Deciphering the regulatory role of matricelluar protein CCN3 in functional collateral blood flow

项目摘要 外周动脉疾病（PAD）影响全球超过2亿成年人。严重肢体缺血（CLI）， PAD的最高级形式，导致显著的发病率、死亡率和卫生保健资源利用。 尽管我们对PAD的病理生物学有了更多的了解，但医学治疗仍然不足， 不幸的是，血管再生（手术和非手术）或截肢是主要的治疗选择。 因此，必须解决这一重要的未满足的临床需求，可能通过开发 新的药物治疗结合更有效的管理策略。但目前的 由于对基本机制的了解不全面，这些领域的工作受到严重阻碍 控制血管功能失调，以及未能产生有效的血管 网络恢复流量。我们最近的观察已经确定了CCN 3（Nov），这是一个特定的成员。 基质细胞蛋白家族CCN（Cyr 61，Ctgf，Nov）是内皮细胞功能的重要调节因子， 新血管形成的背景。发现CCN 3表达在来自CLI的肢体组织中强烈降低。 患者在小鼠后肢缺血（HLI）模型中，CCN 3缺乏导致坏死增强， 伴随着组织再灌注、缺氧诱导因子（HIF）信号传导和VEGF-A的减少， 产生-PAD中功能性侧支血流丧失的关键机制。此外，本发明还 小鼠中CCN 3的细胞类型特异性缺失和基于细胞的初步研究表明， CCN 3缺乏在血流恢复受损中起主要作用。CCN 3的限制性缺失 内皮细胞表型模仿CCN 3基因敲除的效应。这有力地表明， CCN 3是HLI后侧支血流恢复的正调节剂。在细胞水平上，CCN 3的损失 导致内皮迁移和管形成受损，这是血管生成中的关键过程 恢复血流的必要条件。总的来说，这些观察结果导致了一个中心假设，即CCN 3 在驱动新血管形成和伴随的组织灌注中充当关键的生理调节剂。在 目的1，我们将充分表征CCN 3在侧支血流调节中的作用。Aim 2设计 阐明CCN 3缺乏导致受损的新血管形成的机制， 侧支血流。最后，在目标3中，我们计划探索CCN 3在临床前的治疗潜力。 肢体缺血模型。这些研究的结果将阐明CCN 3在控制 缺血中的内皮功能及其促进新血管形成和 功能性侧支血流

PROJECT SUMMARY Peripheral artery disease (PAD) affects more than 200 million adults worldwide. Critical limb ischemia (CLI), the most advanced form of PAD, causes significant morbidity, mortality, and health care resource utilization. Despite our increased understanding of the pathobiology of PAD, medical treatments remain inadequate and revascularization (surgical and non-surgical) or amputation are unfortunately the major therapeutic options. Therefore, it is imperative to address this important unmet clinical need, potentially by the development of novel pharmacologic therapies combined with more effective management strategies. However, current efforts in these areas are significantly hindered due to incomplete knowledge of the fundamental mechanisms that govern the dysregulation of vascular function, as well as the failure to generate an effective vascular network to restore flow. Our recent observations have identified CCN3 (Nov), a specific member of the matricellular protein family, CCN (Cyr61, Ctgf, Nov), as an important regulator of endothelial function in the context of neovascularization. CCN3 expression was found to be strongly reduced in limb tissues from CLI patients. In a murine hind limb ischemia (HLI) model, global CCN3 deficiency resulted in enhanced necrosis concomitant with decreases in tissue reperfusion, hypoxia-induced factor (HIF) signaling and VEGF-A production - key mechanisms responsible for the loss of functional collateral blood flow in PAD. Additionally, cell-type specific deletion of CCN3 in mice and preliminary cell-based studies indicate that endothelial cell CCN3 deficiency plays a major role in the impairment of blood flow recovery. Restrictive deletion of CCN3 in the endothelium phenocopies the effects of the global CCN3 knockout. This strongly suggests that endothelial CCN3 is a positive regulator of collateral blood flow recovery following HLI. At the cellular level, loss of CCN3 results in impaired endothelial migration and tube formation, pivotal processes involved in angiogenesis requisite for blood flow recovery. Collectively, these observations led to the central hypothesis that CCN3 serves as a critical physiological regulator in driving neovascularization and the attendant tissue perfusion. In Aim 1, we will fully characterize the role of CCN3 in the regulation of collateral blood flow. Aim 2 is designed to elucidate the mechanisms by which CCN3 deficiency leads to compromised neovascularization and collateral blood flow. Finally, in Aim 3, we plan to explore the therapeutic potential of CCN3 in preclinical models of limb ischemia. The results of these studies will elucidate the role of CCN3 in controlling endothelial function in ischemia and the mechanisms underlying its ability to promote neovascularization and functional collateral blood flow.