Inhibitory Effect of Shikonin on <i>Candida albicans</i> Growth

Abstract

Our study showed that Shikonin (SK) could provide an action against almost all Candida albicans isolates tested. More importantly, to some Fluconazole (FCZ)-resistant Candida albicans, the action of SK (MIC 80 value 4 µg/mL) was shown to be >16 times higher than that of FCZ (MIC 80 >64 µg/mL). To clarify the mechanism underlying this action, we performed a comparative study in untreated control C. albicans and C. albicans treated with SK. In this study, we found that SK treatment increased generation of endogenous reactive oxygen species (ROS) and decreased mitochondrial membrane potential. Furthermore, anti-oxidants N-acetylcysteine (NAC) and glutathione (GSH) could reduce the antifungal activity of SK significantly in C. albicans. Our analyses also identified 9 differentially expressed genes, which were related to glycolysis-related genes (CDC19 and HXK2), fermentation-related genes (ALD5 and ADH1), antioxidant defense-related genes (SOD2 and SOD5), thioredoxin reductase-related gene (TRR1), mitochondrial respiratory electron transport chain-related gene (MRF1) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidoreductase-related gene (EBP1). These results suggest that mitochondrial aerobic respiration shift and endogenous ROS augmentation contribute to the action of SK against C. albicans. Key words Candida albicans; shikonin; reactive oxygen species; mitochondrial membrane potentialCandida albicans, the major opportunistic fungal pathogen of humans, normally exist as commensal. However, in patients with an impaired immune system such as those suffering from acquired immunodeficiency syndrome or cancer, C. albicans can cause recurrent superficial infection and fatal deep-seated and disseminated candidiasis. 1-4)Treatment for C. albicans includes antifungal drugs, mainly the azoles. Unfortunately, with the increasing clinical use of the azoles, azoles-resistant isolates are occurring more frequently. 5,6) Thus the development of new, effective antifungal agents is strongly needed in medicine.Shikonin (SK) (Fig. 1) is set in the orient, originally in China. This molecule is the major constituent of the red pigment extracts from the roots of the plant Lithospermum erythrorhizon SIEB. et ZUCC (LE). SK is widely used as a material to prepare an ointment called "Shiun-ko" which is used to treat wounds, burn and hemorrhoids in Japan. 7,8) Our study found that SK exhibit significant antifungal activities almost all C. albicans isolates tested. More importantly, to some Fluconazole (FCZ)-resistant C. albicans, the action of SK minimum inhibitory concentration (MIC 80 value 4 µg/mL) was shown to be >16 times higher than that of FCZ (MIC 80 >64 µg/mL). And the antifungal properties may relate to SKmediated oxidative damage and breakdown of mitochondrial membrane potential. Antifungal Susceptibility Testing Assays were performed on 10 isolates of C. albicans according to methods of the CLSI (formerly NCCLS) (M27-A). MATERIALS AND METHODS Strains and Agents9-11) The initial concentration of the fungal sus...

Reactive Oxygen Species-Inducing Antifungal Agents and Their Activity Against Fungal Biofilms

“…ROS accumulation and eventually apoptosis and/or necrosis in fungal pathogens has been demonstrated for several chemical compounds isolated from plants, such as shikonin from roots of Lithospermum erythrorhizon [76], medioresinol from stem bark of Sambucus williamsii [77], styraxjaponoside C from stem bark of Styrax japonica [78], amentoflavone from Selaginella tamariscina [79], dill seed essential oil (Anethum graveolens) [80,81], the polyphenol curcumin [82], plagiochin E from liverwort Marchantia polymorpha [83,84] and baicalein from Scutellaria baicaleinsis roots [85]. Moreover, the plant defensins HsAFP1 (Heuchera sanguinea), RsAFP2 (Raphinus sativus) and PvD1 (Phaseolus vulgaris), active against several yeast and fungal species were found to induce ROS accumulation in C. albicans and Fusarium oxysporum [86][87][88].…”

Simultaneous Addition of Shikonin and Its Derivatives with Lipopolysaccharide Induces Rapid Macrophage Death

“…These compounds have been shown to exert anti-inflammatory, anti-microbial and anti-tumor effects. 6,8,11,21) In this study, the effect of SHK derivatives on LPS-treated macrophages was examined by using a JA-4 cell line, an LPS-sensitive subline of a murine macrophage-like cell line, J774.1, and by using murine peritoneal primary macrophages. Induction of cytotoxic effects of SHK derivatives at lower concentrations on the macrophages was observed in the presence of LPS (Figs.…”

“…1A), one of the active components of Lithospermi radix (LR), which is a traditional herbal medicine made from the dried root of Lithospermum erythrorhizon SIEB. et ZUCC, has been reported to produce multiple pharmacological effects including antioxidant, 5) antitumor, 6) antifungal, 7) antimicrobial, 8) antiviral, 9) and anti-obesity 10) effects. Some of the latest research has focused on SHK's beneficial effect in anti-inflammatory treatment.…”

Potential Targets for Antifungal Drug Discovery Based on Growth and Virulence in Candida albicans

“…The exact role of ROS in fungal pathogenicity and biofilm development is complex and remains unclear. However, compounds disturbing ROS homeostasis, such as shikonin, show powerful action against C. albicans, which suggests the important role of ROS homeostasis in the pathogenicity of C. albicans (91).…”