Defective PS Exposure in Neutrophil Apoptosis in CGD

CGD 中中性粒细胞凋亡中的 PS 暴露缺陷

• 批准号: 6991217
• 负责人: DONNA L BRATTON
• 金额: $ 25.91万
• 依托单位: NATIONAL JEWISH HEALTH
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-12-15 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6991217
• 关键词: NAD(P)H dehydrogenase; apoptosis; chronic granulomatous disease; clinical research; cytokine; fungal antigens; human subject; immune response; inflammation; laboratory mouse; liposomes; mass spectrometry; matrix assisted laser desorption ionization; mycosis; neutralizing antibody; neutrophil; phagocytes; phosphatidylserines; phospholipids; transforming growth factors; transport proteins

DESCRIPTION (provided by applicant): Chronic granulomatous disease (CGD) is a rare, inherited disorder of various defects of the NADPH oxidase rendering it dysfunctional and predisposing to invasive and life-threatening bacterial and fungal infection. While the role of the defective oxidase in immunodeficiency is undisputed, disordered inflammation in the disease (e.g., poor wound healing, obstructing granulomata and autoimmunity), which may or may not be associated with infection, remains unexplained. It is hypothesized that disordered inflammation in CGD results from defective phosphatidylserine (PS) exposure and accumulation on apoptosing CGD neutrophils that leads to impaired recognition and engulfment by phagocytes. Preliminary data suggest that the functioning NADPH oxidase is required for appropriate PS exposure during neutrophil apoptosis, an event that requires coordinate activation of phospholipid (PL) scrambling to expose PS, and inactivation of the aminophospholipid translocase which returns PS to the plasma membrane inner leaflet. This proposal seeks to define the defect in PS exposure on CGD neutrophils by assessing both PL scrambling and aminophospholipid translocase activity. It is hypothesized that apoptosing CGD neutrophils will have diminished PL scrambling and sustained aminophospholipid translocase activity. Exposure of PS on apoptotic cells is required for recognition by phagocytes and is mediated through a receptor, the PSR, that recognizes PS in a stereospeciflc manner. Engagement of the PSR along with other tethering receptors mediates the ingestion of apoptotic neutrophils by an immunologically "silent" mechanism that results in TGFbeta production and suppression of pro-inflammatory cytokine production from the phagocyte. Thus, we hypothesized that PS deficient apoptotic CGD neutrophils will show impaired phagocytic recognition and engulfment, and promote pro-inflammatory cytokine production. Furthermore, unengulfed CGD neutrophils will undergo post-apoptotic cytolysls releasing injurious proteins. Both in vitro phagocytic engulfment assays and in vivo murine models are proposed. It is predicted that apoptotic CGD neutrophils, relative to apoptotic normal neutrophils, instilled into the inflamed peritonea of wild type mice will show delayed clearance, suppression of TGFbeta production and enhanced pro-inflammatory cytokine production; restoration of PS on the surface of CGD neutrophils will reverse this. Finally, injection of CGD mice with sterile fungal hyphae in the presence of PS liposomes will lead to enhanced clearance of accumulating neutrophils, enhanced TGFbeta production and resolution of inflammation. It is expected that data from this investigation will identify novel potential therapeutic targets to address disordered inflammation of CGD.

描述(申请人提供)：慢性肉芽肿性疾病(CGD)是一种罕见的遗传性疾病，具有NADPH氧化酶的各种缺陷，使其功能失调，容易受到侵袭性和危及生命的细菌和真菌感染。虽然缺陷氧化酶在免疫缺陷中的作用是毋庸置疑的，但疾病中的无序炎症(如伤口愈合不良、肉芽肿阻碍和自身免疫)可能与感染有关，也可能不与感染有关，但仍不清楚。据推测，CGD的炎症障碍是由于磷脂酰丝氨酸(PS)的缺陷暴露和聚集在CGD中性粒细胞上，导致吞噬细胞的识别和吞噬功能受损。初步数据表明，在中性粒细胞凋亡过程中，功能正常的NADPH氧化酶是PS适当暴露所必需的，这一事件需要磷脂(PL)的协调激活才能暴露PS，而将PS返回质膜内叶的氨基磷脂转位酶则需要失活。这项建议试图通过评估PL扰乱和氨基磷脂转位酶活性来确定CGD中性粒细胞PS暴露的缺陷。这是假设，通过使用CGD中性粒细胞可以减少PL的扰乱和维持氨基磷脂转位酶的活性。PS暴露在凋亡细胞上是吞噬细胞识别所必需的，并通过一种以立体特异性方式识别PS的受体PSR介导。PSR与其他拴系受体的结合通过一种免疫“沉默”机制介导了对凋亡中性粒细胞的摄取，从而导致TGFβ的产生和吞噬细胞产生的促炎细胞因子的抑制。因此，我们假设PS缺乏的凋亡的CGD中性粒细胞将表现出吞噬识别和吞噬功能受损，并促进促炎细胞因子的产生。此外，未被包埋的CGD中性粒细胞将经历凋亡后的细胞溶解，释放损伤蛋白。提出了体外吞噬试验和体内小鼠模型。据预测，与正常中性粒细胞相比，野生型小鼠炎症腹膜中注入的凋亡的CGD中性粒细胞将表现出延迟清除、抑制TGFβ的产生和促进促炎细胞因子的产生；恢复CGD中性粒细胞表面的PS将逆转这一趋势。最后，在PS脂质体存在的情况下，给CGD小鼠注射无菌真菌菌丝将导致积累的中性粒细胞被增强清除，增加TGFβ的产生和炎症的消退。预计这项研究的数据将确定新的潜在治疗靶点，以解决CGD的紊乱炎症。