Investigation of molecular mechanisms underlying the role of ICAM-3 in the phagocytic clearance of apoptotic leukocytes

体内受损、感染、老化或不需要的细胞通过称为凋亡的过程被诱导死亡。这个过程的最后一步是通过健康的邻近细胞去除（吞噬）垂死细胞，在那里细胞尸体在安全和受控的环境中被吃掉和降解。一系列的分子已经被认为在这一过程中发挥作用，尽管与垂死细胞上识别的分子相关的信息量是有限的。有人认为，死亡细胞表面的变化导致将细胞识别为死亡并准备移除的信号或标志暴露于有活力的邻居。目前对这种“吃我”信号的性质知之甚少。已知细胞表面的脂质在死亡过程中发生变化，磷脂酰丝氨酸（PS）暴露在细胞表面，而在活细胞中，它通常保持在细胞膜的内小叶内。一旦暴露，PS被活细胞上的受体识别并介导死亡细胞清除。另一种分子ICAM-3（细胞间粘附分子-3）在垂死白细胞表面上的变化与专职吞噬细胞巨噬细胞识别和清除垂死白细胞有关。ICAM-3是仅在白细胞上发现的分子，并且当存在于活细胞上时，通过促进细胞-细胞相互作用在免疫应答的起始中是重要的。为了介导这种相互作用，ICAM-3与包括整联蛋白LFA-1和非整联蛋白DC-SIGN的反受体结合。先前的工作表明，垂死细胞上的ICAM-3失去了与其原型反受体LFA-1结合的能力。这种凋亡相关的功能丧失与替代功能的获得相关，其中ICAM-3能够结合另一种尚未鉴定的受体-一种在介导凋亡白细胞的吞噬清除中重要的功能。去除垂死的白细胞对于炎症的解决和免疫反应的控制特别重要。未能在适当的时间清除白细胞可能会导致慢性炎症和自身免疫性疾病。更好地了解死亡白细胞清除过程中涉及的分子和过程将是未来改善人类健康和福祉的战略的核心。该项目旨在确定ICAM-3中发生的变化，这些变化是细胞凋亡期间报告的功能丧失（LFA-1结合丧失）和功能获得（结合其他受体和介导死亡白细胞清除的能力）的基础。在凋亡细胞表面与ICAM-3相关的功能的变化可能是由于ICAM-3位置、配偶体分子或结构（例如ICAM-3的糖含量）的变化，并且任何这样的变化可能是由于细胞经历凋亡时ICAM-3的迁移率的变化引起的或有助于ICAM-3的迁移率的变化。在这个项目中，我们将识别和检测细胞凋亡过程中ICAM-3的结构和/或环境的变化，并将这些变化与死细胞清除联系起来。

Damaged, infected, aged or unwanted cells in the body are induced to die via a process known as apoptosis. The final step in this process is the removal (phagocytosis) of dying cells by healthy neighbouring cells where the cell corpses are eaten and degraded in a safe and controlled environment. A range of molecules have been suggested to play a role in this process though the amount of information relating to molecules recognised on dying cells is limited. It has been suggested that changes at the surface of dying cells result in the exposure of signals or flags that identify the cell, to viable neighbours, as dying and ready for removal. Relatively little is currently known about the nature of such 'eat me' signals. Lipids at the cell surface are known to change during death with phosphatidylserine (PS) becoming exposed at the cell surface where in viable cells it is usually maintained within the inner leaflet of the cell membrane. Once exposed PS is recognised by receptors on viable cells and mediates dying cell clearance. A change in another molecule, ICAM-3 (Intercellular Adhesion Molecule-3) on the surface of dying leukocytes have been implicated in the recognition and removal of dying leukocytes by professional phagocytes, macrophages. ICAM-3 is a molecule found only on leukocytes and, when present on viable cells, is important in the initiation of immune responses by facilitating cell-cell interactions. To mediate this interaction ICAM-3 binds to counter-receptors including the integrin LFA-1 and the non-integrin DC-SIGN. Previous work has indicated that ICAM-3 on dying cells loses its ability to bind to its prototypic counter-receptor LFA-1. This apoptosis-related loss of function is associated with a gain of alternate function where ICAM-3 becomes able to bind to another, as yet unidentified, receptor - a function that is important in mediating the phagocytic clearance of apoptotic leukocytes. The removal of dying leukocytes is especially important for the resolution of inflammation and the control of immune responses. Failure to remove leukocytes at the appropriate time may lead to chronic inflammatory conditions and autoimmune disease. A better understanding of the molecules and processes involved within the clearance of dying leukocytes will be central to future strategies for improving human health and well-being. This project seeks to identify the changes that occur in ICAM-3 that underlie the loss of function (loss of LFA-1 binding) and the gain of function (ability to bind other receptor(s) and mediate clearance of dying leukocytes) that is reported during apoptosis. The change of function associated with ICAM-3 at the apoptotic cell surface may be due to changes in ICAM-3 location, partner molecules or structure (e.g. sugar content of ICAM-3) and any such changes may arise from or contribute to changes in the mobility of ICAM-3 as cells undergo apoptosis. Within this project we will identify and characterise changes in the structure and/or environment of ICAM-3 occurring during apoptosis and will relate these changes to dead cell clearance.