The overall objective of this grant proposal is to define the molecular events associated with the formation of trabeculated, fibrotic bladders. The formation of a noncompliant bladder is a major health problem today and can lead to upper urinary tract problems and ultimately to kidney failure. To begin to understand the nature of this problem, the expression of the major connective tissue proteins and their respective genes will be compared in suitable control and noncompliant tissue samples. Samples will be obtained from patients who have been studied urodynamically as a part of their preoperative care. The ability to correlate biochemical and molecular data with urodynamic studies is an extremely part of this proposal and one which should provide new and important information relating clinical and biochemical changes as fibrosis occurs. Using approaches at both the protein and mRNA levels, the synthesis of the major fiber forming collagens will be qualitatively and quantitatively assessed to determine their contribution to the etiology of bladder fibrosis. Biochemical analysis of type I, III and IV collagens will allow comparison of accumulated protein with appropriate gene expression which will be evaluated by Northern and slot blot hybridization analyses. To begin to associate these changes with a specific cell type, in situ hybridization with cDNA probes for type I and III collagen will be carried out. These studies will permit us to determine which cell type in the bladder wall is likely showing alteration in matrix gene expression as well as its location within the wall, i.e., the lamina propria, the detrusor layer or other areas. These studies will also show is there is a differential expression within discrete areas of the bladder wall or whether the response is a general one shared by all areas or cell types uniformly within an anatomical tissue layer. In the third aim, cells from noncompliant bladder tissue will be isolated and cultured in vitro to begin to investigate the regulatory and control mechanisms which underlie the changes observed clinically. Studies in aims one and two are necessary prerequisites to the studies outlined in aim three because the information obtained will begin to define she molecular mechanisms responsible for the fibrotic changes. Experiments aimed at distinguishing between transcriptional, translational or other mechanisms will provide new information on bladder wall disorders and how matrix gene expression may play a role in altering the normal physiologic compliance.
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