鈣依賴性氯離子通道TMEM16A之抑制與促進對於代謝症候群誘發過動性與不收縮性膀胱平滑肌的影響

洪梵菁、張宏江¹、謝汝敦¹、郭育成

台北市立聯合醫院 泌尿科；¹台灣大學醫學系 泌尿科

The Effects of Inhibition and Activation of TMEM16A Calcium-Activated Chloride Channels on the Smooth Muscle Tissue in Metabolic Syndrome Induced Overactive Bladder and Acontractile Detrusor

Fan-Ching Hung, Hong-Chiang Chang¹, Ju-Ton Hsieh¹, Yuh-Chen Kuo

Department of Urology, Taipei City Hospital, Taipei, Taiwan;

Department of Urology¹, School of Medicine, National Taiwan University, Taipei, Taiwan

Purpose: Current treatment for overactive bladder (OAB) includes oral antimuscarinics, beta 3 agonists and so on. However, there is still some patients having poor therapeutic results and needing new treatment options. The established causes of underactive bladder include neurogenic, myogenic, aging, and medication side effects. Currently, the pharmacotherapy for acontractile detrusor (AD) is still disappointing. During the past years, our studies have linked chloride channels to OAB. Significantly increased molecular expressions of CLC-3 and CLCA4 chloride channels on bladder smooth muscle has been disclosed using a cyclophosphamide (CYP) induced rat OAB model. Increased voiding frequency and non-voiding contractions in CYP-induced OAB rat could be ameliorated by intravesical instillation of chloride channel blockers, implying the potential therapeutic effect of calcium activated chloride channel blockers (CaCC) on OAB. TMEM16A was recently identified as a CaCC. Evidence has also been reported for TMEM16A involvement in intestinal and vascular smooth muscle contraction. However, the role of TMEM16A on bladder smooth muscle has not been elucidated. We therefore implement a fructose feeding rat (FFR) model of metabolic syndrome to investigate the effects of inhibition or activation of TMEM16A CaCC on bladder smooth muscle tissue between normal, metabolic syndrome induced OAB and AD in rats.

Materials and Methods: 50 FFRs were fed a fructose rich diet while 25 control animals received standard rat chow for 6 months. Based on the results of cystometric presentation at month 6, FFRs were categorized into 3 groups: NDF group (normal detrusor function), OAB group and AD group. Then the urinary bladder was harvested. We conducted a strategy using organ bath isometric tension experiments to involve: ①Inhibitory concentration-response curve (CRC) of traditional chloride channel blockers (niflumic acid (NFA)) and TMEM16A blockers (T16inh-A01 and N-((4-methoxy)-2-naphthyl)-5-nitroanthranilic acid (MONNA)) on carbachol (CCh)-induced tonic contractions in control, NDF and OAB rats. ②CRC of TMEM16A activator (N-(2-methoxyethyl)-N-(4-phenyl-2-thiazolyl)- 2,3,4-trimethoxybenzeneacetamide (Eact ))-induced tonic contractions in control, NDF and AD rats. The results of inhibitory CRC and IC50 for TMEM16A blockers and CRC for TMEM16A activator among control, NDF, OAB and control, NDF, AD groups were compared.

Results: ①The CCh-induced contractions (at ED80) could be inhibited by pretreatment of NFA, T16inh-A01 and MONNA in a concentration-dependent manner in control, NDF and OAB groups. The IC50 of NFA, T16inh-A01 and MONNA in the three groups are shown in Table. The inhibitory effects of TMEM16A blockers (T16inh-A01 and MONNA) on CCh-induced contraction are more potent than those of traditional chloride channel blockers (NFA) In addition, there was a significant greater inhibition of tension for NFA (10^-4~10^-1M), T16inh-A01 (10^-5~10^-2M) and MONNA (10^-4~10^-1M) in OAB group than in control group. Also, there was a significant greater inhibition of contraction percentage in OAB group than in control group using T16inh-A01, MONNA (at the concentration of 10^-4M) but not NFA. ②Eact could induce tonic contraction on isolated rat detrusor in control, NDF, and AD groups. The contractility increased as the concentration of Eact increased. The ED80 for Eact was 1.7x10-5M.

Conclusions: This study demonstrated the inhibitory effects of TMEM16A blockers and the activating effects of Eact on bladder smooth muscle contraction in metabolic syndrome induced OAB and AD rats respectively, indicating the novel therapeutic target of TMEM16A CaCC for OAB and AD.