Glomerular endothelial cells play pivotal roles in the maintenance of the normal structure and function of the healthy glomerulus, and also in the development of glomerular injury in kidney diseases. However, our knowledge of the mechanisms that constantly maintain and regenerate the glomerular endothelium after injury have been limited, due to the inaccessibility of this cell type and lack of in vivo research technologies. Our laboratory has recently developed novel transgenic mouse models and an intravital imaging approach using serial multiphoton microscopy (MPM) of the same glomerulus over several days-weeks to quantitatively visualize and track the fate and function of the same single glomerular endothelial cells in the intact living kidney. This technical advance provided visual clues on the development of clonal endothelial cell clusters at the glomerular vascular pole in response to dietary salt restriction and various types on glomerular injury, and suggested its control by the juxtaglomerular apparatus (JGA). In addition, the foundation of this proposal is our recent preliminary findings which suggested that macula densa (MD) cells of the JGA are novel chief organizers and master regulators of glomerular remodeling in the adult kidney. Gene profiling showed that MD cells produce novel secreted tissue remodeling factors that act in a paracrine fashion on resident progenitor cells. One of the top new MD genes responsible for this new glomerular remodeling program was identified as CCN1 (also known as Cyr61), coding for a secreted angiogenic protein that is known to target endothelial cells as well as other cell types. Our main hypothesis is that MD cells, via Wnt signaling-mediated synthesis and secretion of angiogenic proteins including CCN1, are major paracrine regulators of the glomerular endothelium. This project will use comprehensive experimental approaches including in vitro cell culture, new transgenic mouse models, fluorescently tagged cell lineages, cell fate tracking, in vivo disease models and serial MPM to investigate the angiogenic role, mechanisms, and importance of MD cells and CCN1 in glomerular endothelial maintenance and repair. The specific aims are to (i) Examine the role and mechanism of MD cells in the maintenance of normal glomerular capillary structure and function, (ii) Study the regulation of the MD cell angiogenesis promoting program, and (iii) Test for renal and cardiovascular disease modifying roles and mechanisms of MD cells.