title: TGFβ signalling plays an important role in IL4-induced alternative activation of microglia.
abstract: BACKGROUND
  Microglia are the resident immune cells of the central nervous system and are accepted to be involved in a variety of neurodegenerative diseases. Several studies have demonstrated that microglia, like peripheral macrophages, exhibit two entirely different functional activation states, referred to as classical (M1) and alternative (M2) activation. TGFβ is one of the most important anti-inflammatory cytokines and its effect on inhibiting microglia or macrophage classical activation has been extensively studied. However, the role of TGFβ during alternative activation of microglia has not been described yet.
  
  
  METHODS
  To investigate the role of TGFβ in IL4-induced microglia alternative activation, both, BV2 as well as primary microglia from new born C57BL/6 mice were used. Quantitative RT-PCR and western blots were performed to detect mRNA and protein levels of the alternative activation markers Arginase1 (Arg1) and Chitinase 3-like 3 (Ym1) after treatment with IL4, TGFβ or both. Endogenous TGFβ release after IL4 treatment was evaluated using the mink lung epithelial cell (MLEC) assay and a direct TGFβ2 ELISA. TGFβ receptor type I inhibitor and MAPK inhibitor were applied to address the involvement of TGFβ signalling and MAPK signalling in IL4-induced alternative activation of microglia.
  
  
  RESULTS
  TGFβ enhances IL4-induced microglia alternative activation by strongly increasing the expression of Arg1 and Ym1. This synergistic effect on Arg1 induction is almost completely blocked by the application of the MAPK inhibitor, PD98059. Further, treatment of primary microglia with IL4 increased the expression and secretion of TGFβ2, suggesting an involvement of endogenous TGFβ in IL4-mediated microglia activation process. Moreover, IL4-mediated induction of Arg1 and Ym1 is impaired after blocking the TGFβ receptor I indicating that IL4-induced microglia alternative activation is dependent on active TGFβ signalling. Interestingly, treatment of primary microglia with TGFβ alone results in up regulation of the IL4 receptor alpha, indicating that TGFβ increases the sensitivity of microglia for IL4 signals.
  
  
  CONCLUSIONS
  Taken together, our data reveal a new role for TGFβ during IL4-induced alternative activation of microglia and consolidate the essential functions of TGFβ as an anti-inflammatory molecule and immunoregulatory factor for microglia.
authors.affiliation: Institute for Anatomy and Cell Biology, Department of Molecular Embryology, Albert-Ludwigs-University Freiburg, Albertstraße 17, Freiburg 79104, Germany.
authors.forename: Xiaolai
authors.initials: X
authors.lastname: Zhou
authors.forename: Björn
authors.initials: B
authors.lastname: Spittau
authors.forename: Kerstin
authors.initials: K
authors.lastname: Krieglstein
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D001119:Arginase / Q000235:genetics / Q000378:metabolism; D001921:Brain / Q000166:cytology; D002478:Cells, Cultured; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D015847:Interleukin-4 / Q000494:pharmacology*; D037102:Lectins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000187:drug effects*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects*; D001619:beta-N-Acetylhexosaminidases / Q000235:genetics / Q000378:metabolism; D053662:Interleukin-4 Receptor alpha Subunit
chemical_list: D001119:Arginase; D016207:Cytokines; D004791:Enzyme Inhibitors; D015847:Interleukin-4; D037102:Lectins; D020928:Mitogen-Activated Protein Kinases; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D016212:Transforming Growth Factor beta; D001619:beta-N-Acetylhexosaminidases; C504002:Il4ra protein, mouse; D053662:Interleukin-4 Receptor alpha Subunit; C429541:Chil3 protein, mouse; C544448:Arg1 protein, mouse