METHODS: Five patients with PCR-confirmed P. knowlesi malaria were recruited into the study with consent between April and August 2010. Pre-treatment venous blood was washed and cultured ex vivo to increase the proportion of schizont-infected erythrocytes. Cultured blood was seeded into Petri dishes with triplicate areas coated with ICAM-1, VCAM and CD36. Following incubation at 37°C for one hour the dishes were washed and the number of infected erythrocytes bound/mm2 to PBS control areas and to recombinant human ICAM-1 VCAM and CD36 coated areas were recorded. Each assay was performed in duplicate. Assay performance was monitored with the Plasmodium falciparum clone HB3.
RESULTS: Blood samples were cultured ex vivo for up to 14.5 h (mean 11.3 ± 1.9 h) to increase the relative proportion of mature trophozoite and schizont-infected red blood cells to at least 50% (mean 65.8 ± 17.51%). Three (60%) isolates bound significantly to ICAM-1 and VCAM, one (20%) isolate bound to VCAM and none of the five bound significantly to CD36.
CONCLUSIONS: Plasmodium knowlesi infected erythrocytes from human subjects bind in a specific but variable manner to the inducible endothelial receptors ICAM-1 and VCAM. Binding to the constitutively-expressed endothelial receptor CD36 was not detected. Further work will be required to define the pathological consequences of these interactions.
方法:2010年4月至8月,5名经PCR证实的诺氏疟原虫患者被纳入研究。洗涤预处理静脉血并离体培养以增加裂殖体感染的红细胞的比例。将培养的血液接种到具有用ICAM-1、VCAM和CD36包被的一式三份区域的培养皿中。在37°C下孵育1小时后,洗涤培养皿,并记录与PBS对照区域和重组人ICAM-1VCAM和CD36包被区域结合的感染红细胞数/mm2。每个测定一式两份进行。用恶性疟原虫克隆HB3监测测定性能。
结果:将血液样品离体培养长达14.5h(平均11.3±1.9h),以将成熟滋养体和裂殖体感染的红细胞的相对比例增加到至少50%(平均65.8±17.51%)。三个(60%)分离株与ICAM-1和VCAM显著结合,一种(20%)分离物结合VCAM,并且五种中没有一个显著结合CD36。
结论:来自人类受试者的诺氏疟原虫感染的红细胞以特异性但可变的方式与诱导型内皮受体ICAM-1和VCAM结合。未检测到与组成型表达的内皮受体CD36的结合。需要进一步的工作来定义这些相互作用的病理后果。