FAN Wenting,ZHONG Shimin,HU Qi,et al.Tryptophan metabolites regulate Th17/Treg differentiation to alleviate airway inflammation in asthmatic mice receiving allergen-specific immunotherapy[J].J Third Mil Med Univ,2018,40(08):658-665.

色氨酸代谢物调控Th17/Treg分化在小鼠哮喘变应原特异性免疫治疗中的作用及机制研究(/HTML )




Tryptophan metabolites regulate Th17/Treg differentiation to alleviate airway inflammation in asthmatic mice receiving allergen-specific immunotherapy
FAN Wenting ZHONG Shimin HU Qi ZENG Jing LIAO Wei WANG Jinyu  

Department of Pediatrics, First Affiliated Hospital,  Army Medical University (Third Military Medical University), Chongqing, 400038, China

tryptophan metabolites mouse models bronchial asthma allergenspecific immunotherapy Th17/Treg

目的    对色氨酸代谢物调控Th17/Treg分化在小鼠哮喘变应原特异性免疫治疗中的作用及机制进行研究。方法     将30只BALB/c 鼠随机法分为5组:对照组、哮喘组、OVASIT组(OVA:鸡卵白蛋白;SIT:变应原特异性免疫治疗)、OVASIT+1-MT组(1-MT:IDO抑制剂,1-甲基色氨酸),OVASIT+1MT+KYN组(KYN:色氨酸代谢产物,犬尿氨酸)。哮喘组:第0、7 d予OVA致敏,第6周每天予1% OVA雾化激发,50 d予10% OVA 加强激发;OVASIT组:第4周每天予大剂量OVA皮下注射,余同哮喘组;OVASIT+1MT组:第4周每天在腹腔内注入1-MT,1 h后予大剂量OVA皮下注射h,余同哮喘组;OVASIT+1-MT+KYN组:第3周每天加入1-MT,第4周每天加入 KYN,末次加入KYN后1 h予大剂量OVA行免疫治疗,余同哮喘组。末次激发6 h内检测气道高反应性;对支气管肺泡灌洗液(BALF )进行细胞计数分析;ELISA检测血清IgE及BALF中IL-5、IL-10、IL-17;流式细胞技术检测脾脏CD4+RORγt+T及CD4+Foxp3+T 细胞分化情况。结果     OVA-SIT+1-MT+KYN组较OVA-SIT+1-MT组酸性粒细胞浸润减少,炎症反应明显减轻。前者BALF中Il5为74.8~86.8(83.48±6.02)pg/mL,IL-17为33.8~46.5(38.72±4.61)pg/mL,CD4+RORγt+T 细胞为2.45~2.82(2.60±0.14)%,CD4+Foxp3+T 细胞为7.83~9.09(8.36±0.53)%;后者BALF中Il-5为240.3~285.1(259.65±16.27)pg/mL, IL-17为55.2~65.8(59.97±3.76)pg/mL,CD4+RORγt+T 细胞为4.31~5.34(4.94±0.38)%,CD4+Foxp3+T 细胞为5.93~6.59(6.33±0.28)%,因此OVASIT+1-MT+KYN组中Il-5、IL-17细胞因子以及脾脏中CD4+RORγt+T 细胞均明显低于OVA-SIT+1-MT组,差异均有统计学意义(P<0.01),而前组脾脏中CD4+Foxp3+T 细胞水平明显高于后者,差异有统计学意义(P<0.01)。结论    色氨酸代谢产物有助于特异性免疫治疗减轻气道炎症作用,其机制与通过调控Th17及Treg分化有关。


Objective    To study the role of tryptophan metabolites in regulating Th17/Treg differentiation to alleviate airway inflammation in asthmatic mice receiving allergen-specific immunotherapy (SIT). Methods    Thirty BALB/c mice were randomized equally into control group, asthma group, chicken ovalbumin (OVA)SIT group, OVA-SIT+1-methyltryptophan (1-MT, an indoleamine 2,3 dioxygenase inhibitor) group, and OVASIT+1-MT+kynurenine (KYN, a tryptophan metabolite) group. Except for those in the control group, all the mice were sensitized with OVA on days 0 and 7 followed by daily challenge with 1% OVA in the 6th week and then with 10% OVA on day 50 to induce asthma; In OVA-SIT group, the mice were treated in the 4th week with daily subcutaneous injection of high-dose OVA, and in OVA-SIT+1-MT group, the mice received daily intraperitoneal injection of 1-MT 1 h before subcutaneous high-dose OVA injection; in OVA-SIT+1-MT+KYN group, the mice were treated with daily 1-MT injection in the third week and daily KYN injection in the 4th week, and high-dose OVA immunotherapy was administered at 1 h after the last KYN injection. Airway hyperresponsiveness of the mice was measured within 6 h after the last challenge. Serum levels of IgE and levels of IL-10, IL-5 and IL-17 in bronchoalveolar lavage fluid (BALF) were detected with ELISA, and flow cytometry was performed to analyze the differentiation of CD4+RORγt+and CD4+Foxp3+T cells in the spleen. Results     Compared with those in OVA-SIT+1-MT group, the infiltration of acidic granulocytes and neutrophils decreased and the inflammatory response was significantly reduced in OVA-SIT+1-MT+KYN group. In OVASIT+1-MT group, the mean IL-5 level in BALF was (83.48±6.22) pg/mL, IL-17 level was (38.72±4.61) pg/mL, CD4+Foxp3+T cell percentage was (2.60±0.14)%, and CD4+Foxp3+T cell percentage was (8.36±0.53)%, as compared with the levels of (259.65±16.27) pg/mL, (59.97±3.76) pg/mL, (4.94±0.38)%, and (6.33±0.28)% in OVASIT+1-MT+KYN group, respectively; IL5 and IL-17 levels and spleen CD4+RORγt+T cell percentage was significantly lower (P<0.01) while CD4+Foxp3+T cell percentage was significantly higher  (P<0.01) in OVA-SIT+1-MT+KYN group than in OVA-SIT+1-MT group. Conclusion     Tryptophan metabolites contribute to the efficacy of specific immunotherapy in reducing airway inflammation in asthmatic mice, the mechanism of which may involve the regulation of Th17/Treg cell differentiation.


[1]SIMPSON J L, SCOTT R, BOYLE M J, et al. Inflammatory subtypes in asthma: assessment and identification using induced sputum[J]. Respirology, 2006, 11(1): 54-61. DOI: 10.1111/j.14401843.2006.00784.x.
[2]AKDIS C A, AKDIS M. Mechanisms of allergenspecific immunotherapy and immune tolerance to allergens[J]. World Allergy Organiz J, 2015, 8(1): 1-12. DOI: 10.1186/s40413-015-0063-2.
[3]WILSON R H, WHITEHEAD G S, NAKANO H, et al. Allergic sensitization through the airway primes Th17-dependent neutrophilia and airway hyperresponsiveness[J]. Am J Respir Crit Care Med,2009, 180(8): 720-730. DOI: 10.1164/rccm.2009040573OC.
[4]CHOY D F, HART K M, BORTHWICK L A, et al. TH2 and TH17 inflammatory pathways are reciprocally regulated in asthma[J]. Sci Transl Med, 2015, 7(301): 301ra129. DOI: 10.1126/scitranslmed.aab3142.
[5]ZHAO J, LLOYD C M, NOBLE A. Th17 responses in chronic allergic airway inflammation abrogateregulatory T-cell-mediated tolerance and contribute to airway remodeling[J]. Mucosal Immunol, 2013, 6(2): 335-346. DOI: 10.1038/mi.2012.76.
[6]BETTELLI E, CARRIER Y, GAO W, et al. Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells[J]. Nature, 2006, 441(7090): 235-238.DOI: 10.1038/nature04753.
[7]BABAN B, CHANDLER P R, SHARMA M D, et al. IDO activates regulatory T cells and blocks their conversion into Th17-like T cells[J]. J Immunol, 2009, 183(4): 2475-2483. DOI: 10.4049/jimmunol.0900986.
[8]SINGH N, DALAL V, KUMAR P. Structure based mimicking of Phthalic acid esters (PAEs) and inhibition of hACMSD, an important enzyme of the tryptophan kynurenine metabolism pathway[J]. Int J Biol Macromol, 2017, 108: 214-224. DOI: 10.1016/j.ijbiomac.2017.12.005.
[9]VAN DER SLUIJS K F, VAN DE POL M A, KULIK W, et al. Systemic tryptophan and kynurenine catabolite levels relate to severity of rhinovirus-induced asthma exacerbation: a prospective study with a parallelgroup design[J]. Thorax, 2013(12): 1122-1130. DOI: 10.1136/thoraxjnl-2013-203728.
[10]ODEMUYIWA S O, EBELING C, DUTA V, et al. Tryptophan catabolites regulate mucosal sensitization to ovalbumin in respiratory airways[J]. Allergy, 2009, 64(3): 488-492. DOI: 10.1111/j.13989995.2008.01809.x.
[11]BANSI J, KOLIAMITRA C, BLOCH W, et al. Persons with secondary progressive and relapsingremitting multiple sclerosis reveal different responses of tryptophan metabolism to acute endurance exercise and training[J]. J Neuroimmunol, 2018, 314: 101-105. DOI: 10.1016/j.jneuroim.2017.12.001.
[12]RAITALA A, KARJALAINEN J, OJA S S, et al. Indoleamine 2,3-dioxygenase (IDO) activity is lower in atopic than in nonatopic individuals and is enhanced by environmental factors protecting from atopy[J]. Mol Immunol, 2006, 43(7): 1054-1056. DOI: 10.1016/j.molimm.2005.06.022.
[13]SCHRCKSNADEL K, WIRLEITNER B, WINKLER C, et al. Monitoring tryptophan metabolism inchronic immune activation[J]. Clin Chim Acta, 2006, 364(1-2): 82-90. DOI: 10.1016/j.cca.2005.06.013.
[14]HAYASHI T, BECK L, ROSSETTO C, et al. Inhibition of experimental asthma by indoleamine 2,3dioxygenase[J]. J Clin Invest, 2004, 114(2): 270-279. DOI: 10.1172/JCI21275.
[15]HAYASHI T, MO J H, GONG X, et al. 3Hydroxyanthranilic acid inhibits PDK1 activation and suppresses experimental asthma by inducing T cell apoptosis[J]. Proc Natl Acad Sci USA, 2007, 104(47): 1861918624. DOI: 10.1073/pnas.0709261104.
[16]YAN Y, ZHANG G X, GRAN B, et al. IDO upregulates regulatory T cells via tryptophan catabolite and suppresses encephalitogenic T cell responses in experimental autoimmune encephalomyelitis[J]. J Immunol,2010, 185(10): 5953-5961. DOI: 10.4049/jimmunol.1001628.
[17]SHARMA M D, BABAN B, CHANDLER P, et al. Plasmacytoid dendritic cells from mouse tumordraining lymph nodes directly activate mature Tregs via indoleamine 2,3-dioxygenase[J]. J Clin Invest, 2007,117(9): 2570-2582. DOI: 10.1172/JCI31911.
[18]TAHER Y A, PIAVAUX B J, GRAS R, et al. Indoleamine 2,3-dioxygenase-dependent tryptophan metabolites contribute to tolerance induction during allergen immunotherapy in a mouse model[J]. J Allergy Clin Immunol, 2008, 121(4): 983-991. DOI: 10.1016/j.jaci.2007.11.021.
[19]胡琦, 钟世民, 汪金玉, 等. 皮下注射大剂量卵白蛋白诱导小鼠哮喘免疫耐受模型的建立及机制初步研究[J]. 第三军医大学学报, 2016, 38(12): 1404-1409. DOI: 10.16016/j.10005404.201602024.
HU Q, ZHONG S M, WANG J Y, et al. Establishment of mouse model of immune tolerance of asthma through subcutaneous inject-on of high dose of ovalbumin[J]. J Third Mil Med Univ, 2016, 38(12): 1404-1409. DOI: 10.16016/j.10005404.201602024.
[20]HELLINGS P W, KASRAN A, LIU Z, et al. Interleukin-17 orchestrates the granulocyte influx into airways after allergen inhalation in a mouse model of allergic asthma[J]. Am J Respir Cell Mol Biol, 2003, 28(1): 42-50. DOI: 10.1165/rcmb.4832.
[21]YEH Y C, YEN H R, JIANG R S, et al. Dose-response relationship of specific allergen exposureinduced immunological tolerance: a mouse model[J]. Int Forum Allergy Rhinol, 2015, 5(9): 784-793. DOI: 10.1002/alr.21551.
[22]MEHLHOP P D, VAN DE RIJN M, GOLDBERG AB, et al. Allergen-induced bronchial hyperreactivity and eosinophilic inflammation occur in the absence of IgE in a mouse model of asthma[J]. Proc Natl AcadSci USA, 1997, 94(4): 1344-1349. DOI: 10.1073/pnas.94.4.1344.
[23]MAAZI H, SHIRINBAK S, WILLART M, et al. Contribution of regulatory T cells to alleviation of experimental allergic asthma after specific immunotherapy[J]. Clin Exp Allergy, 2012, 42(10): 1519-1528. DOI: 10.1111/j.13652222.2012.04064.x.
[24]B-HM L, MAXEINER J, MEYERMARTIN H, et al. IL-10 and regulatory T cells cooperate in allergenspecific immunotherapy to ameliorate allergic asthma[J]. J Immunol, 2015, 194(3): 887-897. DOI: 10.4049/jimmunol.1401612.


[1]杜晓兰,陈志,尹良军,等.欧几里德几何距离矩阵法对FGFR2 Ser252Trp点突变小鼠头颅形状特征的分析[J].第三军医大学学报,2009,31(08):655.
 DU Xiao-lan,CHEN Zhi,YIN Liang-jun,et al.Skull morphology of mice carrying gain-of-function mutation of FGFR2 Ser252Trp by Euclidean distance matrix analysis[J].J Third Mil Med Univ,2009,31(08):655.
 DU Xiao-lan,CHEN Zhi,YIN Liang-jun,et al.Quantitative analysis on developmental morphologic changes on mandibles of Apert syndrome mice[J].J Third Mil Med Univ,2009,31(08):1625.
 LI Lan-shuang,HU Li-na.Model establishment of cervical cancer lymphatic metastasis and clinical significance of Prox-1[J].J Third Mil Med Univ,2009,31(08):1585.
 Hu Qi,Zhong Shimin,Wang Jinyu,et al.Establishment of mouse model of immune tolerance of asthma through subcutaneous injection of high dose of ovalbumin[J].J Third Mil Med Univ,2016,38(08):1404.
 Yang Zifeng,Yang Chunguang,Wang Yutao,et al.Screening of mouse-lung adapted strain from clinical influenza B viruses[J].J Third Mil Med Univ,2014,36(08):446.

更新日期/Last Update: 2018-04-27