(Directly taken from the application) Polycystic kidney disease (PKD) is marked by several abnormalities, including tubular epithelial cell hyperplasia, abnormal differentiation, and altered tubular cell transport. The cpk strain has been utilized as a model of autosomal recessive polycystic kidney disease where the phenotypic expression of cystic renal disease occurs in the homozygous mice. Sphingolipids, in particular ceramides, have been implicated as important mediators in a variety of cellular processes. These include growth, differentiation, interaction with extracellular matrix, and ion transport. Our recent work has demonstrated a role for ceramides in growth and signaling events in several renal epithelial cell lines, including the Madin-Darby canine kidney and MCT mouse proximal tubule cell lines. MDCK cells have been utilized as an in vitro model of cyst formation. We have studied the glycolipid content and associated enzymatic activities of the kidneys from cpk/cpk mice and their phenotypically normal litter mates. The neutral glycolipids, including glucosylceramide and lactosylceramide, displayed a striking increase in 3 week old cpk/cpk mice as did the acidic lipid, ganglioside GM3. However, a correspondingly significant decrease in sulfoglycolipid and ceramide concentration was also observed in the cpk/cpk kidneys. Glucosylceramide synthase activity was higher in the kidneys of the cpk/cpk mice than in those of the controls. While sulfolipid and glucosylceramide concentrations were not different at one and two weeks of age, ceramide concentrations were significantly reduced prior to the development of severe azotemia. This alteration may be associated with the delayed differentiation of cystic collecting duct epithelia. These results suggest that ceramide may play a potentially important role in the proliferative and transport abnormalities associated with cystic renal disease and the development of azotemia. In the present application we propose to investigate the biochemical basis for the decrease in ceramide content in the kidneys of cpk/cpk mice. We propose as the primary hypothesis for these studies that ceramide regulates the arrested differentiation observed in the cystic phenotype of the cpk/cpk mouse kidney.
The Specific Aims are: 1. identify the metabolic basis for the lower levels of ceramide in the cpk kidneys; 2. evaluate the role of ceramide in growth of cultured cystic epithelium; 3. determine the role of ceramide signaling in physiologically important events in cell cycle regulation of cultured cystic epithelia; and 4. evaluate the effects of the modulation of ceramide levels in cpk mice in vivo by the use of novel inhibitors of ceramide metabolism including blockers of glucosylceramide synthase, 3-ketodihydrosphingosine synthase, ceramide synthase and ceramidase.
