Davis R E, Thalhofer C J, Wilson M E
Interdisciplinary Program in Immunology, University of Iowa, Iowa City, IA, USA.
Agonox Inc., Portland, OR 97213.
J Immunol Tech Infect Dis. 2016;5(3). doi: 10.4172/2329-9541.1000146. Epub 2016 May 10.
Neutrophils (PMNs) are recruited in high numbers to sites of host infection by the protozoan parasites of the genus Leishmania. Although PMNs are capable of phagocytizing Leishmania parasites and are potent producers of anti-microbial compounds including reactive oxygen species (ROS), they are unable to control the establishment of infection. Prior studies document production of ROS in isolated PMNs incubated with Leishmania under conditions allowing phagocytosis, but without a measure of single cells' responses it cannot be discerned whether PMN activation and ROS production is suppressed or ineffective in the cells that internalize the parasite. To address these interactions, we engineered a strain of fluorescent, mCherry-expressing (mCherry-Li). By infecting isolated human PMNs in vitro with mCherry-Li, we observed ready association of the parasites with PMNs in a time- and dose-dependent fashion. We also examined production of PMN ROS (using the fluorescent compound DHR123) and PMN activation (as evidence by loss of surface CD62L expression). Whereas many Li-associated (mCherry) PMNs responded to parasite interactions and uptake with ROS production and/or activation, a proportion exhibited neither response. Furthermore, a large proportion of mCherry - "bystander" PMNs displayed both ROS production and activation. The heterogeneous response of PMNs to Leishmania exposure leads us to hypothesize, first, that some PMNs exhibit decreased activation upon phagocytosis of Leishmania, and could support their maintenance. Second, responses of bystander PMNs may contribute to a local inflammatory environment that is ineffective at parasite clearance.
中性粒细胞(PMNs)会大量募集到利什曼原虫属原生动物的宿主感染部位。尽管中性粒细胞能够吞噬利什曼原虫寄生虫,并且是包括活性氧(ROS)在内的抗菌化合物的强大生产者,但它们无法控制感染的建立。先前的研究记录了在允许吞噬作用的条件下,与利什曼原虫一起孵育的分离中性粒细胞中ROS的产生,但如果没有对单细胞反应的测量,就无法辨别在吞噬寄生虫的细胞中,中性粒细胞的激活和ROS的产生是受到抑制还是无效。为了解决这些相互作用,我们构建了一种表达荧光mCherry的菌株(mCherry-Li)。通过在体外用人源分离的中性粒细胞感染mCherry-Li,我们观察到寄生虫与中性粒细胞以时间和剂量依赖性方式迅速结合。我们还检测了中性粒细胞ROS的产生(使用荧光化合物DHR123)和中性粒细胞的激活(以表面CD62L表达的丧失为证据)。虽然许多与利什曼原虫相关(mCherry)的中性粒细胞对寄生虫的相互作用和摄取有反应,产生ROS和/或激活,但有一部分没有反应。此外,很大一部分mCherry“旁观者”中性粒细胞既产生ROS又被激活。中性粒细胞对利什曼原虫暴露的异质性反应使我们推测,首先,一些中性粒细胞在吞噬利什曼原虫后激活能力下降,并可能支持它们的存活。其次,旁观者中性粒细胞的反应可能会导致局部炎症环境,而这种环境在清除寄生虫方面无效。
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