Peralta Mariana Andrea, da Silva María Angel, Ortega María Gabriela, Cabrera José Luis, Paraje María Gabriela
Departamento de Farmacia, Facultad de Ciencias Químicas, Instituto Multidisciplinario de Biología Vegetal (IMBIV) - Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Córdoba, Haya de la Torre y Medina Allende, 5000 Córdoba, Argentina.
Facultad de Ciencias Exactas Físicas y Naturales, Instituto Multidisciplinario de Biología Vegetal (IMBIV) - Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET); Cátedra de Microbiología, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 299, Córdoba, Argentina.
Phytomedicine. 2015 Oct 15;22(11):975-80. doi: 10.1016/j.phymed.2015.07.003. Epub 2015 Jul 26.
The continuing emergence of infections with antifungal resistant Candida strains requires a constant search for new antifungal drugs, with the plant kingdom being an important source of chemical structures.
The present study investigated the antifungal effect of 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-8-prenylpinocembrin (8PP, formerly 6PP), a natural prenylflavonoid, on Candida albicans biofilms, and compared this with an azole antifungal (fluconazole) by studying the cellular stress and antioxidant response.
STUDY DESIGN/METHODS: The fluconazole sensitive (SCa) and azole-resistant (RCa) C. albicans strains were used, with biofilm formation being studied using crystal violet (CV) and confocal scanning laser microscopy (CSLM). The minimal inhibitory concentration for sessile cells (SMIC) was defined as the concentration of antifungal that caused a 50% (SMIC 50) and 80% (SMIC 80) reduction of treated biofilms. The reactive oxygen species (ROS) were detected by the reduction of nitro blue tetrazolium (NBT), and reactive nitrogen intermediates (RNI) were determined by the Griess assay. The activities of the superoxide dismutase (SOD) and catalase (CAT) antioxidant enzymes and the total antioxidant capacity of the biofilms were measured by spectrophotometric methods. ROS accumulation was also detected inside biofilms by using the fluorogenic dye 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA), which was visualized by CSLM.
The SCa and RCa biofilms were strongly inhibited by 8PP at 100 µM (SMIC 80). We observed that cellular stress affected biofilms growth, resulting in an increase of ROS and also of reactive nitrogen intermediates (RNI), with SOD and CAT being increased significantly in the presence of 8PP. The basal level of the biofilm total antioxidant capacity was higher in RCa than SCa. Moreover, in SCa, the total antioxidant capacity rose considerably in the presence of both 8PP and fluconazole.
Our data suggest that 8PP may be useful for the treatment of biofilm-related Candida infections, through an accumulation of endogenous ROS and RNI that can induce an adaptive response based on a coordinated increase in antioxidant defenses. 8PP may also have a therapeutic potential in C. albicans infections.
对抗真菌耐药念珠菌菌株感染的持续出现，需要不断寻找新的抗真菌药物，植物界是化学结构的重要来源。
本研究调查了天然异戊烯基黄酮2',4'-二羟基-5'-(1''',1'''-二甲基烯丙基)-8-异戊烯基松属素（8PP，原6PP）对白色念珠菌生物膜的抗真菌作用，并通过研究细胞应激和抗氧化反应，将其与唑类抗真菌药（氟康唑）进行比较。
研究设计/方法：使用氟康唑敏感（SCa）和唑类耐药（RCa）的白色念珠菌菌株，通过结晶紫（CV）和共聚焦扫描激光显微镜（CSLM）研究生物膜形成。无浮游细胞的最小抑菌浓度（SMIC）定义为导致处理后生物膜减少50%（SMIC 50）和80%（SMIC 80）的抗真菌药物浓度。通过硝基蓝四唑（NBT）还原检测活性氧（ROS），通过格里斯（Griess）测定法测定活性氮中间体（RNI）。通过分光光度法测量超氧化物歧化酶（SOD）和过氧化氢酶（CAT）抗氧化酶的活性以及生物膜的总抗氧化能力。还使用荧光染料2',7'-二氯二氢荧光素二乙酸酯（DCFH-DA）在生物膜内检测ROS积累，并通过CSLM进行可视化。
100μM的8PP（SMIC 80）对SCa和RCa生物膜有强烈抑制作用。我们观察到细胞应激影响生物膜生长，导致ROS以及活性氮中间体（RNI）增加，在8PP存在下SOD和CAT显著增加。RCa生物膜的总抗氧化能力基础水平高于SCa。此外，在SCa中，8PP和氟康唑存在时总抗氧化能力显著上升。
我们的数据表明，8PP可能通过内源性ROS和RNI的积累，诱导基于抗氧化防御协同增加的适应性反应，从而用于治疗与生物膜相关的念珠菌感染。8PP在白色念珠菌感染中也可能具有治疗潜力。
