Determining tile in vivo role of Nck in endothelial cells during cardiovascular development

Cardiovascular development is a complex process regulated by intercellular signaling mechanisms that coordinate cellular patterning during embryogenesis. The family of Nck adaptor proteins link incoming chemical stimuli at receptor tyrosine kinases on the cell surface to downstream effector molecules that regulate organization of the actin cytoskeleton network. Considering that endothelial cell migration is essential for proper vascular development, and Nck has been shown to regulate this process, we hypothesize that Nck is required for cardiovascular development. Utilizing the Cre-LoxP genetic system, we have demonstrated that deleting Nck2 expression in embryonic endothelial cells of Nck1-null mice results in lethality between embryonic day 10 and 11. These mice are characterized by specific defects to the vitelline vessel structure of the yolk sac, and the endocardial layering of the primitive heart tube. In order to complement the in vivo mutant phenotype, we have optimized a protocol for isolation of primary endothelial cell cultures from the lungs of Cre-, Nck1-/-, Nck2flx/flx adult mice, which will be used for further characterization. These results demonstrate the crucial role of Nck in endothelial cells during cardiovascular development, and may serve to further elucidate the signaling mechanisms that underlie this complex process.