Interaction of microvesicles and bacterial toxins with immune cells

描述（由申请方提供）：避免免疫应答是许多病原体生物学的一个重要方面，并且已经定义了多种机制。对于革兰氏阳性细菌，属于胆固醇依赖性溶细胞素（CDC）家族的蛋白质的分泌可导致高浓度的某些细胞类型的裂解，但在较低浓度下也可通过TLR 4的参与使细胞（如巨噬细胞）对促凋亡诱导刺激物敏感。最近报道了由B产生的一种CDC--蒽溶素O。炭疽，以及其他一些CDC家族成员。然而，TLR的刺激可以被认为是一把双刃剑，因为这也可能刺激免疫应答，特别是当涉及有效的ARC如树突状细胞（DC）时。我们已经研究了三种CDC蛋白质，anthrolysin O，tetanolysin O和streptolysin O的作用，并观察到人DC通过流动钙，然后分泌或脱落其表面上具有II类MHC蛋白的微泡（MV），快速响应于用无毒水平的这些蛋白质的治疗。同时，DC失去了上调II类MHC的能力，以响应IPS。基于这些结果，我们假设CDC通过诱导含有II类MHC和其他对抗原呈递功能很重要的蛋白质的MV的分泌来损害DC的抗原呈递。此外，基于与称为外泌体的囊泡的相似性，我们认为释放的MV本身可能具有免疫抑制特性，并且CDC毒素可能散布在全身并被吞噬细胞吸收，以这种方式比感染部位更广泛地影响抗原呈递。我们提出进一步表征代表性CDC蛋白TLO对人单核细胞衍生DC功能的这些作用，并研究TLO诱导的MV对免疫应答的影响，目的如下：1.将确定TLO如何影响moDC中的II类本地化; 2.将表征TLO诱导的MV的蛋白质含量; 3.探讨细胞产生和摄取MV的机制; 4.将确定TLO诱导的和组成型产生的MV对CD 4和CD 8 T细胞功能的影响。外行陈述;了解细菌的毒素如何影响免疫反应，有助于设计出治疗破伤风、链球菌病和炭疽的新方法。

Description (provided by applicant): Avoidance of the immune response is an important aspect of the biology of many pathogens, and a wide variety of mechanisms have been defined. For Gram-positive bacteria, secretion of proteins belonging to the family of cholesterol dependent cytolysins (CDC) may lead to lysis of some cell types at high concentrations, but at lower concentrations may also sensitize cells such as macrophages to apoptosis-inducing stimuli by engagement of TLR4. This was recently reported for anthrolysin O, a CDC produced by B. anthracis, and for a number of other CDC family members. Stimulation of TLR however could be considered a double edged sword, since this might also stimulate the immune response, particularly when involving potent ARC such as dendritic cells (DC). We have investigated the effects of three CDC proteins, anthrolysin O, tetanolysin O and streptolysin O, and observed that human DC rapidly respond to treatment with non-toxic levels of these proteins by fluxing calcium and then secreting or shedding microvesicles (MV) that have on their surface class II MHC proteins. At the same time, DC lose the ability to up-regulate class II MHC in response to IPS. Based on these results, we hypothesize that CDC act to impair antigen presentation by DC by inducing secretion of MV containing class II MHC and other proteins important to antigen presenting function precisely at the time of pathogen encounter. Furthermore, we suggest that released MV may themselves have immunosuppressive properties, based on similarities to vesicles called exosomes, and that CDC toxins might be disseminated throughout the body and taken up by phagocytes, in this way impacting antigen presentation more widely than at the site of infection. We propose to further characterize these effects of a representative CDC protein, TLO, on the function of human monocyte-derived DC and to investigate the impact of TLO-induced MV on immune reponses in the following aims: 1. how TLO affects class II localization in moDC will be determined; 2. the protein contents of TLO-induced MV will be characterized; 3. the mechanism of MV production and uptake by cells will be probed; 4. the effect of TLO-induced and constitutively produced MV on CD4 and CDS T cell function will be determined. Lay statement; Understanding how toxins from bacteria affect the immune response could help devise new treatments for tetanus, streptococcal disease, and anthrax.