage cell culture (7 days). It is relatively straightforward to genetically manipulate ES cells such that single or multiple genes may be overexpressed or silenced in the subsequently generated macrophages. The work outlined in this proposal will lay the experimental foundation for using specifically designed ES-derived anti-inflammatory macrophages to ameliorate acute renal injury and inflammation and will have direct relevance to IRI affecting other organs such as liver, heart and brain.