The Effect Of Surfactant Protein D Combined With Voriconazole On Invasive Pulmonary Aspergillosis In Immunosuppressive Mice

ObjectiveAspergillus fumigatus is a ubiquitous saprophytic fungi which can causes a variety of clinical syndromes depend on different immune status.In the immunocompromised individuals A.fumigatus often cause fatal invasive pulmonary aspergillosis.Although the development of diagnostic methods and treatments,IPA morbidity and mortality are still high.Over the past few decades,triazole antifungal-resistant Aspergillus strains have emerged in the world,which may lead to the reduction of treatment options and the failure of treatment.It prompted us to find new treatment methods.Immunocompetence individuals inhale a large number of spores daily without disease,indicating that the host immune function play an important role in resistance to aspergillus.Pulmonary-associated surfactant protein D(SP-D)has shown important role in enhancing host immune defense against invading pathogens,which may help cure IPA.In this study,we examined the therapeutic effect of voriconazole in combination with SP-D versus voriconazole alone,to further study the immune mechanism of SP-D to enhance the efficacy of voriconazole in mice IPA model.MethodsTreatment effect was evaluated on day 4 and day 15 in terms of animal survival rate,pulmonary fungal load,cytokine level,pathological changes and T cell differentiation in spleen.Immunosuppressive mice were established by cyclophosphamide,and then infected Aspergillus fumigatus by nasal administration.The mouse IPA model was successfully established and validated.In order to determine the dose of voriconazole and SP-D,we observe the 15-day survival rates of IPA mice treated with different concentrations of voriconazole and SP-D.The animals were treated with voriconazole(load dose 30 mg/kg/d,maintain dose 15 mg/kg/d)or SP-D(10 μg/d)alone,or voriconazole plus SP-D on day 0,day 1 and day2.Treatment effect was evaluated on day 4 in terms of pulmonary fungal load,cytokine level,pathological changes,T cell differentiation in spleen,and on day 5 in terms of animal survival rate.ResultsPAS staining showed less budding hyphal growth and inflammatory cell infiltration in combined treatment group than in voriconazole or SP-D alone group and IPA group.Compared with alone group,combined treatment group significantly reduced fungal load in lung tissues(P<0.01).Compared with voriconazole alone group and IPA group,combined treatment group significantly increased the level of IFN-γ(463±31.5pg/ml,297.9±16.1pg/ml,552.9±28.4 pg/ml,P<0.01),decreased the level of IL-4(129.3±15.2 pg/ml,200±16.5 pg/ml,83.8±10.9 pg/ml,P<0.01)and the level of IL-17(164.9±21.6 pg/ml,203.9±50.2 pg/ml,97.8±6.6pg/ml,P<0.01).Compared with voriconazole alone group,SP-D alone group and IPA group,combined treatment group significantly increased the expression level of TH1(5.65±0.41%,6.86±0.75%,4.2±0.21%,9.87±1.1%,P<0.01),decreased the expression level of TH2(7.01±0.58%,6.10±0.61%,8.64±0.18%,4.82±0.27%,P<0.01)and decreased the expression level of TH17(1.63±0.11%,2.23±0.25%,2.17±0.24%,3.02±0.24%,P<0.01).The result also showed that survival rate in combination group was significantly higher than that of other groups on day 15(combination group 80%;voriconazole group 40%;SP-D group 30%;IPA group 10%).ConclusionThe combination treatment group showed better balance between the response regulator Th1,Th2 and TH17 cells compared with the other treatment groups.It is concluded from these findings that the pulmonary immunomodulator SP-D can boost the antifungal effect of voriconazole by improving immunity in mice IPA model,which provides new insight into IPA therapy.