Therapeutic Effect And Mechanism Of 5-aminolevulinic Acid Photodynamic Therapy For Majocchi Granuloma

Background: Dermatophytes are a group of filamentous fungi that are pathogens causing dermatophytosis and often cause skin,hair and nail lesions.Dermatophytes can produce enzymes that digest keratin.Fungus serves as a food source.Generally does not invade subcutaneous or deep skin tissue.Changes in the system and local factors can affect the body's resistance to dermatophytes,including systemic immune dysfunction,such as oral glucocorticoids,immunosuppressive agents,and trauma,local radiotherapy,long-term external use of glucocorticoid drugs,etc.Individuals with poor immune function have an increased risk of dermatophytosis.The main method of traditional treatment is the oral treatment with standard antifungal drugs.However,there are shortcomings such as long treatment period,impaired liver and kidney function,and induction of drug-resistant bacteria.Trichophyton belongs to the family of dermatophytes of the trichophyton genus and can cause scalp,smooth skin,or superficial fungal infections.The incidence rate has increased year by year in recent years.Photodynamic therapy(PDT)is a new type of non-invasive treatment.The photosensitizer selectively accumulates in the target cell,and under the irradiation of a specific light source,a cytotoxic substance such as singlet oxygen and free radicals is generated,so that the target cell is killed and has no effect on normal tissues.This topic was inspired by the successful photodynamic treatment of patients with clinical Majocchi granulomatosis by the isolation of clinical strains(confirmed as Trichophyton tonsurans),aimed at verifying the inhibition and treatment of ALA-PDT against Trichophyton tonsurans through in vitro and in vivo experiments.The effect provides further theoretical basis for later application of ALA-PDT to fungal infectious skin diseases.Objectives:(1)To investigate the killing effect of 5-Aminolevulinic acid photodynamic therapy(ALA-PDT)on Trichophyton tonsurans in vitro.(2)To observe the efficacy of 5-aminolevulinic acid photodynamic therapy for deep infection of dermatophytes in guinea pigs caused by Trichophyton tonsurans.Methods:(1)The clinical isolates of trichophyton tonsurans were cultured in potato dextrose agar medium to prepare the suspension.After 6 hours of incubation period,the production of PPIX was observed by fluorescence microscopy.Following PDT(at light energy density of 50,100,150,175,or 200J/cm2),the growth inhibitory effect of Trichophyton tonsurans was observed by colony cultivation.Inhibitory effect and mechanism of transmission electron microscopy(ALA-PDT)on the growth of Trichophyton tonsurans.(2)To establish an animal model of dermatophyte deep infection of guinea pigs by intramuscular injection of triamcinolone acetate and intraperitoneal injection of dexamethasone by inducing immunosuppressed state in guinea pigs,fungal microscopy and culture,pathological validation Is the module successful.Explore the best ALA incubation time.Observing the effect of ALA-PDT on dermatophyte deep infection of guinea pigs,mainly including microscopic examination of culture and skin lesion scores,comparing it with the pure light group,simply applied ALA group.Results:(1)Fluorescence microscopy showed obvious Pp IX production of brick red fluorescent substance.When the light energy density was 175 and 200J/cm2,the photos showed that the growth of colonies were significantly restricted which was evidenced by decreased concentration of suspension.The suspension concentrations were(0.53 ± 0.058)× 105 CFU / ml and(0.13 ± 0.057)× 105 CFU / ml at light energy density of 175 and 200J/cm2 respectively,which was significantly lower than that of control group,single drug group,single light group and other low energy density ALA-PDT group(P <0.05).Transmission electron microscopy: The cell wall of the mycelium section of the control group was intact,thick and thin,and the texture was dense: the cell matrix was uniform,the density was high,the organelles were abundant,the nucleus and lipid droplets were clearly visible in the cytoplasm;the cells in the PDT group were swollen and deformed,and the cells were visible In a vacuolar structure,most organelles collapsed and high electron density aggregates appeared in the cells.(2)Fungal microscopic examination and culture combined with pathological validation model success.Blood stasis was observed on the 4th day after inoculation of guinea pigs,and erythema,desquamation,erosion,and scab were observed on the 7th to 8th day.Positive on the 5th day after inoculation.Microscopic examination revealed hyphae and spores and fungi.Culture is a positive result.Pathological examination revealed hyphae or spores in the dermal follicles.Fluorescence detection showed that within 1-7h,with the extension of time,the fluorescence intensity at 5h reached the peak,and Pp IX began to decline after 6h.Therefore,5h can theoretically be the best incubation time for photodynamic therapy.3.Efficacy evaluation: In the ALA-PDT group on the 8th day after treatment,the skin lesion score on the 11 th day was significantly lower than that of the simple phototherapy group and the pure ALA incubation group.Conclusion:(1)ALA-PDT was effective for Majocchi granuloma.(2)Trichophyton tonsurans can absorb ALA and be converted to Pp IX,and the energy density of ALA-PDT has a dose-dependent effect on its killing effect.(3)Glucocorticoids can assist in the construction of a model of deep skin infections in guinea pigs caused by Trichophyton tonsurans.ALA-PDT can effectively treat deep skin infections in guinea pigs.The therapeutic effect is better than that of pure light and ALA alone.