PMN MOTILITY AND ADHERENCE IN NEONATES

人类新生儿和遗传性Mac-1，LFA1缺乏症患者 由以下原因引起的感染性并发症的高风险 吞噬细胞的组织动员受损。这些研究将 评估可能导致中性粒细胞受损或 这些人群中的单核白细胞动员。主要侧重点 将包括以下方面的确定：a)功能关系 细胞黏附和迁移特性，b)调节 细胞黏附的诱导、调节和功能激活 炎症刺激，以及c)细胞骨架可能的病理影响 蛋白质(微管)对细胞移位的影响。他们将专注于 粘附性糖蛋白(GP)家族的贡献(Mac-1，LFA1， P150，95)对白细胞的黏附依赖特性；比较 利用新生儿和Mac-1缺陷细胞的研究将允许一种独特的 了解异常的功能和临床后果的机会 表情。趋化因子介导的Mac-1糖蛋白的表面表达 分泌刺激将使用免疫荧光流进行定量 应用单抗的细胞计数和125I免疫沉淀技术 抗体(MAb)；它们与其他表面的地形关系 IIF将对爬行动物(FCR和CR1)和“黏附部位”进行评估 显微镜。每个MAC-1子单元的MAB将在阻塞中使用 评估它们对粘附性依赖的功能贡献的实验 细胞功能包括微管和微丝组装(IIF 微管蛋白和肌动蛋白探针)。Mac-1gp“上调”与 将研究过度坚持和分泌性事件以及可能的分泌性 新生儿Mac-1糖蛋白诱导和过度黏附受损的决定因素 将对PMN进行评估。亚细胞分离和一种新的刀豆蛋白A 将使用结合分析来确定细胞内的位置 MAC-1池并评估其表面表达的调节机制， 再分配和/或回收。选定致病机理会计 新生儿中性粒细胞MT组装紊乱包括：细胞异常 黏附依赖的细胞骨架激活，翻译后异常 微管蛋白酪氨酸化，或阿尔茨海默病的固有免疫化学异常 或β微管蛋白将通过细胞内免疫荧光进行评估 在促进MT组装或拆卸的条件下进行探测。这些 研究应提供一个基础，以了解功能和 慢性阻塞性肺疾病患者黏附依赖性细胞功能紊乱的临床后果 炎症反应，并有可能导致 在这些和其他高危儿科模型中的治疗策略。

Human neonates and individuals with heritable Mac-1, LFA1 deficiency are at high risk for the development of infectious complications resulting from impaired tissue mobilization of phagocytic cells. These studies will evaluate pathogenic mechanisms that may account for impaired PMN or mononuclear leukocyte mobilization in these populations. Major emphases will include determinations of: a) functional relationships between cellular adherence and migratory properties, b) mechanisms regulatin the induction, modulation and functional activation of cellular adherence by inflammatory stimuli, and c) possible pathologic influences of cytoskeletal proteins (microtubules) on cell translocation. They will focus on the contributions of a family of "adhesive" glycoproteins (GPs) (Mac-1, LFA1, p150,95) to adherence-dependent properties of leukocytes; comparative studies employing neonatal and Mac-1 deficient cells will allow a unique opportunity to understand functional and clinical consequences of abnormal expression. Surface expression of Mac-1 GP when mediated by chemotactic or secretory stimuli will be quantitated using immunofluorescence flow cytometry and 125I immunoprecipitation techniques employing monoclonal antibodies (MAb); their topographical relationships to other surface reptors (FcR and CR1) and to "adhesion sites" will be evaluated by IIF microscopy. MAbs to each Mac-1 subunit will be employed in blocking experiments to assess their functional contributions to adhesion-dependent cell functions including microtubule and microfilament assemblage (IIF tubulin and actin probes). The relationships of Mac-1 GP "up regulation", hyperadherence and secretory events will be studied and possible secretory determinants of impaired Mac-1 GP induction and hyperadherence of neonatal PMNs will be assessed. Subcellular fractionation and a new concanavalin A binding assay will be employed to determine the location of intracellular Mac-1 pools and to evaluate mechanisms regulating their surface expression, redistribution and/or recycling. Selected pathogenic mechanisms accounting for disturbed MT assembly of neonatal PMN including: abnormal cell adherence-dependent cytoskeletal activation, abnormal post-translational tubulin tyrosylation, or intrinsic immunochemical abnormalities of or Alpha or Beta tubulin will be evaluated with intracellular immunofluorescence probes under conditions promoting MT assembly or disassembly. These studies should provide a basis for understanding the functional and clinical consequences of disturbed adhesion-dependent cellular functions in the inflammatory response, and potentially allow the development of therapeutic strategies in these and other high risk pediatric models.