Development of circulatory microRNAs as markers of organ injury and mediators of inter-organ signalling
Morrison, Emma Elisabeth
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Plasma contains small, non-protein coding RNA species, microRNAs (miRNAs). Circulating miRNAs originate from tissues throughout the body and circulate in the blood bound to proteins or encapsulated in extracellular vesicles (EVs). The pattern of circulating miRNAs changes in different pathological states, leading to the hypothesis that they could act as biomarkers or mediators of inter-organ signalling. Acute kidney injury (AKI) is associated with high morbidity worldwide. Recent work has highlighted a potential role for EV signalling in the delivery of functional exogenous miRNA into kidney cells, which may contribute to the pathogenesis of AKI. The studies described in this thesis investigate the effects of circulating miRNAs on renal proximal tubular (PT) cells. Utilising next generation sequencing technology, circulating miRNA profiles were demonstrated to change significantly following myocardial injury. These findings were translated from humans into a mouse model of myocardial injury. Investigation of EV cell signalling, using flow cytometry and nanoparticle tracking analysis, demonstrated that PT cell EV uptake was not affected by known physiological agonists. By contrast, EV release from PT cells was regulated by purinergic P2Y1 and dopamine D1 receptors. Toxic cisplatin injury of PT cells resulted in increased EV release and reduced EV uptake in a dose-dependent manner. Cisplatin toxicity in PT cells was unaffected by EVs from mice with myocardial injury, but toxicity was reduced by EVs from mice with drug-induced liver injury (DILI). Circulating EVs from mice with DILI transferred the liver specific miRNA, miR-122, into PT cells in both in vivo and in vitro models. The consequence of miR-122 transfer was modulation of downstream target genes including Foxo3 which has been implicated in cell injury by apoptosis. These findings therefore show that circulatory miRNA profiles change in different models of organ injury and suggest miRNAs can be transferred to PT cells in vivo and in vitro. The improved viability of injured PT cells following co-incubation with DILI EVs, and subsequent transcriptomic work, suggests this may be as a consequence of miRNA transfer. In conclusion, circulatory miRNAs may act as mediators of inter-organ signalling and could play a crucial role in the propagation of systemic illness.