This grant focuses on one of the most critical yet most poorly understood mechanistic events in urolithiasis, the attachment of urinary crystals by the epithelium of the kidney papillary tip which then serves as maturation sites for urinary stone growth. Stone research literature supports a relationship between tissue injury and stone disease. Their studies on crystal attachment to inner medullary collecting duct cells (IMCD) have demonstrated that calcium oxalate monohydrate (COM), hydroxyapatite (HA), and uric acid (UA) crystals attach to IMCD cells cultures in a structurally specific, saturable and inhibitable manner. Crystal attachment appears to be based on specific atomic interactions between molecular arrays on the surfaces of stone crystals and on the surfaces of lipid rich domains on the surfaces of IMCD cells. They hypothesize that renal epithelial cells will only bind kidney stone crystals if there is a specific perturbation in outer plasma membrane leaflet phospholipid composition and/or structure resulting in the formation of phospholipid domains which are structurally complementary to the molecular arrays on crystal surfaces. Such changes might be associated with renal epithelial injury.
Specific Aim I. They propose to correlate the attachment of COM, calcium oxalate dihydrate (COD), HA, and UA crystals to IMCD cells with specific phospholipid composition and structure in the plasma membrane of these cells. They also propose to define the changes in the composition and structure of the phospholipid domains as a function of crystal attachment following PS and PI phospholipid enrichment.
Specific Aim II. They propose to study crystal attachment to IMCD cells as a function of I) prior exposure of IMCD cells to ionic conditions at or near supersaturation conditions as would be expected in stone forming patients demonstrating recurrent episodes of crystalluria, ii) exposure of IMCD cells to crystals coated with selective lipids or urinary protein macromolecules identified as potential stone growth inhibitors or promoters, and iii) exposure of IMCD cells to one stone crystal type followed by a second exposure of a second crystal type to assay for lipid domain composition or structure perturbations as primers for more effective crystal attachment.
Specific Aim III. They propose to crystallographically model the molecular structures on the surfaces of all five crystals as well as the structures of an assembly of phospholipid headgroups both as single as well as admixed phospholipid structures. They also propose to epitaxially analyze the dimensional basis as well as the specific molecular bonding matches that would support long range interactions between the various stone crystals and molecular structure in lipid-rich domains in the outer leaflet of IMCD cell plasma membranes.
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