Abstract/Summary

Necrotising enterocolitis (NEC) is a devastating disease that can affect pre-term infants and if survived can lead to long term intestinal damage. Currently a variety of antibiotics and in extreme cases surgery is required to manage the disease. However, no treatment exists that attenuates the widespread cell death and suppresses the innate pro-inflammatory response whilst promoting regeneration in the intestine thus reducing long term damage and the need for surgery. The discovery of stem cells, in particular multipotent stem cells (MSCs) has shown them to have potent regenerative therapeutic properties. In particular, C-kit+ foetal derived MSCs from amniotic fluid (amniotic fluid stem cells - AFSCs) when introduced into a rat model of NEC increased the survival of animals affected whilst also promoting the repair of damaged epithelial layers. Nevertheless, the ability of stem cells to promote regeneration was independent of transplanted cell engraftment. This led to a paradigm shift in which it was proposed that regeneration was via the secretion of factors acting in a paracrine manner with target cells. This was proven by the introduction of stem cell conditioned media (CM), which mimicked the beneficial effect seen when whole MSCs were introduced. However, much of the MSC CM is produced from non-stressed MSCs. Stressing MSCs has been shown to increase the secretion of paracrine factors and the extracellular vesicles that contain them. Creation of an AFSC CM (AF-CM) under conditions of stress using a protocol created by industrial partners was therefore hypothesised to produce a more potent AF-CM beneficial for regeneration and in particular for the treatment of NEC. Here we characterise the AF-CM under conditions of stress but also in a clinically compliant manner. The AF-CM was characterised for protein and RNA content, which elucidated to a multitude of proteins present but interestingly showed that the majority of RNA present was miRNA. In vitro investigation into AF-CM treatment also showed significant improvements in cellular proliferation and migration capacity whilst protecting from senescence. Furthermore, I showed that AF-CM treatment accelerated the regeneration of skeletal muscle fibres following an established mouse model of acute cardiotoxin damage in vivo, of which could be partly attributed to increased blood vasculature.