Marcil A, Harcus D, Thomas D Y, Whiteway M
Genetics Group, Biotechnology Research Institute, National Research Council of Canada, Montreal, Quebec H4P 2R2, USA.
Infect Immun. 2002 Nov;70(11):6319-29. doi: 10.1128/IAI.70.11.6319-6329.2002.
Phagocytic cells such as neutrophils and macrophages are potential components of the immune defense that protects mammals against Candida albicans infection. We have tested the interaction between the mouse macrophage cell line RAW 264.7 and a variety of mutant strains of C. albicans. We used an end point dilution assay to monitor the killing of C. albicans at low multiplicities of infection (MOIs). Several mutants that show reduced virulence in mouse systemic-infection models show reduced colony formation in the presence of macrophage cells. To permit analysis of the macrophage-Candida interaction at higher MOIs, we introduced a luciferase reporter gene into wild-type and mutant Candida cells and used loss of the luminescence signal to quantify proliferation. This assay gave results similar to those for the end point dilution assay. Activation of the macrophages with mouse gamma interferon did not enhance anti-Candida activity. Continued coculture of the Candida and macrophage cells eventually led to death of the macrophages, but for the RAW 264.7 cell line this was not due to apoptotic pathways involving caspase-8 or -9 activation. In general Candida cells defective in the formation of hyphae were both less virulent in animal models and more sensitive to macrophage engulfment and growth inhibition. However the nonvirulent, hypha-defective cla4 mutant line was considerably more resistant to macrophage-mediated inhibition than the wild-type strain. Thus although mutants sensitive to engulfment are typically less virulent in systemic-infection models, sensitivity to phagocytic macrophage cells is not the unique determinant of C. albicans virulence.
吞噬细胞,如中性粒细胞和巨噬细胞,是免疫防御的潜在组成部分,可保护哺乳动物免受白色念珠菌感染。我们测试了小鼠巨噬细胞系RAW 264.7与多种白色念珠菌突变菌株之间的相互作用。我们使用终点稀释法监测在低感染复数(MOIs)下白色念珠菌的杀伤情况。在小鼠全身感染模型中显示毒力降低的几种突变体,在巨噬细胞存在的情况下菌落形成减少。为了在更高的MOIs下分析巨噬细胞与念珠菌的相互作用,我们将荧光素酶报告基因导入野生型和突变型念珠菌细胞,并利用发光信号的丧失来量化增殖。该检测结果与终点稀释法相似。用小鼠γ干扰素激活巨噬细胞并没有增强抗念珠菌活性。念珠菌和巨噬细胞的持续共培养最终导致巨噬细胞死亡,但对于RAW 264.7细胞系来说,这不是由于涉及半胱天冬酶-8或-9激活的凋亡途径。一般来说,在菌丝形成方面有缺陷的念珠菌细胞在动物模型中毒力较低,对巨噬细胞吞噬和生长抑制更敏感。然而,无毒力、菌丝缺陷的cla4突变株对巨噬细胞介导的抑制作用比野生型菌株有更强的抗性。因此,尽管在全身感染模型中,对吞噬敏感的突变体通常毒力较低,但对吞噬性巨噬细胞的敏感性不是白色念珠菌毒力的唯一决定因素。