The long term objective of this proposal is to elucidate the mechanisms by which the mammalian urinary bladder epithelium maintains the urine composition near constant (bladders barrier function) without having its viability or underlying tissues' viability compromised. The mammalian urinary bladder has adopted at least two strategies to perform this function. The first is that the urine facing membrane (apical membrane) of this epithelium and the tight junctions (which bind the individual cells together into a planar array at the apical-lateral membrane interface) are relatively impermeable and resistant to the components found in urine. These two structures (apical membrane and tight junctions) effectively isolate the cell cytoplasm, epithelial basolateral membrane and the underlying cell layers from the urine. Second the bladder epithelium maintains a minimum ratio for apical membrane surface area to urine volume, by the withdrawal and insertion of cytoplasmic vesicles at the apical membrane. Interstitial cystitis is a bladder disorder of unknown etiology, but characterized by inflammation, glomerulations, Hunner's ulcers, fibrosis, loss of urothelial barrier function and in some instances the presence of lymphocytes or lymphocyte secretory products.
The specific aim of this grant is to investigate the effect of extracellular proteins (e.g., those secreted by eosinophils) on urothelial integrity. Previous studies have established that cationic proteins (proteins with a net positive charge such as protamine and polylysine) can compromise the barrier function of the mammalian urinary bladder. In addition, preliminary results (from this laboratory) have demonstrated that cationic proteins which can be secreted by eosinophils, found in semen (histones) or instilled in the bladder as an antibiotic, compromise the barrier function of the urinary bladder epithelium. The working hypothesis is that these proteins alter the apical membrane permeability, making it leaky to both cations and anions. This results in an increase influx of cations and anions, an obligate influx of water which results in epithelial cell lysis. Electrophysiological methods and confocal microscopy will be used to determine the mechanism(s) by which these proteins cause cell lysis, investigate other possible sites of protein action and then develop a method to counter the effects of these proteins on the urinary bladder epithelial cells.
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