Uptake of Aspergillus fumigatus conidia by phagocytic and nonphagocytic cells in vitro:: Quantitation using strains expressing green fluorescent protein
Uptake of Aspergillus fumigatus conidia by phagocytic and nonphagocytic cells in vitro:: Quantitation using strains expressing green fluorescent protein
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DOI：
10.1128/iai.70.6.3156-3163.2002
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发表时间：
2002-06-01
影响因子：
3.1
通讯作者：
Moore, MM
中科院分区：
文献类型：
--
作者：
Wasylnka, JA;Moore, MM
几种病原真菌进入真核细胞并操纵宿主细胞环境以有利于其自身的生长和生存。烟曲霉是一种腐生真菌，可在免疫功能低下的宿主中引起侵袭性肺病。为了确定烟曲霉是否可以进入真核细胞，我们在体外研究了两种不同的表达 GFP 的烟曲霉菌株对 A549 肺上皮细胞、人脐静脉内皮 (HUVE) 细胞和 J774 鼠巨噬细胞的摄取。 A549 细胞内化 30% 的结合分生孢子，而 HUVE 和 J774 细胞分别内化 50 和 9011。 A549细胞内的分生孢子保持活力6小时；然而，仅 4 小时后，J774 细胞内 60% 至 80% 的分生孢子就被杀死，活的和热杀死的分生孢子被 A549 细胞内化到相同的程度。 6小时后，A549细胞内的分生孢子几乎没有萌发，而细胞外的分生孢子已发育出芽管。 A549 细胞对分生孢子的内化是一个温度依赖性过程，需要重新排列底层宿主细胞的细胞骨架； 0.5 μM 细胞松弛素 D 的摄取被抑制 75%，5 μM 秋水仙碱的摄取被抑制 65%。用罗丹明鬼笔环肽对感染的 A549 细胞进行荧光标记，提供了细胞骨架改变的明显证据，因为许多细胞内分生孢子包含在肌动蛋白包被的吞噬体中。这些数据提供的证据表明，大量的烟曲霉分生孢子可以在体外被非专业吞噬细胞内化，并且这些细胞可以作为免疫细胞逃避和在整个宿主中传播的储存库。
Several pathogenic fungal organisms enter eukaryotic cells and manipulate the host cell environment to favor their own growth and survival. Aspergillus fumigatus is a saprophytic fungus that causes invasive lung disease in the immunocompromised host. To determine whether A. fumigatus could enter eukaryotic cells, we studied the uptake of two different GFP-expressing A. fumigatus strains into A549 lung epithelial cells, human umbilical vein endothelial (HUVE) cells, and J774 murine macrophages in vitro. A549 cells internalized 30% of the bound conidia whereas HUVE and J774 cells internalized 50 and 9011, respectively. Conidia within A549 cells remained viable for 6 h; however, 60 to 80% of conidia within J774 cells were killed after only 4 h, Live and heat-killed conidia were internalized to the same extent by A549 cells. After 6 h, almost none of the conidia inside A549 cells had germinated, whereas extracellular conidia had developed germ tubes. Internalization of conidia by A549 cells was a temperature-dependent process and required rearrangement of the underlying host cell cytoskeleton; uptake was inhibited by 75% with 0.5 muM cytochalasin D and by 65% with 5 muM colchicine. Fluorescent labeling of infected A549 cells with rhodamine phalloidin provided visible evidence of cytoskeletal alteration as many of the intracellular conidia were contained in actin-coated phagosomes. These data provide evidence that significant numbers of A. fumigatus conidia can be internalized by nonprofessional phagocytes in vitro and these cells may serve as reservoirs for immune cell evasion and dissemination throughout the host.