MECHANISMS FOR ALTERED DETRUSOR CONTRACTION IN DIABETES

描述（改编自应用程序） 人类糖尿病与一系列排尿功能障碍有关 其特征在于膀胱感觉受损， 逼尿肌收缩力下降，逼尿肌可能进展 反射消失和尿流减少。动物模型研究， 特别是链脲佐菌素（STZ）诱导的糖尿病大鼠， 患有遗传性尿崩症（di/di）的布拉特博罗（BB）大鼠显示， 类似于人类糖尿病的变化，例如，大膀胱 排尿功能受损的能力。拟议研究的数据 将提供有关糖尿病对健康影响的基本信息， 逼尿肌收缩力和膀胱功能，并测试潜在的 假设糖尿病对分子机制有影响， 调节清空所需的力的产生和维护 膀胱为了解决这一假设，我们建议使用逼尿肌 来自糖尿病（STZ诱导的糖尿病BB大鼠）的平滑肌，胰岛素 治疗组和对照组大鼠（非糖尿病渗透性利尿和正常）， 以确定调节机制如何控制（通过蛋白质 蛋白质相互作用或酶促作用）被这些蛋白质改变。 具体来说我们将确定：（1）是否缺乏膀胱灌注 糖尿病膀胱的压力和过度膨胀是由于低水平的 肌球蛋白轻链（MLC）磷酸化或肌球蛋白轻链（MLC）磷酸化位点的变化 静息时逼尿肌平滑肌中MLC的磷酸化 （2）活动或活动过程中， MLC激酶和/或蛋白C激酶（PCK）同种型的表达， 参与信号通路，在糖尿病逼尿肌中改变; (3)平滑肌特异性钙调蛋白表达的变化 （h-钙调素），细丝组成部分，调节肌动蛋白-肌球蛋白 糖尿病患者逼尿肌中的相互作用导致 糖尿病平滑肌无力维持力量;（4）是否 这是平滑肌肌球蛋白亚型表达的变化， 对糖尿病的反应;（5）肌动蛋白/激活的ATP酶活性是否 糖尿病患者膀胱逼尿肌肌球蛋白的含量发生改变。我们建议 使用来自糖尿病患者、非糖尿病渗透性 利尿剂和正常大鼠。肌球蛋白的表达将使用 逆转录聚合酶链反应（PCR），定量 比较PCR和北方和西方印迹分析。的功能 通过测量肌球蛋白ATP酶活性，分析肌球蛋白亚型 活动和运动的肌动蛋白丝的肌球蛋白头中， 体外运动性测定。逼尿肌收缩力将通过力分析 测量完整的肌肉条。收缩力的改变 收缩的装置和调节，独立于膜， 肌浆网，将使用化学"皮肤" 肌肉条对导致 糖尿病的收缩功能障碍是至关重要的， 药物治疗的分子步骤 糖尿病性膀胱病

DESCRIPTION (adapted from application) Diabetes in humans is associated with a spectrum of voiding dysfunctions characterized by impaired bladder sensation, increased postvoidal residual volume and decreased detrusor contractility that may progress in detrusor areflexia and diminished urinary flow. Studies on animal models, particularly the streptotzotozin (STZ) induced diabetic rats and Brattleboro (BB) rats with hereditary diabetic insipidus (di/di), show changes similar to those in human diabetes, e.g., a greater bladder capacity with impaired voiding function. Data from the proposed studies will provide the basic information regard the effect of diabetes on detrusor contractility and bladder function, and test the underlying hypothesis that diabetes has an effect on the molecular mechanisms that regulate force generation and maintenance that are required to empty the urinary bladder. To address this hypothesis, we propose to use detrusor smooth muscle from diabetic (STZ induced diabetes BB rats), insulin treated, and control rats (non diabetic osmotic diuresis, and normal) and to determine how the regulatory mechanisms controlled (through protein protein interaction or enzymatically) by these proteins are altered. Specifically. we will determine: (1) whether the lack of intravesical pressure and over distension of the diabetic bladder is due to a low level of myosin light chain (MLC) phosphorylation or a change in the site of phosphorylation of the MLC in the detrusor smooth muscle at the resting tone or during force development; (2) whether the activities and expression of MLC kinases and/or the protein C kinase (PCK) isoforms, implicated in signaling pathways, are altered in the detrusor in diabetes; (3) whether changes in the expression of smooth muscle specific caldesmon (h-caldesmon), the thin filament component that regulates actin-myosin interactions, in the detrusor in diabetes is responsible for the inability for the inability of diabetic smooth muscle to maintain force; (4) whether thre is a change in the expression of smooth muscle myosin isoforms in response to diabetes; and (5) whether the actin/activated ATPase activity of myosin in the detrusor is altered in diabetic bladders. We proposed to use muscles from bladders and urethras of diabetic, non diabetic osmotic diuretic and normal rats. The expression of myosin will be studied using reverse transcribed polymerase chain reaction (PCR), quantitative comparative PCR, and Northern and Western blot analyses. The function of myosin isoforms will be analyzed by measuring the action-myosin ATPase activity and the movement of actin filaments over myosin heads in the in vitro motility assays. Detrusor contractility will be analyzed by force measurements of intact muscle strips. Alterations in the contractile apparatus and regulation of contraction, independent of the membranes and sarcoplasmic reticulum, will be determined using chemically "skinned" muscle strips. An understanding of the molecular events that lead to contractile dysfunctions in diabetes is crucial in order to target the molecular steps for the development for pharmacological gents to treat diabetic cystopathy.