title: Neutralization of Interleukin-9 Decreasing Mast Cells Infiltration in Experimental Autoimmune Encephalomyelitis.
abstract: BACKGROUND
  Th9 cells are a newly discovered CD4+ T helper cell subtype, characterized by high interleukin (IL)-9 secretion. Growing evidences suggest that Th9 cells are involved in the pathogenic mechanism of multiple sclerosis (MS). Mast cells are multifunctional innate immune cells, which are perhaps best known for their role as dominant effector cells in allergies and asthma. Several lines of evidence point to an important role for mast cells in MS and its animal models. Simultaneously, there is dynamic "cross-talk" between Th9 and mast cells. The aim of the present study was to examine the IL-9-mast cell axis in experimental autoimmune encephalomyelitis (EAE) and determine its interaction after neutralizing anti-IL-9 antibody treatment.
  
  
  METHODS
  Female C57BL/6 mice were randomly divided into three groups (n = 5 in each group): mice with myelin oligodendrocyte glycoprotein (MOG)-induced EAE (EAE group), EAE mice treated with anti-IL-9 antibody (anti-IL-9 Abs group), and EAE mice treated with IgG isotype control (IgG group). EAE clinical score was evaluated. Mast cells from central nervous system (CNS) were detected by flow cytometry. The production of chemokine recruiting mast cells in the CNS was explored by reverse transcription-polymerase chain reaction (RT-PCR). In mice with MOG-induced EAE, the expression of IL-9 receptor (IL-9R) complexes in CNS and spleen mast cells was also explored by RT-PCR, and then was repeating validated by immunocytochemistry. In vitro, spleen cells from EAE mice were cultured with anti-IL-9 antibody, and quantity of mast cells was counted by flow cytometry after co-culture.
  
  
  RESULTS
  Compared with IgG group, IL-9 blockade delayed clinical disease onset and ameliorated EAE severity (t = -2.217, P = 0.031), accompany with mast cells infiltration decreases (day 5: t = -8.005, P < 0.001; day 15: t = -11.857, P < 0.001; day 20: t = -5.243, P = 0.001) in anti-IL-9 Abs group. The messenger RNA expressions of C-C motif chemokine ligand 5 (t = -5.932, P = 0.003) and vascular cell adhesion molecule-1 (t = -4.029, P = 0.004) were significantly decreased after IL-9 neutralization in anti-IL-9 Abs group, compared with IgG group. In MOG-induced EAE, the IL-9R complexes were expressed in CNS and spleen mast cells. In vitro, splenocytes cultured with anti-IL-9 antibody showed significantly lower levels of mast cells in a dose-dependent manner, compared with splenocytes cultured with anti-mouse IgG (5 μg/ml: t = -0.894, P = 0.397; 10 μg/ml: t = -3.348, P = 0.019; 20 μg/ml: t = -7.639, P < 0.001).
  
  
  CONCLUSIONS
  This study revealed that IL-9 neutralization reduced mast cell infiltration in CNS and ameliorated EAE, which might be relate to the interaction between IL-9 and mast cells.
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Jun-Jie
authors.initials: JJ
authors.lastname: Yin
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Xue-Qiang
authors.initials: XQ
authors.lastname: Hu
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Zhi-Feng
authors.initials: ZF
authors.lastname: Mao
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Jian
authors.initials: J
authors.lastname: Bao
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Qiu
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Zheng-Qi
authors.initials: ZQ
authors.lastname: Lu
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Hao-Tian
authors.initials: HT
authors.lastname: Wu
authors.affiliation: Department of Neurology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, China.
authors.forename: Xiao-Nan
authors.initials: XN
authors.lastname: Zhong
mesh_terms: D000818:Animals; D000906:Antibodies / Q000627:therapeutic use*; D002490:Central Nervous System / Q000378:metabolism; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000188:drug therapy* / Q000378:metabolism*; D005260:Female; D007150:Immunohistochemistry; D016906:Interleukin-9 / Q000037:antagonists & inhibitors* / Q000276:immunology / Q000378:metabolism*; D008407:Mast Cells / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000906:Antibodies; D016906:Interleukin-9; D012333:RNA, Messenger