Primary cilia are small microtubule-based appendages expressed on nearly every cell type. Genetic mutations that affect the construction or function of the cilium result in a spectrum of disorders termed ciliopathies, which manifest in a range of phenotypes including neural defects, skeletal defects, situs inversus, and renal, hepatic, and pancreatic cysts. The most common ciliopathy, Polycystic Kidney Disease (PKD), affects more than 13 million individuals worldwide and is characterized by the formation of large renal and hepatic cysts. Despite decades of research, treatment of PKD in the U.S remains limited to dialysis and kidney transplants, necessitating more research into the disease. Recently, it has emerged that macrophages and the inflammatory response promote PKD progression, raising the possibility that renal primary cilia may regulate the innate immune response. Our study aims to determine how mutations to the primary cilium affect kidney function and the local immune response to promote cyst formation. To study the effects of ciliary dysfunction, we utilize mice with conditional alleles for polycystin-2 (PC2), a cationic channel mutated in human PKD, as well as IFT88, a protein necessary for cilium biogenesis and homeostasis. Previous work has demonstrated that deletion of either of these genes prior to postnatal day 12 (P12) in mice induces rapid cyst formation, while deletion after P14 results in slow, focal cyst formation that takes several months to form. This has gone on to suggest the presence of a ?critical window? in which functional primary cilia are necessary for proper kidney function. Interestingly, rapid cyst formation is restored to adult induced mice following renal injury. Our lab has recently shown that specific subpopulations of macrophages are present in the juvenile kidney and that this macrophage profile shifts around the critical window. Moreover, the juvenile profile returns to the adult induced kidney following injury, suggesting that this subpopulation associates with rapid cyst formation and may promote cyst growth. These data lead to our overall hypothesis that disruption of cilia structure or function leads to a dysregulated intercellular signaling microenvironment and inflammatory response, which drives cyst formation and disease progression. We intend to test this hypothesis through utilization of novel intravital microscopy techniques.
Aim 1 will determine how dysfunctional primary cilia affect the repair processes following injury in the kidney through the use of mice with fluorescently labeled primary cilia (sstr3GFP) and through the use of dextran injection that will label actively filtrating nephrons.
Aim 2 will elucidate the effects of ciliary mutations on macrophage profile and their intercellular actions following kidney injury. This will be accomplished by injuring transgenic mice with fluorescently labeled macrophages (Ccr2RFP;Cx3cr1GFP), while using dextran injection to label active tubules. Data gathered from this research will be beneficial in determining how mutations to the cilium exert global effects in the kidney to influence renal function, macrophage accumulation, inflammation, and cyst formation.
Polycystic Kidney Disease (PKD) is a common genetic disorder that affects more than 13 million people worldwide and leads to end stage renal disease in 50% of those affected. Despite clinical need, the mechanisms of PKD remain incompletely understood and the only treatments available are dialysis and organ transplant. Data generated from this work will help address how cells of the immune system are dysregulated in PKD and how they are promoting cyst growth and disease, thus providing possible targets for therapeutic treatments.
