Release of find-me signals during apoptotic cell clearance

描述(申请人提供)：在几乎所有的组织中，细胞都有持续的更替，通常是通过细胞凋亡的过程。在健康的组织中，死亡的细胞很快就会被吞噬细胞识别和清除。然而，未能及时清除凋亡细胞会导致继发性坏死，释放有毒的细胞质内容物，并导致组织内的炎症。目前的证据表明，在死亡过程的早期，凋亡细胞通过被称为“Find-Me信号”的可溶性因子来吸引吞噬细胞，从而促进它们的迅速清除。这一提议的主要研究人员的初步研究发现，核苷酸ATP和UTP是一种‘Find-Me信号’，对体外和体内的细胞凋亡清除至关重要；随后的研究发现，膜蛋白PAnnexin 1(Panx1)是介导凋亡细胞核苷酸释放的通道。这一建议检验的总体假设是，依赖于pAnnexin通道的核苷酸Find-Me信号从凋亡细胞中释放，随后吞噬细胞的感知对于体内适当的细胞清除是重要的，而Panx1介导的Find-Me信号通路的中断将导致疾病。在我们的初步研究的基础上，在目标1中，我们研究了一种新的Panx1激活机制的分子基础，以寻求新的理解，从而允许通过这些渠道操纵Find-Me信号的释放。在目标2中，我们使用条件性细胞特异性Panx1基因敲除小鼠来确定Panx1通道的操作是否会影响体内的细胞清除。为此，我们利用了两个模型，新的证据表明，细胞清除在正常组织稳态和疾病中都很重要--即胸腺发育和呼吸道炎症。总而言之，我们希望这些研究能够对Find-Me信号的调节释放如何影响细胞清除产生新的机械性理解，并更好地定义死亡细胞和吞噬细胞之间的这种细胞间通信模式，从而对自身免疫和呼吸道炎症产生影响。这些研究还可以为将Panx1通道作为治疗开发的合适靶点提供理论依据。

DESCRIPTION (provided by applicant): In nearly all tissues, there is a continual turnover of cells, usually by the process of apoptosis. In healthy tissues, the dying cells are quickly recognized and cleared by phagocytes. However, failure to promptly clear apoptotic cells leads to their secondary necrosis, release of toxic cytoplasmic contents, and inflammation within tissues. Current evidence suggests that the apoptotic cells 'advertise' their presence early on in the death process, via soluble factors termed 'find-me signals' to attract phagocytes, and thereby promote their prompt clearance. Initial studies from the Principal Investigators of this proposal have identified the nucleotides ATP and UTP as one type of 'find-me signal' that is critical for apoptotic cell clearance in vitro and in vivo; subsequent studies led to a key discovey that the membrane protein pannexin 1 (Panx1) is the channel mediating nucleotide release from apoptotic cells. The overall hypothesis tested in this proposal is that pannexin channel-dependent release of nucleotide find-me signal from apoptotic cells, and subsequent sensing by phagocytes is important for proper cell clearance in vivo, and that disruption of the Panx1-mediated find-me signal pathway would contribute to diseases. Based on our preliminary studies, in Aim 1, we study the molecular basis of a novel Panx1 activation mechanism, seeking new understanding that will allow manipulation of 'find-me' signal release via these channels. In Aim 2, we use conditional cell-specific Panx1 knockout mice to determine if manipulation of Panx1 channels affects cell clearance in vivo. For this, we take advantage of two models for which new evidence suggests cell clearance is important in normal tissue homeostasis and in disease - i.e., thymic development and airway inflammation. Collectively, we expect these studies to yield new mechanistic understanding on how the regulated release of find-me signals influence cell clearance, and better define this mode of inter-cellular communication between dying cells and phagocytes, with implications for autoimmunity and airway inflammation. These studies can also provide a rationale for considering Panx1 channels as a suitable target for therapeutic development.