Saibabu Venkata, Singh Shweta, Ansari Moiz A, Fatima Zeeshan, Hameed Saif
Amity Institute of Biotechnology, Amity University Haryana, Gurugram (Manesar), India.
Rev Soc Bras Med Trop. 2017 Jul-Aug;50(4):524-529. doi: 10.1590/0037-8682-0114-2017.
Citronellal (Cit) possesses antifungal activity and has possible implications for reactive oxygen species (ROS) generation in Candida albicans. In this study, the effects of Cit on ROS generation and the mechanisms by which Cit exerts anti-Candida effects were examined.
A 2',7'-dichlorodihydrofluorescein diacetate assay was used to assess oxidative damage. Cell necrosis was determined by flow cytometry after FITC-Annexin V staining. Mitochondrial function was studied based on mitochondrial potential, metabolic activity (MTT assay), and phenotypic susceptibility on a non-fermentable carbon source. Membrane intactness and DNA damage were estimated by a propidium iodide (PI) uptake assay and 4',6-diamidino-2-phenylindole (DAPI) staining.
ROS generation was enhanced in response to Cit, leading to necrosis (2%). Additional hallmarks of cell death in response to Cit, such as mitochondrial membrane depolarization and DNA damage, were also observed. Cit treatment resulted in dysfunctional mitochondria, as evidenced by poor labeling with the mitochondrial membrane potential-sensitive probe rhodamine B, reduced metabolic activity (61.5%), and inhibited growth on a non-fermentable carbon source. Furthermore, Cit induced DNA damage based on DAPI staining. These phenotypes were reinforced by RT-PCR showing differences in gene expression (30-60%) between control and Cit-treated cells. Finally, PI uptake in the presence of sodium azide confirmed non-intact membranes and suggested that Cit activity is independent of the energy status of the cell.
Cit possesses dual anticandidal mechanisms, including membrane-disruptive and oxidative damage. Taken together, our data demonstrated that cit could be used as a prominent antifungal drug.
香茅醛（Cit）具有抗真菌活性，可能与白色念珠菌中活性氧（ROS）的产生有关。在本研究中，检测了香茅醛对ROS产生的影响以及其发挥抗念珠菌作用的机制。
采用2',7'-二氯二氢荧光素二乙酸酯检测法评估氧化损伤。通过FITC-Annexin V染色后用流式细胞术测定细胞坏死情况。基于线粒体电位、代谢活性（MTT检测）以及在非发酵碳源上的表型敏感性研究线粒体功能。通过碘化丙啶（PI）摄取检测和4',6-二脒基-2-苯基吲哚（DAPI）染色评估膜完整性和DNA损伤。
香茅醛刺激后ROS生成增加，导致坏死（2%）。还观察到香茅醛引起的细胞死亡的其他特征，如线粒体膜去极化和DNA损伤。香茅醛处理导致线粒体功能障碍，这可通过线粒体膜电位敏感探针罗丹明B标记不佳、代谢活性降低（61.5%）以及在非发酵碳源上生长受抑制来证明。此外，基于DAPI染色，香茅醛诱导了DNA损伤。这些表型通过RT-PCR得到加强，显示对照细胞和经香茅醛处理的细胞之间基因表达存在差异（30 - 60%）。最后，在叠氮化钠存在下的PI摄取证实了膜不完整，并表明香茅醛的活性与细胞的能量状态无关。
香茅醛具有双重抗念珠菌机制，包括膜破坏和氧化损伤。综上所述，我们的数据表明香茅醛可作为一种重要的抗真菌药物。
