INFLUENCE OF NADPH OXIDASES ON IPS DIFFERENTIATION AND FUNCTION OF IPS-DERIVED ENDOTHELIAL CELLS 

Induced pluripotent stem (iPS) cell-derived endothelial cells (ECs), which display characteristic morphology and marked vasoreparative capacity both in vitro and in vivo, hold clear potential for therapeutic angiogenesis in ischaemic disease. Oxidative stress and reactive oxygen species (ROS), which are central features of ischaemic disease, are established regulators of both EC and stem cell biology, with recent evidence specifically highlighting NADPH oxidases. Notably, we have identified a key role for the NOX4 isoform in regulating iPS differentiation to iPS-ECs and its importance in maintening the iPS-EC phenotype. Specifically, we investigated the influence of NOX4 NADPH oxidase on iPS differentiation and function of iPS-ECs. Our data indicate that NOX4 (mRNA) is progressively induced during iPS to iPS-EC differentiation. Following NOX4 overexpression, endothelial progenitor markers were significantly increased, suggesting NOX4 as a potential target for promoting EC differentiation efficiency. Interestingly, while endogenous ROS actions mediated by NOX4 activity enhanced differentiation, exogenous ROS induced by phorbol 12-myristate 13-acetate (PMA), a PKC activator, were shown to inhibit this process. The precise mechanisms through which ROS regulate cellular differentiation remain unknown, although it appears likely that these may incorporate important yet-to-be-explored epigenetic changes. Therefore, together with detailed investigation of the specific influence of NOX4-derived ROS on iPS differentiation through extensive genomic/proteomic studies, wide-screen epigenetic profiling is critical to identify key ROS-induced remodelling events relating to the DNA (methylation) and chromatin (histone methylation, acetylation etc) which are likely to regulate this dynamic process.