MOLECULAR MECHANISM FOR ERECTILE FUNCTION & DYSFUNCTION

勃起功能的分子机制

• 批准号: 2822707
• 负责人: Michael Edward DiSanto
• 金额: $ 15.95万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-29 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2822707
• 关键词: RNA splicing; RNase protection assay; caldesmon; clinical research; enzyme activity; gel electrophoresis; human subject; human tissue; immunocytochemistry; isozymes; laboratory rabbit; male; muscle cells; muscle contraction; muscle relaxation; myosin light chain kinase; myosins; penis disorder; penis erection; protein isoforms; smooth muscle; southern blotting

Erection(tumescence) is a complex neurophysiologic event that leads to relaxation of the smooth muscle cells in the corpus cavernosum penis which allows filling of blood into the cavernous spaces causing an increase in the size and rigidity of the penis. Flaccidity (detumescence) is initiated and maintained by contraction of the corpus cavernosum smooth muscle (CCSM) cells. The CCSM is unique since it remains in the contracted state most of the time. It is not know whether the Ca2+-calmodulin-MLCK pathway or other then filament- mediated regulation (via the actin-binding protein caldesmon) plays a role in keeping these cells in the contracted state, for the penis to remain flaccid. A third possibility is that the myosin isoforms in CCSM play a role in determining the velocity of force generation and the tonicity. A recent study conducted by this investigator showed that the aortic smooth muscle, which is considered tonic, contains one type of myosin isoform whereas the smooth muscle cells in small muscular arteries contain a different myosin isoform. The latter isoform is encoded by a myosin heavy chain mRNA that is alternatively spliced to insert a 21-nucleotide sequence that encodes a region near the ATP-binding site on the myosin head. This insert encodes a 7-amino acid peptide, and the resulting myosin isoform has a two-fold increase in the actin-activated myosin ATPase activity and an associated increase in shortening velocity (Vmax). Similar studies conducted on CCSM in our lab show that the CCSM contains both inserted (as in small muscular arteries and all visceral smooth muscles which show phasic characteristics) and non-inserted (as in aortic smooth muscle which is tonic) myosin heavy chain isoforms. Specially, using CCSM from normal rabbits, normal humans, and patients with erectile dysfunctions, we will address the following questions: (1) Are the differences in the contractile characteristics of CCSM compared to smooth muscle from other sources due to a difference in the composition of smooth muscle myosin isoforms that are produced by alternative splicing of the myosin pre-mRNA at the 3' end (resulting in SM1 and SM2) or the 5' end (resulting in inserted myosin)? (2) What role does myosin light chain phosphorylation play in erectile function and do CCSM cells have higher kinase or lower phosphatase activity compared to other smooth muscle cells (e.g. bladder smooth muscle)? (3) What is the level of expression of myosin light chain kinase and caldesmon in the CCSM and is it altered in patients with erectile dysfunction? Thus, the results from experiments proposed in this application will provide the molecular mechanisms for the regulation of CCSM contraction and delineate specific steps in the pathways for cross-bridge cycling and contractility of smooth muscle cells in the normal corporus cavernosum penis. It will also provide and explanation for changes in contractility of smooth muscle cells seen in patients with erectile dysfunction.

勃起（肿胀）是一种复杂的神经生理事件，导致 松弛海绵体内的平滑肌细胞 阴茎允许血液充满海绵体空间，导致 阴茎的尺寸和硬度增加。 弛缓性 （消肿）是通过收缩来启动和维持的 海绵体平滑肌（CCSM）细胞。 CCSM 是独一无二的 因为它大部分时间都处于契约状态。 它不是 知道 Ca2+-钙调蛋白-MLCK 途径或其他细丝- 介导的调节（通过肌动蛋白结合蛋白 caldesmon）起着 使这些细胞保持收缩状态的作用，使阴茎 保持松弛状态。 第三种可能性是肌球蛋白亚型 CCSM 在确定力产生的速度和 张力。 该研究者最近进行的一项研究表明，主动脉 平滑肌被认为是补品，含有一种肌球蛋白 同种型，而小肌动脉中的平滑肌细胞 含有不同的肌球蛋白亚型。 后一种异构体编码为 肌球蛋白重链 mRNA，选择性剪接以插入 编码 ATP 结合附近区域的 21 个核苷酸序列 位于肌球蛋白头部。 该插入片段编码 7 个氨基酸肽， 由此产生的肌球蛋白同种型的 肌动蛋白激活的肌球蛋白 ATP 酶活性和相关的增加 缩短速度（Vmax）。 我们对 CCSM 进行了类似的研究 实验室表明 CCSM 包含两种插入（如小肌肉 动脉和所有内脏平滑肌表现出阶段性 特征）和非插入（如主动脉平滑肌， 是补品）肌球蛋白重链亚型。 特别是，使用来自正常兔子、正常人类和 对于患有勃起功能障碍的患者，我们将解决以下问题 问题：（1）收缩特性是否有差异？ CCSM 与其他来源的平滑肌相比，由于 平滑肌肌球蛋白亚型的组成差异 由肌球蛋白前体 mRNA 在 3' 处的选择性剪接产生 端（导致 SM1 和 SM2）或 5' 端（导致插入 肌球蛋白）？ （二）肌球蛋白轻链磷酸化起什么作用 CCSM 细胞是否具有较高的激酶或较低的激酶 与其他平滑肌细胞（例如平滑肌细胞）相比，磷酸酶活性 膀胱平滑肌）？ (3) 表达水平如何 CCSM 中的肌球蛋白轻链激酶和 caldesmon 是否发生改变 勃起功能障碍患者？ 因此，结果为 本申请中提出的实验将提供分子 CCSM 收缩的调节机制和描绘 过桥骑行路线的具体步骤和 正常体中平滑肌细胞的收缩性 阴茎海绵体。 它还将提供更改的解释 患者平滑肌细胞的收缩性 勃起功能障碍。