Exogenous glycosaminoglycans coat damaged bladder surfaces in experimentally damaged mouse bladder

Abstract

Background: Interstital cystitis is often treated with exogenous glycosaminoglycans such as heparin, chondroitin sulphate (Uracyst), hyaluronate (Cystistat) or the semi-synthetic pentosan polysulphate (Elmiron). The mechanism of action is presumed to be due to a coating of the bladder surface to replace the normally present chondroitin sulphate and heparan sulphate lost as a result of the disease. This study used fluorescent labelled chondroitin sulphate to track the distribution of glycosaminoglycans administered intravesically to mouse bladder that had been damaged on the surface.

Methods: The surfaces of mouse bladders were damaged by 3 mechanisms -- trypsin, 10 mM HCl, and protamine sulphate. Texas Red-labeled chondroitin sulphate was instilled into the bladders of animals with damaged bladders and controls instilled only with saline. Bladders were harvested, frozen, and sectioned for examination by fluorescence.

Results: The normal mouse bladder bound a very thin layer of the labelled chondroitin sulphate on the luminal surface. Trypsin- and HCl-damaged bladders bound the labelled chondroitin sulphate extensively on the surface with little penetration into the bladder muscle. Protamine produced less overt damage, and much less labelling was seen, presumably due to loss of the label as it complexed with the protamine intercalated into the bladder surface.

Conclusion: Glycosaminoglycan administered intravesically does bind to damaged bladder. Given that the changes seen following bladder damage resemble those seen naturally in interstitial cystitis, the mechanisms proposed for the action of these agents is consistent with a coating of damaged bladder.

Figure 1

Damage produced by each model of bladder damage and binding of Texas Red-labelled chondroitin sulphate to mouse bladder. Images in the first two columns are transmitted light images of bladder sections stained with H&E or Acid Alcian Blue to demonstrate bladder damage. Acid Alcian Blue binds to glycosaminoglycans and therefore stains the connective tissue and the "GAG Layer," which is indicated by arrows in the control image. The fluorescence images are digitally combined images of Hoechst 33258 fluorescence (blue) to show nuclei and Texas Red-labelled chondroitin sulphate (red). Arrows have been added to the fluorescence image of the protamine-treated bladder to show the location of the urothelium. From top to bottom, the rows show the controls, bladders treated with 1 mg/ml trypsin (30 min), 1 mg/ml protamine (10 min) and 10 mM HCl (10 min.) respectively. The fluorescence image of the control was digitally enhanced to demonstrate the small amount of binding that occurred, but which was undetectable at the settings used for the other images (Table 1).