The Mechanism Of Tryptophan Metabolism Regulated By IDO1 In Airway Remodeling Of Allergic Asthma

ObjectiveRecently,as the most common type of asthma clinically,both the incidence and the medical burden of allergic asthma are increasing year by year.In addition,some patients are insensitive to conventional treatment.Therefore,the studies on the mechanism and latent treatment target are meaningful.In recent years,it has been found that tryptophan metabolism is involved in the development of a variety of diseases,including cancers,autoimmune diseases,infectious diseases,and central nervous system diseases.Indoleamine2,3-dioxygenase 1(IDO1),highly expressed in respiratory system,is one of key enzymes that regulates tryptophan metabolism,and its downstream product is kynurenine(Kyn).Previous studies have reported abnormal serum tryptophan metabolism in patients with allergic asthma,which may be linked to the development of asthma,but the specific mechanism have not been illustrated.Additionally,Kyn is the endogenous ligand of transcription factor aromatics receptor(Ah R),inducing Ah R translocated into the nucleus to regulate the expression of key genes.By screening the key genes,we focused on CTH gene,which encodes cystathionine-γ-lyase,the main synthase of endogenous hydrogen sulfide.Meanwhile,endogenous hydrogen sulfide has airway protective effects on asthma.Therefore,this study intends to clarify the level of the tryptophan metabolism and its key enzyme IDO1 in allergic asthma and explore the specific mechanism.Additionally,we are aimed to illustrate the effect and mechanism of tryptophan metabolism abnormalities(especially IDO1)on allergic asthma and make it clear whether IDO1-regulated tryptophan metabolism plays a role in airway remodeling in allergic asthma by affecting the synthesis of endogenous hydrogen sulfide.This study is helpful to gain insight into allergic asthma and can provide clues for the new therapy.Methods1.Serum of patients with allergic asthma and healthy controls were collected,and the concentration of tryptophan and kynurenine in serum were detected by metabolic mass spectrometry;Luminex assays were used to detect the concentrations of Th1 and Th2 cytokine.Asthmatic and control(the distal normal lung tissues from patients with lung cancer)lung tissue samples were collected,and the histopathology of asthmatic lung and the expression of IDO1 were detected by HE staining and immunohistochemical staining of lung tissue sections,respectively.2.The human airway epithelial cell line Beas-2B was stimulated with Th1 and Th2 cytokines,and the expression of IDO1 was detected by q PCR and WB;The changes of tryptophan and its metabolites were detected by targeted tryptophan metabolic mass spectrometry.3.Beas-2B cells were stimulated with Kyn to clarify the subcellular location of Ah R by immunofluorescence and nuclear-cytoplasmic separation experiments;The transcription of Ah R classical regulatory target genes CYP1B1 was detected by q PCR.The Ah R downstream regulatory gene CTH was screened out by CUT&Tag and Ch IP experiments.IFN-γ and Kyn were used in combination with IDO1 inhibitors and Ah R nucleustransportation inhibitors to stimulate BEAS-2B cells,and the expression of CTH was detected by q PCR and WB.4.HDM-induced allergic asthma were established.Serum cytokine levels were detect by Luminex technology;The expression levels of IDO1 and CTH were detected by q PCR,WB and immunohistochemical staining;Inflammatory infiltration and airway remodeling of lung tissue were observed by lung tissue section staining.5.Lung tissue was specifically infected with adeno-associated virus carrying the Ido1 gene.The expression levels of IDO1 and CTH were detected by q PCR,WB,and immunohistochemical staining;Inflammatory infiltration and airway remodeling of lung tissue were observed by lung tissue section staining.6.To interfere the level of hydrogen sulfide in vivo,the mice with HDM-induced allergic asthma were treated with hydrogen sulfide sustained-release agent GYY4137 and CTH inhibitor PAG,and the inflammatory infiltration and airway remodeling of lung tissue were observed by lung tissue section staining.7.To interfere the concentration of hydrogen sulfide in cultural medium,the human bronchial smooth cell line HBSMC were treated with hydrogen sulfide sustained-release agent GYY4137 and CTH inhibitor PAG,and the effect of hydrogen sulfide on airway smooth muscle cell contraction was verified by collagen disc contraction experiment.Results1.Tryptophan metabolism was inhibited in patients with allergic asthma,and the expression of IDO1 in the airway epithelium was weakened in asthmatic patients.2.Th1 cytokine,such as IFN-γ,promoted IDO1 expression,Th2 cytokines,such as IL-4,IL-5 and IL-13,damaged IDO1 expression.3.Th1 cytokines promote kynurenine pathway in airway epithelial cells,while Th2 cytokines inhibit it.Meanwhile,IDO1 induced by IFN-γ-catalyze the process of the metabolism of tryptophan into kynurenine.4.Kyn promoted Ah R nucleus-transportation in airway epithelial cells,which led to the regulation of expression of CTH,which is the endogenous hydrogen sulfide synthetase.5.The expression levels of IDO1 and CTH in the lungs of animal models of allergic asthma were reduced,and increasing the expression of IDO1 in lung tissues could improve airway remodeling.6.Impaired CTH activity aggravated inflammatory infiltration and airway remodeling of allergic asthma in mice,and the supplement of hydrogen sulfide can improve the pathological changes in allergic asthma.7.Inhibition of CTH aggravates airway smooth muscle contraction in vitro.ConclusionTryptophan metabolism is impaired in the presence of allergic asthma,and the mechanism is probably associated with the Th2 inflammation.During the occurrence of allergic asthma,the Th2 inflammation is activated in lung,and the concentration of Th2 cytokines,such as IL-4,IL-5 and IL-13,increases,which inhibits the expression of IDO1 in airway epithelial cells maintained by IFN-γ.Consequently,the tryptophan metabolism is impaired,which further results in the low level of Kyn in cells.In turn,the abnormality of tryptophan metabolism further induces the exacerbation of allergic asthma.Kyn promotes Ah R nucleus-transportation and then induces the expression of endogenous hydrogen sulfide synthase CTH to maintain a certain level of endogenous hydrogen sulfide.In allergic asthma,the level of Kyn is decreased.Consequently,the Kyn-Ah R-CTH pathway is inhibited and the production of endogenous hydrogen sulfide is reduced,which causes increasing inflammatory infiltration,airway hyperresponsiveness,and aggravated airway remodeling.Additionally,activating tryptophan metabolism in lung tissue or elevating the level of exogenous hydrogen sulfide can effectively alleviate the pathological process of asthma.