Elevated polyamines in urothelial cells from OAB subjects mediate oxotremorine-evoked rapid intracellular calcium rise and delayed acetylcholine release

Abstract

Increased polyamine signaling in bladder urothelial cells (BUC) may play a role in the pathophysiology of overactive bladder (OAB). We quantitated intracellular polyamine levels in cultured BUC from OAB and asymptomatic (NB) subjects. We assessed whether polyamines modulated rapid intracellular calcium ([Ca(2+)]i) changes and delayed acetylcholine (ACh) release evoked by oxotremorine (OXO, a muscarinic agonist). BUC were cultured from cystoscopic biopsies. High-performance liquid chromatography (HPLC) quantitated intracellular putrescine, spermidine, and spermine levels. Five-millimeter difluoromethylornithine (DFMO), and one-millimeter methylglyoxalbisguanylhydrazone (MGBG) treatments were used to deplete intracellular polyamines. Ten micrometers of OXO were used to increase [Ca(2+)]i levels (measured by fura 2 microfluorimetry) and trigger extracellular ACh release (measured by ELISA). Polyamine levels were elevated in OAB compared with NB BUC (0.5 ± 0.15 vs. 0.16 ± 0.03 nmol/mg for putrescine, 2.4 ± 0.21 vs. 1.01 ± 0.13 nmol/mg for spermidine, and 1.90 ± 0.27 vs. 0.86 ± 0.26 nmol/mg for spermine; P < 0.05 for all comparisons). OXO evoked greater [Ca(2+)]i rise in OAB (205.10 ± 18.82% increase over baseline) compared with in NB BUC (119.54 ± 13.01%; P < 0.05). After polyamine depletion, OXO evoked [Ca(2+)]i rise decreased in OAB and NB BUC to 43.40 ± 6.45 and 38.82 ± 3.5%, respectively. OXO tended to increase ACh release by OAB vs. NB BUC (9.02 ± 0.1 vs. 7.04 ± 0.09 μM, respectively; P < 0.05). Polyamine depletion reduced ACh release by both OAB and NB BUC. In conclusion, polyamine levels were elevated twofold in OAB BUC. OXO evoked greater increase in [Ca(2+)]i and ACh release in OAB BUC, although these two events may be unrelated. Depletion of polyamines caused OAB BUC to behave similarly to NB BUC.

Alterations in Nerve-Evoked Bladder Contractions in a Coronavirus-Induced Mouse Model of Multiple Sclerosis

“…The urothelium is known to release various substances, such as ACh, ATP, nitric oxide [24] , prostaglandins, and nerve growth factor [25] , [26] . These substances could have either inhibitory or augmenting effects on muscarinic receptor-mediated detrusor contractions.…”

Mitogen‐Activated Protein Kinase and Intracellular Polyamine Signaling Is Involved in TRPV1 Activation–Induced Cardiac Hypertrophy

“…According to our previously reported method, 16 the samples were homogenized in 700 lL of 0.9% sodium chloride mixed with 10 lL (200 nmol/mL) internal standard (1, 7-diaminoheptane). To precipitate proteins, 300 lL of trichloroacetic acid (0.5 mol/L) was added to the homogenate.…”

Mouse urothelial genes associated with voiding behavior changes after ovariectomy and bladder lipopolysaccharide exposure

“…S‐adenosylmethionine decarboxylase (SAM‐DC) is an enzyme that catalyzes the decarboxylation of S‐adenosylmethionine to decarboxylated S‐adenosylmethionine, which is a substrate for production of the polyamines spermine and spermidine. The relevance of SAM‐DC in OAB was demonstrated by the finding that cultured human OAB urothelial cells contained significantly higher amounts of spermine and spermidine compared to non‐OAB urothelial cells . While increased intracellular urothelial polyamines have not been studied in the context of UTI pathogenesis, there is a potential correlate in pathophysiology of Helicobacter pyloriHelicobacter pylori gastric infections.…”

“…The relevance of SAM-DC in OAB was demonstrated by the finding that cultured human OAB urothelial cells contained significantly higher amounts of spermine and spermidine compared to non-OAB urothelial cells. 25 While increased intracellular urothelial polyamines have not been studied in the context of UTI pathogenesis, there is a potential correlate in pathophysiology of Helicobacter pyloriHelicobacter pylori gastric infections. Investigators showed that the gastric innate immune response against H. pylori infection resulted in increases in gastric epithelial polyamines leading to increased nitric oxide and reactive oxygen species which were toxic to H. pylori.…”