Danley D L, Polakoff J
Infect Immun. 1986 Jan;51(1):307-13. doi: 10.1128/iai.51.1.307-313.1986.
To study the interaction between Candida albicans blastoconidia and human phagocytes, we incubated peripheral leukocytes with fungi for 1 h at 37 degrees C and stained the cells with fluorescent vital stains ethidium bromide (EB) and fluorescein diacetate. Fungi that had been phagocytosed showed little staining; however, some leukocytes containing blastoconidia exhibited nuclear staining with EB, even though their cell membranes showed no signs of penetration by fungi. The number of EB-positive leukocytes was related to viability of the yeast cells and the temperature at which they were maintained before use. Because efforts to quantitate EB-positive leukocytes microscopically were frustrated by cell aggregation, we labeled the leukocytes with 51Cr and measured isotope release. We determined that leukocytes incubated with viable fungi released significantly more isotope than cells incubated alone or with killed blastoconidia. Furthermore, 51Cr release correlated directly with concentration of fungi in the assay, time of incubation, and temperature at which fungi were maintained before use. Using a number of isolates of C. albicans and several other species of Candida, we found that all exhibited cytotoxic activity against leukocytes, but the level of activity varied among organisms. Finally, we depleted or enriched peripheral leukocytes for specific cell populations and determined that only monocytes released more 51Cr after incubation with viable blastoconidia. Blastoconidia can lyse phagocytic cells through germination and penetration of cell membranes within 1 to 2 h, but the cytotoxic phenomenon we describe occurs within 15 to 30 min after yeast cells have been phagocytosed. Therefore, this capacity may represent a more immediate response by blastoconidia against phagocytosis and killing by monocytes.
为研究白色念珠菌芽生孢子与人类吞噬细胞之间的相互作用，我们将外周血白细胞与真菌在37℃孵育1小时，并用荧光活体染料溴化乙锭（EB）和荧光素二乙酸酯对细胞进行染色。被吞噬的真菌几乎没有染色；然而，一些含有芽生孢子的白细胞却显示出EB核染色，尽管其细胞膜没有真菌穿透的迹象。EB阳性白细胞的数量与酵母细胞的活力以及使用前保存的温度有关。由于细胞聚集使通过显微镜定量EB阳性白细胞的努力受挫，我们用51Cr标记白细胞并测量同位素释放。我们确定，与活真菌孵育的白细胞比单独孵育或与灭活芽生孢子孵育的细胞释放出显著更多的同位素。此外，51Cr释放与检测中真菌的浓度、孵育时间以及使用前真菌保存的温度直接相关。使用多种白色念珠菌分离株和其他几种念珠菌属菌种，我们发现所有菌株均对白细胞表现出细胞毒性活性，但活性水平在不同菌株间有所差异。最后，我们对外周血白细胞进行特定细胞群的耗竭或富集，确定只有单核细胞在与活芽生孢子孵育后释放出更多的51Cr。芽生孢子可在1至2小时内通过发芽和穿透细胞膜来裂解吞噬细胞，但我们描述的细胞毒性现象发生在酵母细胞被吞噬后15至30分钟内。因此，这种能力可能代表芽生孢子对单核细胞吞噬和杀伤的一种更即时的反应。
