Mechanisms of contractile dysfunction in the obstructed bladder: Role of desmin and vimentin

摘要 下尿路功能障碍(LUTD)是一种常见疾病，其发病率和患病率随着年龄的增加而增加。 人口年龄和目前的治疗方法都是无效的。LUTD与以下因素密切相关 良性前列腺增生症(BPH)部分继发性膀胱平滑肌(BSM)肥大 膀胱出口梗阻(PBOO)。BPH和BSM功能障碍的分子发病机制尚不清楚 理解；填补我们知识中的这一空白可能会导致新的治疗靶点的确定和 更有效的LUTD药物或管理策略。我们和其他人已经证明了收缩 梗阻性膀胱功能障碍、线粒体ROS增加和BSM ATP水平降低 与中间丝(IF)蛋白结蛋白和波形蛋白的过度表达有关。上一首 研究和初步数据表明，JNK2是BSM收缩的主要效应器 结蛋白和波形蛋白过度表达所致的功能障碍，并证明了一种IF/JNK2- 依赖性机制参与了膀胱出口梗阻的收缩功能障碍。在当前 我们通过基因芯片发现线粒体蛋白G0S2在人类体内有显著的诱导作用 小鼠BSM条带和细胞过度表达结蛋白和波形蛋白。我们的额外初步数据进一步 证明抑制G0S2的表达降低了磷酸化JNK的水平，增加了线粒体的ATP， 并减少结蛋白和波形蛋白过表达的小鼠和人BSM条中ROS的产生 和细胞。进一步，我们证明了G0S2与电压依赖阴离子通道(VDAC)的相互作用。 VDAC促进线粒体外膜与腺嘌呤结合的ATP/ADP交换 核苷酸易位酶和线粒体肌酸激酶。G0S2与VDAC的绑定可能会扰乱 从而降低线粒体的ATP水平，增加线粒体的ROS 制作。我们假设线粒体G0S2介导结蛋白和波形蛋白诱导的BSM收缩 通过磷酸化JNK2引起的功能障碍。我们进一步假设，G0S2与VDAC的相互作用促进了 线粒体ROS的产生和ROS诱导的磷酸化JNK2参与了收缩功能障碍。 为了解决这些假设，我们设计了三个具体目标。在目标1中，我们将确立 线粒体G0S2在IF蛋白过度表达诱导的BSM收缩功能障碍中的作用在《目标2》中我们将 确定蛋白质过度表达引起的BSM收缩功能障碍是否与G0S2和VDAC有关 互动。在目标3中，我们将使用活体模型来建立G0S2和JNK2在小鼠PBOO中的作用 诱发的BSM收缩功能障碍：我们的研究有望描绘出一种收缩的机制 G0S2介导的功能障碍，从而确定治疗PBOO/LUTD的靶点。

Abstract Lower urinary tract dysfunction (LUTD) is a common disease whose incidence and prevalence increase as the population ages and the the current therapeutic approaches are ineffective. LUTD is strongly associated with bladder smooth muscle (BSM) hypertrophy secondary to benign prostatic hyperplasia (BPH)-induced partial bladder outlet obstruction (PBOO). The molecular pathogenesis of BPH and BSM dysfunction is poorly understood; filling this gap in our knowledge will likely lead to the identification of new therapeutic targets and more effective LUTD drugs or management strategies. We and others have demonstrated that the contractile dysfunction, increased mitochondrial ROS and reduced ATP levels of BSM in the obstructed bladder is associated with the overexpression of the intermediate filament (IF) proteins desmin and vimentin. Previous studies as well as preliminary data presented herein suggest JNK2 as a major effector of the BSM contractile dysfunction induced by the overexpression of desmin and vimentin and demonstrate that an IFs/JNK2- dependent mechanism contributes to the contractile dysfunction in bladder outlet obstruction. In the current proposal we have identified via microarray significant induction of the mitochondrial protein G0S2 in human and murine BSM strips and cells overexpressing desmin and vimentin. Our additional preliminary data further demonstrate that inhibition of G0S2 expression decreases phospho-JNK levels, increases mitochondrial ATP, and decreases the ROS production in desmin- and vimentin- overexpressing murine and human BSM strips and cells. Further, we demonstrate the interaction of G0S2 with voltage-dependent anion channel (VDAC). VDAC promotes ATP/ADP exchange across the mitochondrial outer membrane in association with adenine nucleotide translocase and mitochondrial creatine kinase. G0S2 binding to VDAC presumably disrupts the ATP/ADP exchange thereby, reducing the mitochondrial ATP level and increasing the mitochondrial ROS production. We hypothesize that mitochondrial G0S2 mediates desmin- and vimentin-induced BSM contractile dysfunction via phospho-JNK2. We further hypothesize that G0S2 interaction with the VDAC promotes the mitochondrial ROS production and the ROS-induced phospho-JNK2 contributes to the contractile dysfunction. Three Specific Aims are designed to address these hypotheses. In Aim 1, we will establish the role of mitochondrial G0S2 in IF protein overexpression-induced BSM contractile dysfunction. In Aim 2 we will determine whether IF protein overexpression-induced BSM contractile dysfunction is due to G0S2 and VDAC interaction. In Aim 3 we will employ in vivo models to establish the role of G0S2 and JNK2 in murine PBOO induced BSM contractile dysfunction: We expect our studies to delineate a mechanism of contractile dysfunction mediated by G0S2 and thus identify therapeutic targets for the treatment of PBOO/LUTD.