Inhibition of pressure induced bladder smooth muscle cell hyperplasia using CRM197

Purpose: In vivo the effects of sustained hydrostatic pressure on the bladder wall and its components are evident under physiological and pathological conditions. We previously reported that exposure of bladder smooth muscle cells to 20 and 40 cm, H(2)O hydrostatic pressure for as little as 1 hour resulted in the up-regulation of heparin binding epidermal growth factor messenger RNA in a time dependent fashion as well as in activation of the heparin binding epidermal growth factor growth factor gene. In our current study we investigated the use of CRM197 as an agent for blocking undesirable cellular level events, such as smooth muscle cell hyperplasia, eliminating the irreversible alterations in bladder and kidney function that result from chronic and/or severe bladder outlet obstruction. Materials and Methods: Control and experimental neonatal ovine smooth muscle cells were exposed to 0.3 pressure and 8.5 cm. H(2)O, respectively, for 7 days. We evaluated the mitogenic activity of the supernatant medium from bladder smooth muscle cells exposed to 8.5 cm. H(2)O for 5 days (conditioned medium) before and after the addition of 0.1 mg./ml, CRM197. Bladder smooth muscle cell apoptosis was also assessed after CRM197 exposure. Statistical analysis was performed using the Student t test with p <0.05 considered significant. Results: Exposing bladder smooth muscle cells to sustained 8.5 cm. H(2)O hydrostatic pressure for 7 days resulted in increased cell proliferation. Conditioned medium contained mitogenic activity, which was ablated after CRM197 was added. No direct toxic effect of CRM197 on bladder smooth muscle cell growth was appreciated (no apoptosis). Conclusions: We demonstrated a proliferative response of neonatal bladder smooth muscle cells after exposure to sustained hydrostatic pressure. This response was partially due to the release of heparin binding epidermal growth factor and was blocked by adding CRM197. These data support the potential use of CRM197 in drug targeted therapy for diseases involving bladder outlet obstruction.