The major goal of this proposal is to determine the cellular and molecular mechanisms responsible for the up-regulation of extracellular matrix genes and proteins, specifically types I and III collagen, in bladder fibrosis. Congenital and acquired obstructive uropathies, as the sequelae to neurologic lesions (myelomeningocele, tethered cord syndrome) or physical alteration (posterior urethral valves, BPH) can result in fibrosis of the bladder wall. We have previously demonstrated that the terminal response is an accumulation of collagens in abnormal locations and an alteration in the type III :l collagen ratio. We have shown that angiotensin II can act through a TGF-beta1 -dependent mechanism in the up-regulation of collagens, and that this occurs through another down-stream protein factor. We will test the hypothesis that extracellular matrix changes in human bladder smooth muscle cells and fibroblasts in vitro are regulated by a TGF-beta1 via a cascade mechanism involving down-stream mediator, connective tissue growth factor CTGF. We will determine if the effect of TGF-beta1 on regulation of collagenous protein occurs via CTGF RNA and protein expression, and if the CTGF effect occurs through not only synthesis, but also involves alteration in the degradation/inhibition pathway through analysis of matrix metalloproteinases and tissue inhibitors of metalloproteinases (TIMPs). In vivo, murine partial bladder outlet obstruction studies will determine if the fibrotic response is owing to a TGF-beta and CTGF dependent mechanism, using conditional gene targeting experiments whereby the CTGF gene is ablated in smooth muscle cells, CTGF transgenic animals, and in vivo antibody inhibition experiments. We will also determine whether treatment with HMG-CoA reductase inhibitors (statins) can ameliorate the fibrotic response by inhibiting the CTGF pathway, both in in vitro and in vivo models. This research seeks to target translational therapeutic strategies to attenuate or block the fibrotic response in obstructive, and possibly inflammatory, uropathies. The statins are currently in use as cholesterol lowering agents in the general population, with side effects which have been rigorously documented. The potential use of statins would be of enormous benefit to all patients with, or having the potential to develop, bladder fibrosis, since the drug has few serious side effects and has been evaluated and approved by the FDA for other uses.
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