The role of parasitic phosphatidylserine in the infection of human polymophonuclear neutrophil granulocytes (PMN) by Leishmania major

专性胞内病原体大利什曼原虫以吞噬细胞为宿主细胞。为了建立一种有效的感染，吞噬细胞的防御被灭活。为了繁殖，寄生虫使用它的最终宿主细胞--巨噬细胞。然而，在到达最终目的地之前，主要是招募和感染中性粒细胞(PMN)。然后，寄生虫利用感染的凋亡PMN作为运输工具进入巨噬细胞。因此，PMN防御必须首先被灭活，才能使寄生虫存活。非炎症性进入吞噬细胞的最好例子是吞噬凋亡的细胞。免疫反应是通过识别表达在凋亡细胞外膜上的磷脂酰丝氨酸(PS)来抑制的。我们发现，从体外培养或从沙蝇载体中提取的有感染性的前鞭毛体种群中含有凋亡的PS阳性寄生虫。在体内，在皮肤感染模型中，感染纯化的PS阴性寄生虫的敏感BALB/c小鼠没有发生疾病。疾病的发展依赖于PS阳性寄生虫的存在。这些数据表明PS在疾病发展中是一个关键的媒介。在体外，PS阳性寄生虫通过一种未知的机制使PS阴性寄生虫在PMN中存活。在这个项目中，我们的主要目标将是阐明PS介导的抑制PMN激活的机制。我们将比较PS在前鞭毛体和凋亡细胞上的表达。我们将研究PMN摄取PS依赖和PS非依赖寄生虫的分子。在PS阳性寄生虫不存在或存在的情况下，我们将评估PMN对PS阴性寄生虫的免疫反应。最后，我们将产生一种不能表达PS的突变寄生虫菌株。我们的研究将有助于了解利什曼原虫感染的病理生理机制，并可能为新的治疗策略提供基础。

The obligate intracellular pathogen Leishmania major (L. major) uses phagocytes as host cells. To establish a productive infection phagocyte defenses are inactivated. For multiplication the parasite uses its final host cell, the macrophage. However, before reaching its final destination L. major first recruits and infects polymorphonuclear neutrophil granulocytes (PMN). Infected apoptotic PMN are then used by the parasite as transport vehicle to enter macrophages. Consequently, PMN defenses have to be inactivated first to enable parasite survival. The best example of a non inflammatory entry into phagocytes is the uptake of apoptotic cells. Immune responses are suppressed by the recognition of phosphatidyl serine (PS) expressed on the outer membrane of apoptotic cells. We found that infective promastigotes populations taken either from in vitro cultures or from the sand fly vector contain apoptotic PS-positive parasites. In vivo, in a cutaneous infection model susceptible BALB/c mice infected with purified PS-negative parasites did not develop disease. Disease development depended on the presence of PS-positive parasites. These data suggest that PS is a key mediator in disease development. In vitro, PS-positive parasites enable survival of PS-negative parasites in PMN by a still unknown mechanism. In this project, our main objective will be to elucidate the mechanism of PS-mediated suppression of PMN activation. We will compare PS expression on promastigotes with apoptotic cells. We will study the molecules involved in both PS-dependent and PS-independent uptake of parasites by PMN. In the absence or presence of PS-positive parasites, we will assess PMN immune responses towards viable PS-negative parasites. Finally we will generate a mutant parasite strain unable to express PS. Our investigations will help to understand the pathophysiological mechanisms of Leishmania infections and may provide the base for new therapeutic strategies