Slit/Robo Signaling On Vasculogenesis During Gastrulation

The neurogenesis and vasculogenesis are the two key events in the early embryonic development. Both primary blood vessels plexus and axon extension are guided to their termination by "specific" signaling such as early-expressing growth factor genes. And interestingly it is found that some guidance signaling molecules are shared by the both developmental processes. Slit-Robo signaling as a major repellent molecular was initially discovered to mediate axon crossing the midline of the central nervous system. The following investigation suggests that this pathway might also play an important role on other system developmental process including vasculogenesis. However, it remains unclear whether Slit-Robo signaling involve in the process of embryonic vasculogenesis.In the present study we investigated the effect of Slit-Robo on gastrula vasculogenesis using early chick embryo as an in vivo model. At first we made the detection for the protein and mRNA expression pattern of Slit/Robo in gastrulation and confirmed the positive expression of Slit-Robo in early embryo using in situ hybridization and immunohistochemistry. Blocking Slit-Robo via the knockdown of Robo1 expression with siRNA leaded to poor embryonic development especially significantly reduced the area and number of vascular plexus and converted shape of blood island. The cell-cell junction was disarranged too in presence of Robo1 siRNA. To analyze the development blood island in detail we employed a specific staining method with carmine for evaluating the developing blood island, in which the size and number of blood island in Robo1 knockdown group was significantly reduced compared with the one in control group. The result is verified with immunohistochemistry staining with VE-cadherin, a specific blood island marker.In summary, we revealed that Slit-Robo signaling plays an important role in embryonic blood vessel development. This finding will broaden our understanding of Slit-Robo in development, and the investigation of mechanism about Slit/Robo mediating formation of blood vessel plexus is carrying on in our laboratory.ConclusionsBlocking Slit/Robo signaling by the knockdown of Robo1 expression with siRNA leaded to poor embryonic vascular plexus development, suggesting that Slit-Robo signaling plays an important role in embryonic blood vessel formation.