Identification and suppression of specific genes of urinary incontinence using RNAi

脑梗死（CI）引起的膀胱过度活动（BO）的发展被认为需要脑桥排尿中心（PMC）的RNA转录。我们之前报道过环氧化酶-2 (COX-2) mRNA的表达是由PMC中NMDA （n -甲基- d -天冬氨酸）受体的活性介导的，并且是BO发展所必需的。本研究旨在探讨前列腺素（PG） E或D合成酶、COX-2下游基因的表达以及PGE2和D水平是否与BO有关。首先，我们使用COX-2敲除的背景小鼠（C57-BL/6J）来检验其对CI是否具有与大鼠模型相同的反应。为了证实COX-2的表达对BO的发展至关重要，我们认为在小鼠PMC和左大脑中动脉（MCA）闭塞中，体内RNAi是必要的。方法：对雌性SD大鼠和C57-BL/6J小鼠进行MCA闭塞诱导脑梗死（CI）。对清醒的动物进行膀胱造影检查。在中动脉闭塞或假手术（SO）后0.25、1、3、5、12和24小时，从桥背被（DPT）获得更多的标本。实时荧光定量PCR检测PGES和PGDS在DPT中的表达。为了确定PGE2是否对BO的诱导至关重要，我们在氨基甲酸乙酯麻醉大鼠的PMC或导水管周围灰质（PAG）中微量注射PGE2。此外，EP1受体拮抗剂ONO-8711在PGE2滴注前静脉或脑室内注射。结果：MCA闭塞后1-24小时，CI大鼠和小鼠膀胱容量明显降低。MCA闭塞1小时后，与SO相比，PGES和PGDS mRNA表达显著增加。在MCA闭塞后至少12小时，PGES的表达始终高于SO大鼠。MK-801预处理可抑制膀胱过度活动的发生，并显著降低DPT中PGES mRNA的表达。MCA闭塞后3 ~ 5小时PGE2水平升高。脑室和脑桥背被微量注射PGE2可增加膀胱容量。PGE2进入PAG显著降低膀胱容量，ONO-8711可拮抗膀胱容量。MCA闭塞导致C57-BL/6J小鼠DPT膀胱容量明显减少，COX-2/PGES表达增加。结论：MCA闭塞后BO的发生与DPT中COX-2和PGES mRNA表达的增加有关。PAG中PGE2水平升高被认为在CI所致BO的发展中起重要作用。少

Development of bladder overactivity (BO) caused by cerebral infarction (CI) is believed to require RNA transcription in the pontine micturition center (PMC). We previously reported that the expression of cyclooxygenase-2 (COX-2) mRNA was mediated by the activity of an NMDA (N-methyl-D-aspartate) receptor in the PMC and necessary for the development of BO. This study was undertaken to examine whether the expression of prostaglandin (PG) E or D synthase, downstream gene of COX-2, and levels of PGE2 and D were related to BO. To begin with, background mouse (C57-BL/6J) of COX-2 knockout was used to examine whether it has the same response to CI as a rat model. To confirm that the expression of COX-2 is essential for the development of BO, in vivo RNAi in the mice PMC and left middle cerebral artery (MCA) occlusion are thought to be necessary.Methods : Cerebral infarction (CI) was induced by MCA occlusion in female SD rats and C57-BL/6J mice. Awake animals were cystometrically examined. Spe … More cimens were obtained from the dorsal pontine tegmentum (DPT) 0.25, 1, 3, 5, 12, and 24 hours after MCA occlusion or a sham operation (SO). Expressions of PGES and PGDS in the DPT were monitored with real-time PCR. To determine whether PGE2 is essential for the induction of BO, microinjection of PGE2 into the PMC or periaquaeductal gray (PAG) was performed in urethane anesthetized rats. Furthermore, ONO-8711, EP1 receptor antagonist was administered intravenously or intracerebroventriculaly before PGE2 instillation.Results : Bladder capacity of CI rats and mice was significantly reduced 1-24 hours after MCA occlusion. One hour after MCA occlusion, PGES and PGDS mRNA expression had significantly increased, as compared to that in SO animals. PGES expressions remained consistently higher than those in SO rats at least 12 hours after MCA occlusion. Pretreatment with MK-801 inhibited the development of bladder overactivity and significantly reduced the expression of PGES mRNA in the DPT. The level of PGE2 increased 3 to 5 hours after MCA occlusion at the DPT. Microinjection of PGE2 into the cerebral ventricle and dorsal pontine tegmentum increased bladder capacity. PGE2 into the PAG significantly decreased bladder capacity, which was antagonized by the administration of ONO-8711. MCA occlusion produced a significant reduction in bladder capacity and increased expression of COX-2/PGES in the DPT of C57-BL/6J mice.Conclusion : These results indicate that the development of BO following MCA occlusion is mediated by an increase in COX-2 and PGES mRNA expression in the DPT. Increase in PGE2 level in the PAG is believed to play an important role in the development of BO caused by CI. Less