Mitochondrial Dysfunction in Obstructed Bladder: Role of Desmin and Vimentin

 描述(申请人提供)：下尿路功能障碍(LUTD)与良性前列腺增生症(BPH)引起的部分膀胱出口梗阻(PBOO)继发的膀胱平滑肌(BSM)肥大、神经原性膀胱和糖尿病相关，其发病率和患病率随着人口的老龄化而增加。现有的治疗干预措施既不是高度有效的，也不是持久的，而且有其自身的副作用。生化研究表明，梗阻膀胱肥厚的BSM线粒体ATP减少，活性氧(ROS)产生增加，这与中间丝(IF)蛋白、结蛋白和波形蛋白的过度表达有关，这些蛋白对肌肉和线粒体功能至关重要。线粒体和肌肉的病理生理学研究表明，结蛋白和波形蛋白的过度表达在诱导线粒体和肌肉功能障碍中起一定作用。然而，这些IF蛋白介导BSM功能障碍的潜在机制仍未被探索。我们的初步数据显示，结蛋白和波形蛋白在人和小鼠BSM条和细胞中的过度表达显著减少线粒体ATP的合成，增加ROS的产生，并抑制氨基甲胆碱和KCl介导的收缩，这在患有PBOO的BSM中也可以看到。IF蛋白在BSM条带和细胞中的过表达增强了丝裂原活化蛋白激酶(MAPK)/JNK2的磷酸化，这与其增加线粒体定位和结蛋白和波形蛋白的磷酸化相一致，在患有PBOO的BSM中也可以看到。线粒体结蛋白和波形蛋白可能通过线粒体JNK2抑制线粒体电压依赖性阴离子通道(VDAC)的通透性。根据我们的初步数据，我们假设JNK2的激活及其线粒体定位的增加与结蛋白和波形蛋白诱导的BSM收缩和线粒体功能障碍有关。我们进一步假设，JNK2磷酸化的线粒体结蛋白和波形蛋白与VDAC相互作用，这种相互作用抑制了ATP的合成，增加了ROS的产生。我们设计了三个具体目标来解决这些假设。在目标1中，我们将建立IF表达增加在导致BSM收缩功能障碍中的稳健性，并阐明JNK2是一个关键的效应因子。在目标2中，我们将确定蛋白过度表达是否导致BSM收缩和线粒体功能障碍是由于JNK2依赖的结蛋白和波形蛋白与VDAC的相互作用。在目标3中，我们将确定线粒体JNK2在PBOO诱导的BSM收缩和线粒体功能障碍中的作用。我们期望我们的研究能够揭示线粒体JNK2信号促进VDAC与结蛋白和波形蛋白相互作用的收缩功能障碍的机制，从而确定治疗PBOO/LUTD的新靶点。

 DESCRIPTION (provided by applicant): Lower urinary tract dysfunction (LUTD) is associated with bladder smooth muscle (BSM) hypertrophy secondary to benign prostatic hyperplasia (BPH)-induced partial bladder outlet obstruction (PBOO), neurogenic bladder, and diabetes, and its incidence and prevalence are increasing as the population ages. Existing therapeutic interventions are neither highly effective nor durable and have side effects of their own. Biochemical studies have shown reduced mitochondrial ATP and increased reactive oxygen species (ROS) production in the hypertrophied BSM of obstructed bladders and this is associated with overexpression of the intermediate filament (IF) proteins, desmin and vimentin which are important for muscle and mitochondrial function. Mitochondrial and muscle pathophysiology, including decreased ATP, increased ROS production and diminished contractility of smooth muscle in LUTD suggests a role for desmin and vimentin overexpression in inducing mitochondrial and muscle dysfunction. However, potential mechanisms by which these IF proteins mediate BSM dysfunction remain unexplored. Our preliminary data demonstrate that desmin and vimentin overexpression in human and murine BSM strips and cells significantly reduces mitochondrial ATP synthesis; increases ROS production and inhibits carbachol-and KCl-mediated contraction as also seen in BSM with PBOO. Overexpression of IF proteins in BSM strips and cells enhances mitogen-activated protein kinase (MAPK)/JNK2 phosphorylation which coincides with its increased mitochondrial localization and phosphorylation of desmin and vimentin as also seen in BSM with PBOO. Mitochondrial desmin and vimentin could potentially function to inhibit the mitochondrial voltage-dependent anion channel (VDAC) permeability via mitochondrial JNK2. Based on our preliminary data, we hypothesize that the activation of JNK2 and its increased mitochondrial localization is involved in desmin- and vimentin-induced BSM contractile and mitochondrial dysfunction. we further hypothesize that JNK2 phosphorylated mitochondrial desmin and vimentin interacts with the VDAC and this interaction inhibits ATP synthesis and increases ROS production. We designed three Specific Aims to address these hypotheses. In Aim 1, we will establish the robustness of increased IF expression in inducing BSM contractile dysfunction, and clarify JNK2 as a critical effector. In Aim 2, we will determine whether IF protein overexpression-induced BSM contractile and mitochondrial dysfunction is due to JNK2 dependent interaction of desmin and vimentin with VDAC. In Aim 3, we will ascertain the role of mitochondrial JNK2 in PBOO- induced BSM contractile and mitochondrial dysfunction. We expect our studies to delineate a mechanism of contractile dysfunction mediated by mitochondrial JNK2 signaling promoting the VDAC interaction with desmin and vimentin, and thus identify novel therapeutic targets for the treatment of PBOO/LUTD.