Magnolol Inhibits the Growth of Cryptococcus neoformans by Disrupting the GSH Oxidation-Reduction System

In this study, Cryptococcus neoformans BNCC225501 was used as research objects to explore the antifungal activity of magnolol. The molecular mechanisms underlying magnolol against Cryptococcus neoformans BNCC225501 were explored using metabolomics and transcriptomics. The results offer ideas and directions for developing new antifungal drugs based on magnolol.

The minimum inhibitory concentration (MIC) and the method of drawing the growth curve were used to determine whether magnolol had fungistatic and fungicidal effects on Cryptococcus neoformans. Cryptococcus neoformans was cultured on urea basal medium and Niger seed medium, and different concentrations of magnolol were added to these media to detect the effects of magnolol on the urease and melanin of Cryptococcus neoformans. Network pharmacology was used to screen for potential target genes of magnolol and fungal infection, obtaining common targets. Further molecular docking was performed between magnolol and the proteins corresponding to the common targets.

The MIC of the individual tested agents against Cryptococcus neoformans strain was 8 μg/mL for magnolol. The combination of magnolol and fluconazole revealed good synergistic effects against Cryptococcus neoformans (FIC ≤ 0.5). At a magnolol concentration of 8 μg/mL, the expression of urease was inhibited. Compared to the control group, in the experimental group treated with magnolol, among the differential metabolites, lipids and lipid-like molecules accounted for the highest percentage of differential metabolites, except for unclassified metabolites. A total of 30% of the metabolites belonged to lipids and lipid-like molecules, followed by amino acids, peptides, and analogues (22.63%), organoheterocyclic compounds (7.91%), alkaloids (6.17%), sugars (6.01%), nucleosides, nucleotides and analogues (3.80%), organic oxygen compounds (2.22%), aromatic compounds (1.90%), organic acids (1.90%), and other categories (16.93%). KEGG pathway enrichment analysis results showed that the metabolites were mainly enriched in glyoxylate and dicarboxylate metabolism, arginine biosynthesis, valine, leucine, and isoleucine degradation; arginine and proline metabolism; cysteine and methionine metabolism; pentose and glucuronate interconversions; sphingolipid metabolism; histidine metabolism; and the citrate cycle. Compared to the control group, in the experimental group treated with magnolol, a total of 928 genes were detected to be altered. The GO enrichment results showed that the altered genes were enriched in the component of membrane and oxidation-reduction process. KEGG enrichment analysis showed that the differential genes were mainly enriched in the DNA replication, NOD-like receptor signaling pathway, and fatty acid biosynthesis. Combined analysis of metabolomics and transcriptomics demonstrated that the glycerophospholipid metabolism pathway and glutathione metabolism pathway were significantly changed after treatment with magnolol. Screening identified 31 target genes for magnolol and 969 fungal genes, with three intersecting targets, DPP4, MAPK14, and CCNA2. Molecular docking studies showed that magnolol and the target protein had a good binding ability, a docking binding energy of -7.3 kcal/mol.

Magnolol has fungistatic and fungicidal ability for Cryptococcus neoformans and can inhibit the urease production capacity of Cryptococcus neoformans. The antifungal mechanism of magnolol may be that it disrupts redox homeostasis and inhibits detoxification by affecting Glutathione metabolism.