Effects Of Suloctidil On Biofilm Formation And Growth Of The Candida Albicans

C.albicans is an opportunistic pathogen,according to the Centers for Disease Control,oral candidiasis develops in approximately 7%of infants,31%of AIDS patients,and 20%of cancer patients receiving chemotherapy.Vulvovaginal candidiasis is more common and is estimated to occur more than once in many healthy women.Invasive candidiasis has an impossibly high mortality of 40%-60%.Because C.albicans adapts to various environmental conditions by developing biofilms,which is hardly penetrated by traditional antibiotics and finally prevent fungus from being cleared.The formation of C.albicans biofilm includes the adhesion,initial colonization,production of extracellular material and maturation phases.Biofilm formation is an important virulence factor of C.alicans and occurs in host tissues,prostheses,and indwelling medical devices,such as urinary and vascular catheters,?dental?implants,and heart valves.The biofilm significantly increases the resistance of C.albicans to traditional antifungal drugs,biofilm-associated C.albicans is 4000 times more resistant to fluconazole than its planktonic form.Hence,alternatives to conventional antifungal agents are urgently needed to destroy C.albicans biofilms.Suloctidil?1-?4-isopropylthiophenyl?-2-n-octylaminopropanol?is a calcium-channel blocker,and a vascular antispasmodic and antithrombotic drug with in vitro antifungal properties against C.albicans.The current study found that suloctidil has antifungal activity and synergy with FLU against C.neoformans.However,the antifungal activity of suloctidil against C.albicans biofilm has not been studied.The present study aims to explore the inhibitory effect of suloctidil on the formation of C.albicans biofilm and on preformed biofilm to evaluate its potential therapeutic application in biofilm-associated candidiasis.ObjectiveIn our study,we explored the inhibitory effect of suloctidil on the C.albicans biofilm,evaluated inhibitory effect of suloctidil and relevant mechanisms on C.albicans biofilm in vitro,evaluated therapeutic effect of suloctidil on vaginal candidiasis,in order to provide an improving strategy for superficial biofilm-associated candidal infections.Methods1.The minimum inhibitory concentration(MIC₈₀),biofilm inhibiting concentration(BIC₈₀),biofilm-eradicating concentration(BEC₈₀)of suloctidil to wild type strains of C.albicans YEM30,a clinical isolate of C.albicans LC3.2.The kinetics of the inhibitory effect of suloctidil against C.albicans biofilm formation and preformed biofilm was investigated by the time-kill test.3.The morphological changes of C.albicans biofilm treated by suloctidil were observed by scanning electron microscope?SEM?.4.The ultrastructure changes of C.albicans biofilm treated by suloctidil were observed by transmission electron microscope?TEM?.5.The yeast-to-hypha morphological transition of C.albicans biofilm caused by suloctidil on different adding time was examined by inverted microscope.6.The effect of suloctidil on the expression of hypha-specific genes HWP1,ALS3,and ECE1 was evaluated by quantitative real-time reverse transcription PCR?qRT-PCR?.7.The antifungal activity of suloctidil against C.albicans in vivo was evaluated by murine vaginal candidiasis model.Results1.Suloctidil inhibited the initial colonization of C.albicans and suppressed the maturation of biofilm.The MIC₈₀ of YEM30 and LC30 treated by suloctidil was 4?g/mL,the BIC₈₀ was 16 ?g/mL,The BEC₈₀ was 64 ?g/mL.2.The inhibitory effect of sulodil on the biofilm cells of C.albicans was concentration-dependent.With the prolongation of time,the higher the concentration,the better the inhibitory effect.3.The C.albicans biofilm cells shriveled exposed to suloctidil,with the extension of time,cells were completely shriveled and budding cells had fallen off the hyphae.4.In vitro,the ultrastructure of C.albicans biofilm cells destroyed exposed to suloctidil,the cell membrane shriveled,cell wall fractured,intracellular electron density decreased,cell contents oozed.5.Suoctidil effectively prevented the transformation of C.albicans from yeast to hyphae.The earlier the adding times of suloctidil,the more obvious the effect.6.The expression levels of hypha-specific genes,HWP1,ALS3,andECEl in suloctidil treated cells were significantly reduced.7.Animal experiments showed the reduction in the C.albicans load of mice treated with suloctidil compared with control,suloctidil is effective in the treatment of vaginal candidiasis.ConclusionWe first found the antifungal effect of suloctidil against C.albicans biofilm in vitro and in vivo.Suloctidil can not only broke the morphology and ultrastructure of mature biofilm,inhibited the formation of C.albicans biofilms by down-regulating the expression of HWP1,ALS3,and ECE1 to prevent the morphological transition from yeast to hypha.Therefore,our results revealed that suloctidil is a potential treatment for superficial biofilm-associated candidal infections.