Increase of reactive oxygen species contributes to growth inhibition by fluconazole in Cryptococcus neoformans

Abstract

Background: Cryptococcus neoformans, a basidiomycetous yeast, is a fungal pathogen that can colonize the lungs of humans causing pneumonia and fungal meningitis in severely immunocompromised individuals. Recent studies have implied that the antifungal drug fluconazole (FLC) can induce oxidative stress in C. neoformans by increasing the production of reactive oxygen species (ROS), as presence of the antioxidant ascorbic acid (AA) could reverse the inhibitory effects of FLC on C. neoformans. However, in Candida albicans, AA has been shown to stimulate the expression of genes essential for ergosterol biosynthesis. Hence, the contribution of ROS in FLC-mediated growth inhibition remains unclear. Results: In order to determine whether counteracting ROS generated by FLC in C. neoformans can contribute to diminishing inhibitory effects of FLC, we tested three other antioxidants in addition to AA, namely, pyrrolidine dithiocarbamate (PDTC), retinoic acid (RA), and glutathione (GSH). Our data confirm that there is an increase in ROS in the presence of FLC in C. neoformans. Importantly, all four antioxidants reversed FLC-mediated growth inhibition of C. neoformans to various extents. We further verified the involvement of increased ROS in FLC-mediated growth inhibition by determining that ROS-scavenging proteins, metallothioneins (CMT1 and CMT2), contribute to growth recovery by PDTC and AA during treatment with FLC. Conclusion: Our study suggests that ROS contributes to FLC-mediated growth inhibition and points to a complex nature of antioxidant-mediated growth rescue in the presence of FLC.

Effects of environmental factors on sensitivity of Cryptococcus neoformans to fluconazole and amphotericin B

“… 2018 ; Dbouk et al . 2019 ). It has been shown that co-treatment of C. neoformans with FLC and the antioxidants, retinoic acid (RA), ascorbic acid (AA) and pyrrolidine dithiocarbamate (PDTC) prevents the increase of ROS and has a significant effect by lowering the susceptibility towards FLC (Dbouk et al .…”

“…It has been shown that co-treatment of C. neoformans with FLC and the antioxidants, retinoic acid (RA), ascorbic acid (AA) and pyrrolidine dithiocarbamate (PDTC) prevents the increase of ROS and has a significant effect by lowering the susceptibility towards FLC (Dbouk et al . 2019 ). Surprisingly, the effect of yet another classic antioxidant, glutathione (GSH) had a minimal effect on susceptibility towards FLC (Dbouk et al .…”

“…Surprisingly, the effect of yet another classic antioxidant, glutathione (GSH) had a minimal effect on susceptibility towards FLC (Dbouk et al . 2019 ). Given the reciprocal effects of the nutrient and differences in the effect of temperature between FLC and AMB, we wished to probe the effects of the antioxidants on the susceptibility towards these two drugs.…”

“… 2018 ; Dbouk et al . 2019 ). ROS such as hydrogen peroxide, oxides, or hydroxide are highly unstable cytotoxic molecules that generate free radicals which can cause molecular and cellular damage (Auten and Davis 2009 ).…”

“…Interestingly, while treatment of C. neoformans with FLC and the antioxidants retinoic acid (RA), ascorbic acid (AA) and pyrrolidine dithiocarbamate (PDTC), can lower the sensitivity of C. neoformans to FLC, the antioxidant glutathione (GSH) exhibits little to no rescue from FLC inhibition (Dbouk et al . 2019 ). Whether the efficacy of AMB is altered in the presence of various antioxidants has not been determined.…”