Mechanisms of Inflammation: Role of CD34 and Podocalyxin in Neutrophil Recruitment

炎症是宿主对生物、化学或物理侮辱的先天防御反应。炎症是通过感染细菌、病毒和寄生虫引起的。炎症是许多器官疾病的基础，如肝脏(如肝炎)、关节(如关节炎)和肺部(如肺炎和哮喘)。炎症的特征是血液中中性粒细胞和单核细胞等防御细胞的激活。中性粒细胞等被激活的细胞从血液进入器官以对抗细菌。中性粒细胞通过与血管内皮细胞的相互作用从血管中迁移。内皮细胞排列在血管壁的内侧。激活的中性粒细胞通过产生氧自由基等化学物质杀死细菌。激活的中性粒细胞过度迁移到组织中，会导致组织受到氧自由基和中性粒细胞产生的其他化学物质的严重损害。中性粒细胞产物造成的组织损伤，如果过量，会导致发病率和死亡率。为了保持平衡，确保中性粒细胞利大于弊，我们需要保持中性粒细胞的抗微生物作用，同时微调它们在组织中的迁移，以防止组织损伤。这可能是通过控制中性粒细胞的迁移来实现的。 中性粒细胞的迁移通过各种步骤发生，如减缓，在内皮上滚动，与内皮细胞牢固粘连，最终迁移出血管。中性粒细胞迁移到组织中的早期事件之一是中性粒细胞与内皮细胞之间的相互作用。这种相互作用是通过重组细胞骨架蛋白(如F-肌动蛋白)在中性粒细胞膜上形成突起(称为伪足)来促进的。导致中性粒细胞细胞骨架改变的机制尚不完全清楚。我建议研究CD34和Podocalysin(PC)蛋白在这些过程中的作用，以最终发现控制中性粒细胞迁移的分子机制。这些研究包括对影响中性粒细胞和内皮细胞CD34和PC表达的机制的评估，以及这些蛋白在中性粒细胞与内皮细胞相互作用中的作用的研究。 这些研究将通过一系列体外和体内方法来研究细胞的形态、分子和结果：共聚焦和电子免疫细胞化学；免疫印迹；流式细胞术；内皮细胞和中性粒细胞培养；小鼠提睾肌和肺的炎症模型。总而言之，我和我的受训者将进行从分离培养的中性粒细胞和内皮细胞到两种不同的炎症动物模型的研究，以了解中性粒细胞迁移的基本性质。 这些研究的数据有望显示CD34和PC在中性粒细胞伪足形成过程中的未知角色，中性粒细胞伪足的形成促进了与血管内壁内皮细胞的相互作用。由于这些最初的膜接触对中性粒细胞的最终迁移至关重要，这些数据将促进我们的知识，并有助于开发动物和人类炎症性疾病的新分子管理模型。

Inflammation is a host’s innate defense response to biological, chemical or physical insults. Inflammation is induced through infections with bacteria, viruses and parasites. Inflammation underlies many diseases in organs such as the liver (e.g., hepatitis), joints (e.g., arthritis) and lungs (e.g., pneumonia and asthma). Inflammation is characterized by the activation of defense cells such as neutrophils and monocytes while in the blood stream. Activated cells such as neutrophils move from the blood into the organs to fight bacteria. Neutrophils migrate from the blood vessels through interactions with the endothelial cells. Endothelial cells line the inside of the walls of blood vessels. Activated neutrophils kill bacteria through production of chemicals such as free oxygen radicals. Excessive migration of activated neutrophils into the tissues results in significant tissue damage from free oxygen radicals and other chemicals produced by the neutrophils. The tissue damage caused by products of neutrophils, if excessive, contributes to morbidity and mortality. In order to maintain balance and insure that neutrophils do more good than harm, we need to maintain the anti-microbial actions of the neutrophils, and at the same time, fine-tune their migration into the tissues to prevent tissue damage. This can be potentially accomplished through control of migration of the neutrophils. Neutrophil migration occurs through various steps such as slowing, rolling on the endothelium, firm adhesion with the endothelium and finally migration out of the blood vessels. One of the very early events in the migration of neutrophils into the tissues is the interaction between the neutrophils and the endothelium. This interaction is facilitated through the formation of projections (called pseudopods) on the neutrophil membrane through reorganization of cell skeleton proteins such as F-actin. The mechanisms that lead to the cytoskeletal changes in neutrophils are not fully understood. I propose to study the roles of CD34 and podocalyxin (PC) proteins in these processes to eventually discover molecular mechanisms to control neutrophil migration. The studies include evaluation of the mechanisms that influence the expression of CD34 and PC in neutrophils and endothelial cells as well as the study of the role of these proteins in the interactions of neutrophils with the endothelium. The studies will be conducted with an array of in vitro and in vivo methods to investigate the cell morphology, molecules and outcomes: confocal and electron immunocytochemistry; western blots; flow cytometry; endothelial and neutrophil cultures; mouse models of inflammation in cremaster muscle and lungs. Taken together, my trainees and I will conduct studies that range from isolated cultured neutrophil and endothelial cells to two different animal models of inflammation to understand the fundamental nature of the neutrophil migration. The data from these studies are expected to show hitherto unknown roles for CD34 and PC in the formation of pseudopods on neutrophils, which facilitate interactions with endothelial cells lining the inside walls of the blood vessels. Because these initial membrane contacts are critical for the eventual migration of neutrophils, these data will advance our knowledge and contribute towards the development of new molecular management models of inflammatory diseases for animals and humans.