Signal transduction defects in human platelets

血小板起主要止血作用，先天血小板功能缺陷均与出血表现相关。本项目的目标是明确先天性血小板功能缺陷患者导致血小板功能障碍的生化机制，从而加强我们对正常血小板激活机制的了解，这对于开发新的出血和血栓形成障碍的治疗策略至关重要。绝大多数先天性血小板缺陷症患者一般以聚集反应受损和激活时分泌致密颗粒为特征；大多数患者具有正常的致密颗粒储存。这些患者被归入被称为“血小板分泌缺陷”或“激活缺陷”的宽泛定义的组。在它们中，潜在的生化和分子机制是完全未知的。我们的假设是，这些患者在信号转导机制上存在缺陷。这个项目聚焦于特定的患者，我们在这些患者中发现了两种与信号转导机制有关的主要蛋白质的缺陷，即a)磷脂酶C-β2(Aim 1)和b)GTP结合蛋白Galphaq(Aim 2)。到目前为止，我们的研究表明，这些患者的PLC-β2和Galphaq基因的编码序列正常，但各自的mRNA水平降低。我们将研究PLC-beta2(Aim 1)和Galphaq(Aim 2)的转录调控，以确定其机制。在其他几个有血小板功能缺陷的患者中，我们已经获得了信号事件异常的证据(例如，钙动员、Pleckstrin磷酸化)。在目标3中，我们将在3-4名患者及其家属中进行详细的研究，以确定导致反应受损的机制。这个项目代表了应用最先进的技术来确定一组患者的血小板功能障碍的分子机制，这些患者目前的特征非常差，构成了一个尚未开发的新信息来源。这些研究将为两种主要蛋白质PLC-β2和Galphaq在血小板信号转导机制中的作用提供新的信息。

Platelets play a major role in hemostasis in congenital defects in platelet function are associated with bleeding manifestations. The goal of this project is to define the biochemical mechanisms underlying the platelet dysfunction in patients with congenital platelet function defects and thereby enhance our understanding of the normal platelet activation mechanisms, which are vital to the development of newer therapeutic strategies for both bleeding and thrombotic disorders. The vast majority of patients with congenital platelet defects are generally characterized by impaired aggregation responses and dense granule secretion on activation; most of them have normal dense granule stores. These patients are lumped in the loosely defined group called "platelet secretion defects" or "activation defects". In them, the underlying biochemical and molecular mechanisms are totally unknown. Our hypothesis is that these patients have defects in the signal transduction mechanisms This project focuses on specific patients in whom we demonstrate hitherto undescribed deficiencies in two major proteins involved in signal transduction mechanisms, namely, a) phospholipase C-beta2 (Aim 1), and b) GTP- binding protein Galphaq (Aim 2), respectively. Our studies to date demonstrate that the coding sequence of PLC-beta2 and Galphaq gene is normal in these patients but the respective mRNA levels are decreased. We will study the transcriptional regulation of PLC-beta2 (Aim 1) and Galphaq (Aim 2) to define the mechanisms. In several other patient with platelet function defects, we have obtained evidence for abnormalities in the signaling events (e.g. Ca2+ mobilization, pleckstrin phosphorylation). In Aim 3, we will perform detailed studies in 3-4 patients and their family members to characterize the mechanisms leading to the impaired responses. This project represents application of state-of-the-art techniques to define the molecular mechanisms of platelet dysfunction in a group of patients who are very poorly characterized at present and constituted an untapped source of new information. These studies will provide new information on the role of two major protein, PLC-beta2 and Galphaq, in platelet signal transduction mechanisms.