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

项目总结 外周动脉疾病(PAD)影响着全球2亿多成年人。严重肢体缺血(CLI)， 最先进的PAD形式，导致显著的发病率、死亡率和卫生保健资源的利用。 尽管我们对PAD的病理生物学有了更多的了解，但药物治疗仍然不足和 不幸的是，血运重建(手术和非手术)或截肢是主要的治疗选择。 因此，迫切需要解决这一重要的未得到满足的临床需求，可能是通过开发 新的药物疗法与更有效的管理策略相结合。但是，当前 由于对基本机制的不完全了解，这些领域的努力受到严重阻碍 控制着血管功能的失调，以及无法产生有效的血管 恢复流量的网络。我们最近的观察确定了CCN3(NOV)，一个特定的成员 基质细胞蛋白家族CCN(Cyr61，CTGF，NOV)作为血管内皮细胞功能的重要调节因子 新血管形成的背景。在CLI的肢体组织中发现CCN3的表达显著减少 病人。在小鼠后肢缺血(HLI)模型中，整体CCN3缺乏导致更严重的坏死 伴随着组织再灌注、缺氧诱导因子(HIF)信号和血管内皮生长因子-A的减少 产生--PAD中功能性侧支血流丧失的关键机制。另外， 小鼠CCN3的细胞类型特异性缺失和初步细胞研究表明，内皮细胞 CCN3缺乏在血流恢复障碍中起主要作用。CCN3的限制性缺失 内皮细胞具有全局CCN3基因敲除的作用。这强烈地表明内皮细胞 CCN3是HLI后侧支循环血流恢复的正向调节因子。在细胞水平上，CCN3的丢失 导致内皮细胞迁移和管子形成受损，这是参与血管生成的关键过程 是恢复血流所必需的。总而言之，这些观察结果导致了CCN3的中心假设 在驱动新生血管和伴随的组织灌流中起着关键的生理调节作用。在……里面 目的1、研究CCN3在侧支循环血流调节中的作用。AIM 2是专门设计的 目的：阐明CCN3缺乏导致新生血管受损的机制。 侧支循环血流。最后，在目标3中，我们计划探索CCN3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CCN3在控制 缺血时血管内皮细胞的功能及其促进新生血管和血管生成的机制 功能性侧支循环血流。

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.