Compound 48/80 alters mechanical compliance via activation of Gelatinases (MMP-2 and -9) in an ATP-dependent manner

Abstract ID 92717 Poster Board 347 Many lower urinary tract symptoms (LUTS) that arise due to alteration in mechanical compliance of the bladder are also accompanied by increased urothelial signaling. While traditionally assumed to be a high-resistance barrier, urothelial cells can respond to inflammatory insult, signal to cells in the bladder wall, and potentiate the downstream release and activation of matrix metalloproteases (MMPs). MMPs are responsible for degrading components of the extracellular matrix (ECM) and causing bladder wall remodeling. Previously, we discovered that the Mas-related G protein-coupled receptor B2 (Mrgprb2) agonist Compound 48/80, increased bladder wall compliance in an MMP dependent manner. However, the specific MMPs involved and their activation state post exposure to Compound 48/80 is not understood. We hypothesized that Mrgprb2 activation causes breakdown of collagen in the bladder ECM via activation of MMP-2 and MMP-9. Whole mouse bladders from C57Bl/6 mice were dissected and mounted in a custom designed pentaplanar reflected image macroscopy (PRIM) system for simultaneous measurement of transient pressure events, wall stress, and wall stretch during ex vivo bladder filling. Bladders were exposed to Compound 48/80 (10 μg/mL) in absence or presence of the MMP-9 inhibitor JNJ-0966 (500 nM) or MMP-2 inhibitor ARP-100 (200 nM). MMP-2 and MMP-9 expression and activity in the bladders exposed to either vehicle or Compound 48/80 was also determined using Western blot analysis and gelatin zymography respectively. ARP-100 and JNJ-0966 completely abolished the increase in compliance caused by Compound 48/80. However, they had no effect on the increases in transient pressure events caused by Compound 48/80. Western blot analysis showed no difference in the expression of pro-MMP-2/MMP-2 and pro-MMP-9/MMP-9 between bladders that were exposed to vehicle or Compound 48/80 prior to protein isolation. The activity level of pro-MMP-2 and MMP-2 was increased in bladders that were exposed to Compound 48/80. These findings suggest that Compound 48/80 brings about MMP activation which drives collagen degradation in the bladder wall. Future experiments will study the mechanistic pathway that leads to MMP activation after exposure to Compound 48/80.