Study On Antifungal Activity And Mechanism Of Fufang Xianhecao Combined With Fluconazole On Candida Albicans

Background: With the increasing number of diseases such as cancer,diabetes,the use of broad-spectrum antibiotics and immunosuppressive agents,systemic fungal infections and fungal drug resistance are becoming increasingly prominent and severe.Studies have shown that some Chinese medicines have a synergistic effect against drug-resistant fungi.Compound Xianhecao capsules(FFXHC)have the effects of strengthening the spleen Qi and clearing away heat and dampness,which are similar to the pathogenesis of Candida albicans according to the theory of traditional Chinese medicine.We found that the use of FFXHC combined with fluconazole(FLC)that is one of antifungal drugs has a good prospect for the treatment of fungal infections.Objective: The purpose of this study was to evaluate the antifungal effects of FFXHC combined with FLC on drug-resistant Candida albicans in vivo and in vitro,and the mechanism of the synergistic effects.Method: The position of the Fufangxianhecao combined with fluconazole against Candida albicans was measured by the microdilution method.The fractional inhibitory concentration index was used to evaluate the synergy effect of the two drugs.The time-bactericidal curve method was used to dynamically study the inhibition of the combined use of capsules and fluconazole on drug-resistant strains;Induction of mycelial growth with different media was also observed;The mouse Candida albicans infection model was used to evaluate the antifungal effects of FFXHC and FLC in vivo,The mice were randomly divided into control group(NC)and model Group(Model),Compound Agrimony combined with fluconazole administration group(FFXHC + FLC.H,FFXHC + FLC.M,FFXHC + FLC.L)and berberine combined with fluconazole administration group(BBR + FLC).The changes in the survival rate,body weight,lung and kidney colony number,and immune organ coefficient of mice in different administration groups were observed to determine the effect of FFXHC combined with FLC on Candida albicans infection.The fluorescent probe rhodamine 6G was used to determine the effect of FFXHC and FLC on Candida albicans drug transporter,and the expression of drug resistance genes is determined by RT-PCR.The fluorescent probes H2 DCFDA and JC-1 were used to determine the effects of FFXHC and FLC on the intracellular reactive oxygen species and mitochondrial membrane potential of Candida albicans,and the change of ATP content was also determined.Proteomics studies were used to detect differential protein expression before and after the treatment of FFXHC to explore the antifungal mechanism of FFXHC combined with FLC.Result: The combination of FFXHC and FLC has a good synergistic anti-Candida albicans effect.The combination can significantly improve the survival rate and body weight,and reduce tissue lesions of mice.The number of visceral colonies and the visceral coefficient of immune organs were also improved significantly.Compared with the positive controlberberine(BBR),FFXHC has better therapeutic effects and significantly improves the survival status of mice.Mechanism study found that FFXHC combined with FLC can reduce the efflux pump function of FLC-resistant Candida albicans,but it had no significant effect on the expression of drug resistance genes CDR1,CDR2 and MDR1.Further studies showed that FFXHC combined with FLC can significantly reduce the ATP content and the mitochondrial membrane potential and increase the level of intracellular reactive oxygen species in Candida albicans,indicating that it may affect energy metabolism process.Proteomics studies confirmed the assumption that the treatment of FFXHC can affect the expression of key enzymes in energy metabolism pathways such as glycolysis,fructose and mannose in resistant strains.Our results showed that the combination of FFXHC and FLC can reduce the ATP content to inhibit the efflux function of Candida albicans.Conclusion: FFXHC combined with FLC have a significant anti-Candida albicans effect in vivo and in vivo.Mechanism studies have shown that the drug efflux pump activity can be inhibited by inducing ATP content to achieve restore FLC sensitivity and antifungal effect.