Abstract 15272: Wnt11 Delivered in Mesenchymal Stem Cell-Derived Media Promotes Angiogenesis Through Non-canonical Wnt-PKC-JNK Pathway

Abstract

Background: Wnt11 is involved in many developmental and cellular processes. We have previously reported that Wnt11 overexpressing (MSC Wnt11 ) significantly improved cardiac function and increased angiogenesis. Here, we aimed to elucidate how Wnt11 delivered by MSC increase blood vessel formation. Methods and Results: An acute myocardial infarction (MI) model in SD rats was developed by ligation of the left anterior descending (LAD) coronary artery. Conditioned medium (CdM) collected from MSC Wnt11 (CdM Wnt11 ) was injected immediately into the ischemic border area. Rats treated with CdM Wnt11 showed a significantly improved cardiac function. Furthermore, CdM Wnt11 increased blood vessel density and regional blood flow in ischemic myocardium. Fluorescent immunostaining showed that both vWF-positive single endothelial cell numbers and microvessel numbers were significantly increased in CdM Wnt11 treated myocardium. In vitro studies showed that CdM Wnt11 significantly increased number and length of capillary-like structure (CLS) formation and promoted human umbilical vein endothelial cells (HUVECs) migration. These results were confirmed by directly treating HUVECs with recombinant Wnt11 proteins (5μg/ml) or over-expressing Wnt11 in HUVECs (H Wnt11 ). However, these effects could be abolished by using a Wnt11 neutralizing antibody. Immunostaining and Western blotting results revealed upregulation of p-pan-PKC and p-JNK in H Wnt11 . Real-time PCR analysis indicated that the expression of novel PKCs including PKCδ, PKCε, PKCη, and PKCθ was significantly upregulated in H Wnt11 . Moreover, the effect of Wnt11 on CLT formation and HUVECs migration were abrogated when the JNK inhibitor SP600125 at 5 μM and the PKC inhibitor Calphostin-C at 0.1 μM were added to the culture system. Conclusion: Our data suggests, for the first time, that Wnt11 in MSC-derived media improved cardiac function and promoted angiogenesis in ischemic myocardium. The effect of Wnt11 in angiogenesis is mediated by novel PKCs, with JNK as its downstream mediator.