Abstract 18703: Novel MicroRNA 874 is a Unique Angiogenic Modulator of Stem Cell Paracrine Factor Epidermacan

Abstract

Introduction: We have previously shown that treatment with epidermacan a novel stem cell derived paracrine factor promoted angiogenesis in vitro accompanied with an increase in angiogenic miRNA expression. Here, we identified novel microRNAs regulated by epidermacan and validated their importance in modulating the epidermacan angiogenic effects in vitro and in vivo . Methods and Results: Increased levels of pro-angiogenic miRNAs such as miR-10b, miR-21 and miR-424 were discovered in MicroRNA profiling of HUVEC cells treated with epidermacan and then confirmed by qRT-PCR. We also identified miR-874, a relatively unknown miRNA but for its role as a tumor suppressor. Investigation of the downstream microRNA/gene networks revealed that epidermacan regulated known angiogenic genes such as VEGFC, TBX1, HIF-1a and NRP1 . Importantly, treatment with epidermacan decreased the levels of HoxD10, a known target of miR-10b in thrombin induced angiogenesis. Inhibition of miR-10b or miR-874 via anti-miR transfection repressed the effects of epidermacan in HUVEC tube formation on matrigel. Interestingly inhibition of miR-10b also reversed the angiogenic effects of VEGF; however inhibition of mir-874 only suppressed the effects of epidermacan, suggesting miR-874 as an epidermacan specific target. The fli:GFP transgenic zebrafish model was used to study the role of epidermacan in angiogenesis in vivo . Morpholino against zebrafish epidermacan gene was injected into the yolk of fli:GFP transgenic zebrafish embryos at 1-4 cell stage, and larvae were collected at 4 days post fertilization to monitor vessel development. The trunk vascular network was disorganized in larvae injected with the morpholino of epidermacan compared to uninjected control. The vascular defects were rescued by co-injection of 100 pg mRNA of human NDNF (epidermacan homolog), suggesting that vessel malformation is specifically due to knockdown of epidermacan. Conclusion: Our data confirm the role of epidermacan as an angiogenic factor and identify the novel miR-874 as specific modulator of epidermacan’s angiogenic effect. Further analysis to elucidate the microRNA/gene networks downstream of epidermacan will provide new insights on the regulatory networks of angiogenesis.