 Wouldn't it be great to have a hydrogel, which you could put onto a chronic non-healing wound, which would promote angiogenesis using the body's own growth factors and heparin? Well that's what we've set out to do in this study and I'm going to describe how this article gives us the first stages towards this. First I'm going to explain our thinking and then take you through the article. To get new blood vessels to form in an area you need a combination of things and pro-angiogenic growth factors such as vascular endothelial growth factor are a key part but we also need a better way of delivering these growth factors so that they stay in place until needed, don't break down rapidly and are released in sufficient concentrations to promote neovascularization. In nature there's a slow release of pro-angiogenic peptides from the wound site. We've sought to replicate some of these features using a biomimetic approach which combines polymer chemistry and vascular cell biology. We've developed a hydrogel which has amino acids on it which are naturally charged. These then bind heparin on an electrostatic basis. Heparin is one of the body's major glycozamine glycans capable of binding a range of growth factors and vascular endothelial growth factor. So we used a hydrogel which is itself non-fouling based on PNVP and acrylic acid. This we then aminated comparing lysine and arginine tripeptides. We then used the negative charge of these peptides to bind heparin. We show that the heparin can then bind vascular endothelial growth factor and that the heparin VEGF loaded hydrogel is capable of releasing vascular endothelial growth factor in sufficient quantities to stimulate the proliferation of small vessel endothelial cells. Figure one, an acrylic acid-derivitized poly-vinyl pyrrolidone hydrogel was synthesized and then functionalized with peptides. The hydrogels were reacted with either trilycine or tri-arginine with NHS and DCC for 24 hours at room temperature. The hydrogels were then washed extensively for a week to remove any non-covalently bound peptides. The presence of peptide was confirmed by XPS and TOF-SIMS and a TNBS colorometric assay was also used to give an indication of the bulk concentration of peptides attached to the hydrogels. We also demonstrated that the hydrogels and modified hydrogels were basically non-fouling for protein adhesion with the exception of lysosyme, which was bound to the lysine-modified hydrogels. Demonstration of heparin binding to the hydrogels was undertaken by XPS looking for the sulfur signal present in the heparin. We demonstrated semi-quantitative binding of heparin in these hydrogels using fluorescently labeled heparin and from this it was clear that the lysine-modified hydrogel bound approximately two and a half times as much as the non-modified hydrogel and the arginine-modified hydrogel bound approximately four times as much heparin. Then we looked at the ability of these heparin-modified hydrogels to bind and release vascular endothelial growth factor. The hydrogels were loaded with heparin and then with VEGF and the strength of VEGF binding was examined by extensively washing the hydrogels. The arginine-modified hydrogel retained the most VEGF under washing conditions and we demonstrated that accordingly it released the most VEGF over three days. We next looked at cell biocompatibility for the hydrogels looking at both fibroblasts and endothelial cells. Cells were stained with a vital fluorescent cell tracker dye so that we could image them more readily. We found that fibroblasts on tissue-cultural plastic grew right up to the edge of the piece of hydrogel placed within the well but there was no attachment to fibroblasts to the hydrogel until it was modified with arginine when a few cells did grow on the hydrogel. We demonstrated that the hydrogel had no cytotoxic effects on the fibroblasts when grown next to it but that a few cells were able to grow on the hydrogel modified with arginine. We then looked at the performance of the hydrogels with microvascular endothelial cells as with the fibroblast cells grew up to the edges of the gel but didn't grow on them in any appreciable numbers. Endothelial cells were then incubated for three days with hydrogels, preloaded with heparin and VEGF, and appropriate unloaded hydrogels were included and their effect on cell attachment and total cell viability was examined both in the presence of 5% fetal carcium and 2% fetal carcium. In 5% fetal carcium there was a slight stimulation of around 20% of proliferation with the arginine modified hydrogel but this wasn't significant. However, when we reduced the serum to 2% then the peptide functionalised hydrogels could clearly be seen to stimulate cell viability. We also saw a small number of endothelial cells growing on these modified hydrogels. In summary, we have used the ability of heparin to bind to basic peptide sequences exploring both lysine and arginine and demonstrating that hydrogels modified with lysine and arginine will bind heparin and in this respect arginine is more effective than lysine. In turn, the heparin bound to the hydrogels can bind VEGF and we demonstrated that these preloaded hydrogels will then significantly stimulate proliferation of adjacent endothelial cells which is one of the key steps in neovascularisation. So thank you for listening. I hope you find the paper interesting and useful for your own research.