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POLYCLONAL B CELL ACTIVATION BY F NUCLEATUM CELL WALLS

F 核细胞壁激活多克隆 B 细胞

基本信息

批准号：
3220482
负责人：
Dennis F Mangan
金额：
$ 8.86万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
1984
资助国家：
美国
起止时间：
1984-01-01 至 1989-11-30
项目状态：
已结题

项目摘要

Fusobacterium nucleatum (FN) cells walls are potent stimulants of human polyclonal B lymphocyte activation (PBA). There is increasing evidence that PBA induced by FN and other gram negative plaque bacteria perpetuates chronic inflammation and is responsible for the unusually large number of activated B cells in periodontitis lesions. As part of our continuing study of nonspecific B cell activation in periodontal diseases, we intend to examine isolated macromolecules from FN cell walls and characterize B cell activation by these components. This proposal has two specific objectives. The first objective is to fractionate cell walls of FN by standard biochemical techniques into the following six fractions: cytoplasmic membranes, peptidoglycan, proteins/lipoproteins, lipopolysaccharide, lipid A, and polysaccharide. These fractions in other gram negative bacteria are potent PBA stimulants and, therefore, are likely candidates of FN-induced PBA. Each fraction will be tested for the capacity to induce PBA in human lymphocyte cultures. Active fractions will characterized and used in subsequent studies. The second objective is to investigate at the cellular level polyclonal B cell activation induced by the purified FN cell wall components. Lymphocytes for these studies will be obtained from peripheral blood of healthy donors. Using monoclonal antibodies, fluorescence activated cell sorting (FACS), and standard rosetting procedures, B and T lymphocytes will be separated and thorougly depleted of contaminating cell types. Initial studies will identify which subset(s) of B cells is activated and the percentage of these cells in peripheral blood. Next, multiple assays of B cell growth and differentiation will be utilized to characterize the events which occur during or following stimulation of B cells. Finally, regulation of PBA by resting or activated T cells will be studied by culturing stimulated B cells with graded doses of unseparated T cells, T cell subsets, or T cell-derived lymphokines.

具核梭杆菌（FN）细胞壁是有效的刺激物人多克隆B淋巴细胞活化（PBA）。有越来越多的证据表明，由FN和其他革兰氏阴性杆菌诱导的PBA 阴性菌斑细菌使慢性炎症持续存在，导致异常大量的活化B细胞，牙周炎病变。作为我们继续研究的一部分，非特异性B细胞活化在牙周病，我们打算检查从FN细胞壁分离的大分子，表征这些组分对B细胞的活化。这项建议有两个具体目标。第一个目标是通过标准生物化学技术破碎FN细胞壁分成以下六个部分：细胞质膜，肽聚糖，蛋白质/脂蛋白，脂多糖，脂质A，和多糖。其他革兰阴性菌中的这些分数细菌是有效的PBA刺激物，因此， FN诱导的PBA的候选者。将检测每个馏分的在人淋巴细胞培养物中诱导PBA的能力。活性组分将被表征并用于随后的研究中。问题研究第二个目标是在细胞水平上进行研究纯化的FN细胞壁诱导的多克隆B细胞活化件. 用于这些研究的淋巴细胞将从健康献血者的外周血。使用单克隆抗体，荧光激活细胞分选（FACS）和标准玫瑰花结程序中，将分离B和T淋巴细胞，耗尽了污染细胞类型。初步研究将确定 B细胞的哪个亚群被激活，以及这些亚群的百分比外周血中的细胞。接下来，对B细胞生长的多种测定和差异将被用来表征事件其发生在B细胞刺激期间或之后。最后，将研究静息或活化T细胞对PBA的调节通过用分级剂量的未分离的T细胞培养刺激的B细胞，细胞、T细胞亚群或T细胞衍生的淋巴因子。