ABSTRACT The vasculature consists of vessels of different sizes that are arranged in a hierarchical pattern. Two cell populations work in concert to establish this pattern during embryonic development and adopt it to changes in blood flow demand later in life: endothelial cells that line the inner surface of blood vessels, and adjacent vascular mural cells, including smooth muscle cells and pericytes. Despite recent progress in elucidating the signalling pathways controlling their crosstalk, much debate remains with regard to how mural cells influence endothelial cell biology and thereby contribute to the regulation of blood vessel formation and diameters. In this Review, I discuss mural cell functions and their interactions with endothelial cells, focusing on how these interactions ensure optimal blood flow patterns. Subsequently, I introduce the signalling pathways controlling mural cell development followed by an overview of mural cell ontogeny with an emphasis on the distinguishing features of mural cells located on different types of blood vessels. Ultimately, I explore therapeutic strategies involving mural cells to alleviate tissue ischemia and improve vascular efficiency in a variety of diseases.