Molecular mechanisms of anthrax toxin-mediated edema

炭疽毒素介导的水肿的分子机制

• 批准号: 7678823
• 负责人: JEFFREY M TESSIER
• 金额: $ 10.21万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-03-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7678823
• 关键词: Address; Afferent Neurons; Anthrax disease; BALB/cJ Mouse; Bacillus anthracis; Biological; Bone Marrow; Cells; Cessation of life; Clinical; Conditioned Culture Media; Contracts; Cultured Cells; Cyclic AMP; Dinoprostone; Edema; Endothelial Cells; Escherichia coli; Event; Functional disorder; Funding; Goals; Hand; Histamine; Histamine H1 Antagonists; Histamine Release; Human; In Vitro; Inflammatory; Intervention; Intoxication; K-Series Research Career Programs; Knockout Mice; Laboratories; Measures; Mediating; Mediator of activation protein; Messenger RNA; Molecular; Mus; Neurons; Oryctolagus cuniculus; PTGS2 gene; Pathogenesis; Pathology; Pharmaceutical Preparations; Preparation; Production; Prostaglandin Production; Prostaglandins; Proteins; Pyrilamine; Rattus; Role; Signs and Symptoms; Skin; Source; Spinal Ganglia; Statistically Significant; Substance P; Testing; Time; Toxin; Work; anthrax toxin; base; celecoxib; cell type; ganglion cell; in vivo; inhibitor/antagonist; mRNA Expression; macrophage; mast cell; monocyte; mortality; preprotachykinin A; response

The objectives of this project, which is supported by a MARGE Career Development Award, are: 1) to dentify the cell types through which ET initiates and sustains the release of mediators, such as histamine, prostanoids and neurokinins, that are involved in ET-mediated edema and; 2) determine if these same mediators are involved in/responsible for ET-induced death in mice. Although we have demonstrated that ET s able to elicit COX-2 expression, using HEK cells, we have not observed an increase in COX-2 mRNA over basal, in human monocyte-derived monocytes. On the other hand, ET elicits a small increase in PGE2 synthesis from RAW264.7 macrophages and the effect is synergistic with LPS-induced prostaglandin production in these cells. To address the hypothesis that the mediator cascade initiated by ET might begin with or include direct stimulation of Substance P (SP) release from neuronal cells, dorsal root ganglion (DRG) cells were isolated from rats and rabbits and exposed to ET in vitro. Thus far, we have not detected a statistically significant release of SP from either of these cells types. Based on our earlier work showing a histamine (H1) antagonist (pyrilamine) and an inhibitor of COX-2 (celecoxib) cause a reduction in edema produced in rabbit skin by intradermal ET, we have tested these to drugs for their effect on ET-induced mortality in of BALB/cJ mice. While neither drug had a significant effect alone, the combination resulted in a reduced mortality and prolonged time to death. In these studies, we noted however, that only ET composed of E. coli-derived EF had this effect and the EF produced in B.anthracis did not cause any mortality. In order to address that discrepancy, we obtained funding through an NIAID/BEI contract, for which we are doing a direct comparison of the ET from these two sources for enzymatic and in vitro toxin activities, as well as in vivo effects in mice and rabbits. This work is ongoing, but there appears to be a significant and reproducible difference in the toxin effects (both in vitro and in vivo) between these two preparations of EF. In the second year of this career development award, we will continue to study the effect of ET on COX-2 induction in different cell types, measure histamine release from mast cells (murine and human) that are exposed to the conditioned medium of ET-treated DRG cells, characterize the effects of pharmacological antagonists on ET-associated clinical signs, laboratory abnormalities, pathological changes, and death in mice and utilize COX-2 and preprotachykinin A-knockout mice, and mast cell-deficient mice to investigate the roles of prostanoids, substance P and mast cells in ET-induced murine pathology/mortality.

该项目得到了MARGE职业发展奖的支持，其目标是：1） 确定ET通过其启动和维持介质释放的细胞类型，例如组胺， 前列腺素和神经激肽，参与ET介导的水肿; 2）确定这些相同的 介质参与/负责ET诱导的小鼠死亡。虽然我们已经证明ET 尽管HEK细胞能够诱导考克斯-2表达，但我们没有观察到考克斯-2 mRNA的增加超过 基础，在人单核细胞衍生的单核细胞中。另一方面，ET使PGE 2略有增加， 从RAW264.7巨噬细胞合成，并且与LPS诱导的前列腺素具有协同作用 在这些细胞中生产。为了解决由ET启动的介质级联可能开始的假设， 有或包括直接刺激P物质（SP）从神经元细胞、背根神经节释放 (DRG)从大鼠和兔中分离细胞并在体外暴露于ET。到目前为止，我们还没有发现 从这些细胞类型中的任一种中统计学显著地释放SP。 基于我们早期的工作显示组胺（H1）拮抗剂（吡拉明）和考克斯-2抑制剂 （塞来昔布）引起兔皮肤皮内ET产生的水肿减少，我们已经测试了这些， 药物对ET诱导的BALB/cJ小鼠死亡率的影响。虽然这两种药物都没有明显的效果， 单独使用时，联合用药导致死亡率降低和死亡时间延长。在这些研究中，我们 然而，我们注意到，只有ET由E.大肠杆菌来源的EF具有这种作用， B. anthracis未引起任何死亡。为了解决这一矛盾，我们通过以下方式获得资金： 一个NIAID/BEI合同，我们正在对这两个来源的ET进行直接比较， 酶和体外毒素活性，以及小鼠和家兔体内效应。这项工作正在进行中，但 在毒素效应方面似乎存在显著和可重复的差异（体外和体内） 这两种EF制剂之间的差异。 在这个职业发展奖的第二年，我们将继续研究ET对考克斯-2的影响 在不同细胞类型中诱导，测量从肥大细胞（鼠和人）释放组胺， 暴露于ET处理的DRG细胞的条件培养基，表征药理学作用 拮抗剂对ET相关临床体征、实验室异常、病理学变化和死亡的影响 小鼠，并利用考克斯-2和前速激肽原A基因敲除小鼠和肥大细胞缺陷小鼠进行研究 前列腺素、P物质和肥大细胞在ET诱导的小鼠病理/死亡中的作用。