Soluble guanylyl cyclase (sGC) degradation and impairment of nitric oxide-mediated responses in urethra from obese mice: reversal by the sGC activator BAY 60-2770.

Obesity has emerged as a major contributing risk factor for overactive bladder (OAB), but no study examined urethral smooth muscle (USM) dysfunction as a predisposing factor to obesity-induced OAB. This study investigated the USM relaxant machinery in obese mice and whether soluble guanylyl cyclase (sGC) activation with BAY 60-2770 [acid 4-({(4-carboxybutyl) [2-(5-fluoro-2-{[4-(trifluoromethyl) biphenyl-4-yl] methoxy} phenyl) ethyl] amino} methyl) benzoic] rescues the urethral reactivity through improvement of sGC-cGMP (cyclic guanosine monophosphate) signaling. Male C57BL/6 mice were fed for 12 weeks with a high-fat diet to induce obesity. Separate groups of animals were treated with BAY 60-2770 (1 mg/kg per day for 2 weeks). Functional assays and measurements of cGMP, reactive-oxygen species (ROS), and sGC protein expression in USM were determined. USM relaxations induced by NO (acidified sodium nitrite), NO donors (S-nitrosoglutathione and glyceryl trinitrate), and BAY 41-2272 [5-cyclopropyl-2-[1-(2-fluoro-benzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]-pyrimidin-4-ylamine] (sGC stimulator) were markedly reduced in obese compared with lean mice. In contrast, USM relaxations induced by BAY 60-2770 (sGC activator) were 43% greater in obese mice (P < 0.05), which was accompanied by increases in cGMP levels. Oxidation of sGC with ODQ [1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one] (10 µM) potentiated BAY 60-2770-induced USM responses in the lean group. Long-term oral BAY 60-2770 administration fully prevented the impairment of USM relaxations in obese mice. Reactive-oxygen species (ROS) production was enhanced, but protein expression of ß1 second guanylate cyclase subunit was reduced in USM from obese mice, both of which were restored by BAY 60-2770 treatment. In conclusion, impaired USM relaxation in obese mice is associated with ROS generation and down-regulation of sGC-cGMP signaling. Prevention of sGC degradation by BAY 60-2770 ameliorates the impairment of urethral relaxations in obese mice.

Accumulated endogenous nitric oxide synthase inhibitors in inhibiting urethral relaxation following estrogen supplementation in ovariectomized rabbits.

PURPOSE: We investigated the possible role of the endogenous nitric oxide (NO) synthase (NOS) inhibitors N-monomethyl-L-arginine (L-NMMA) and asymmetrical N, N-dimethyl-L-arginine (ADMA) in inhibiting urethral relaxation following estrogen supplementation in ovariectomized rabbits. MATERIALS AND METHODS: A total of 16 mature Japanese White female rabbits were divided into 2 groups. In the control group rabbits were sacrificed 2 weeks after bilateral ovariectomy. In the estrogen group estradiol was administered subcutaneously for 2 weeks with the aid of sustained release pellet from 2 weeks after ovariectomy until sacrifice. Isolated urethra was cut into transverse strips for functional study and processed to determine endogenous NOS inhibitors, NOS activity, dimethylarginine dimethylaminohydrolase (DDAH) activity as a metabolizing enzyme of endogenous NOS inhibitors and cyclic guanosine monophosphate production. RESULTS: Electrical field stimulation produced NO mediated and neurogenic relaxation of the urethral strip in the presence of guanethidine and atropine under contraction with phenylephrine. Relaxation was significantly decreased in the estrogen group and accompanied by decreased cyclic guanosine monophosphate production. Sodium nitroprusside induced relaxation was not different between the 2 groups. The content of L-NMMA plus ADMA in the urethra was significantly increased in the estrogen group. Ca dependent NOS activity in the urethra remained unaffected. DDAH activity was significantly lower in the estrogen group. CONCLUSIONS: Estrogen supplementation leads to decreased NO mediated and neurogenic urethral relaxation through the accumulation of L-NMMA and ADMA in the urethra. The accumulation of NOS inhibitors is possibly brought about by impaired DDAH activity.

Localization of 11beta-hydroxysteroid dehydrogenase types 1 and 2 in the male reproductive tract.

The action of glucocorticoids in target tissues is dependent on the local expression of glucocorticoid receptors and two 11beta-hydroxysteroid dehydrogenase (11beta-HSD) enzymes, 11beta-HSD1 and 11beta-HSD2, which interconvert active and inactive glucocorticoids. This study examined expression of the 11beta-HSD enzymes in the male reproductive tract of the adult rat. 11beta-HSD1 was immunolocalized to the apical region of principal epithelial cells of the caput epididymis, with the less numerous clear cells devoid of signal. Epididymal 11beta-HSD1 expression was confirmed by Western blot analysis, with immunoreactive species identified at 34 kDa (the expected size for 11beta-HSD1) and at approximately 48 kDa. 11beta-HSD bioactivity was readily detectable in the epididymis, with 11-oxoreductase activity clearly the favored reaction (as observed in liver), consistent with 11beta-HSD1 expression. The epithelium of the vas deferens, seminal vesicle, and penile urethra were also immunopositive for 11beta-HSD1, as were smooth muscle cells of the vas deferens and penile blood vessels. 11beta-HSD2 was also immunolocalized to the epididymal epithelium, but its distribution was complementary to that of 11beta-HSD1 (i.e. clear cells showing intense 11beta-HSD2 staining but principal cells devoid of signal). 11beta-HSD2 was also present in the corpora cavernosa of the penis but not in other tissues. In conclusion, the differential expression of 11beta-HSD1 and 11beta-HSD2 throughout the male reproductive tract suggests that these enzymes locally modulate glucocorticoid and mineralocorticoid actions, particularly in the epididymis and penile vasculature.