Martínez-Esparza M, Sarazin A, Jouy N, Poulain D, Jouault T
Departamento de Bioquímica y Biología Molecular B e Inmunología, Facultad de Medicina, Universidad de Murcia, Campus de Espinardo, 30100 Murcia, Spain.
J Immunol Methods. 2006 Jul 31;314(1-2):90-102. doi: 10.1016/j.jim.2006.06.004. Epub 2006 Jul 7.
The yeast Candida albicans is an opportunistic pathogen, part of the normal human microbial flora that causes infections in immunocompromised individuals with a high morbidity and mortality levels. Recognition of yeasts by host cells is based on components of the yeast cell wall, which are considered part of its virulence attributes. Cell wall glycans play an important role in the continuous interchange that regulates the balance between saprophytism and parasitism, and also between resistance and infection. Some of these molecular entities are expressed both by the pathogenic yeast C. albicans and by Saccharomyces cerevisiae, a related non-pathogenic yeast, involving similar molecular mechanisms and receptors for recognition. In this work we have exploited flow cytometry methods for probing surface glycans of the yeasts. We compared glycan expression by C. albicans and by S. cerevisiae, and studied the effect of culture conditions. Our results show that the expression levels of alpha- and beta-linked mannosides as well as beta-glucans can be successfully evaluated by flow cytometry methods using different antibodies independent of agglutination reactions. We also found that the surface expression pattern of beta-mannosides detected by monoclonal or polyclonal antibodies are differently modulated during the growth course. These data indicate that the yeast beta-mannosides exposed on mannoproteins and/or phospholipomannan are increased in stationary phase, whereas those linked to mannan are not affected by the yeast growth phase. The cytometric method described here represents a useful tool to investigate to what extent C. albicans is able to regulate its glycan surface expression and therefore modify its virulence properties.
白色念珠菌是一种机会致病菌,是人类正常微生物群落的一部分,可在免疫功能低下的个体中引发感染,且发病率和死亡率很高。宿主细胞对酵母的识别基于酵母细胞壁的成分,这些成分被认为是其毒力属性的一部分。细胞壁聚糖在调节腐生和寄生之间、抗性和感染之间平衡的持续交互作用中发挥着重要作用。其中一些分子实体在致病性酵母白色念珠菌和相关的非致病性酵母酿酒酵母中均有表达,涉及相似的分子机制和识别受体。在这项研究中,我们利用流式细胞术方法探测酵母的表面聚糖。我们比较了白色念珠菌和酿酒酵母的聚糖表达,并研究了培养条件的影响。我们的结果表明,使用不同抗体,通过流式细胞术方法可以成功评估α-和β-连接的甘露糖苷以及β-葡聚糖的表达水平,而无需依赖凝集反应。我们还发现,单克隆或多克隆抗体检测到的β-甘露糖苷的表面表达模式在生长过程中受到不同的调节。这些数据表明,暴露在甘露糖蛋白和/或磷脂甘露聚糖上的酵母β-甘露糖苷在稳定期增加,而与甘露聚糖相连的β-甘露糖苷不受酵母生长阶段的影响。这里描述的细胞计数方法是一种有用的工具,可用于研究白色念珠菌在多大程度上能够调节其聚糖表面表达,从而改变其毒力特性。