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The role of microglia in CNS GvHD

Mar 16, 2020

Graft -versus-host disease (GvHD) is a complication observed in patients following allogeneic hematopoietic cell transplantation (allo-HCT). Primarily believed to be restricted to the skin, gastrointestinal tract and liver, pre-clinical and clinical observations have since revealed the effect of GvHD on the central nervous system (CNS). 1

T cell CNS infiltration is a key player in CNS GvHD establishment and progression. As a growing concern in the field of allo-HCT, efforts have been made to determine the mechanisms behind T cell infiltration, neuronal apoptosis and subsequent neurological deficits. 2Microglia are a tissue macrophage population found in the CNS and stand as the main major histocompatibility complex class II (MHC-II)-expressing antigen presenting cell (APC) in the CNS. 3Although microglia are important in mediating CNS immune responses, they have also been linked to the pathogenesis of a number of neurodegenerative diseases. 4A team ledby Mathew R Nimithaand Vinnakota M Janaki, University of Freiburg, Freiburg, DE, investigated microglial activation in CNS GvHD mouse models and patient samples. 2

Study design

  • The bone marrow (BM) of experimental mice was irradiated and GvHD was induced by transplantation of BM cells and CD4 +/CD8 +T cells from a mismatched donor (allo-HCT). As a control, syngeneic stem cell transplantation (syn-HCT) was performed by transplantation of BM and T cells from a donor mouse matching the genetic background of the recipient mouse
  • CNS immune infiltration and activation were measured using:
    • Immunohistochemistry (IHC) and flow cytometric (FC) quantification of CD3 +T cells
    • IHC and FC quantification of CD11b +CD45 highcells (dendritic cells, monocytes, macrophages)
    • IHC quantification of Iba-1, a microglial activation marker
    • IHC and FC quantification of MHC II- and CD80-expressing microglia
    • Transcriptome analysis of Map3k7,which encodes for transforming growth factor beta (TGF-β)-activated kinase 1 (TAK 1)
    • IHC quantification of phosphorylated (p) microglia via the markers: TAK 1, pJNK, pNF-kB, pMAPK
  • The elevated plus maze and novel object recognition tests were implemented to evaluate neurocognitive function in mice
  • GvHD was induced in Cx3cr1 creER: Tnf fl/- mice to investigate the role of tumour necrosis factor (TNF) in CNS tissue damage
  • GvHD was induced in Cx3cr1 creER:Tak1 fl/fl to investigate the role of microglial TAK1 in CNS-GvHD
  • The effect of two TAK1 inhibitors, takinib and 5Z-7-Oxozeaenol, on CNS GvHD-presenting mice was investigated
  • Formalin fixed paraffin embedded (FFPE) patient brain samples ( Institute of Neuropathology, University Hospital Freiburg, Germany) were obtained from patients that had died following allo-HCT and were either asymptomatic or had developed GvHD

Results

  • The abundance of CD3 +T cells was significantly increased in the meninges (p< 0.005) and cortex (p= 0.001) of mice that had undergone allo-HCT but not after syn-HCT
  • The abundance of CD11b +CD45 highcells was significantly greater (p= 0.005) in mice that had undergone allo-HCT compared to syn-HCT
  • The abundance of Iba-1 +microglia cells was increased in mice that had undergone allo-HCT compared to syn-HCT (p= 0.01)
  • Features of microglial activation (reduced filament dendrite length, reduced number of dendrite segments, branching points and dendrite terminal points) were observed in allo-HCT treated, GvHD-developing mice but not in syn-HCT treated or untreated mice
  • Significant upregulation of MHC-II (p< 0.0001) and CD80 (p< 0.0001) on CD11b +CD45 highcells was observed in allo-HCT treated mice compared to syn-HCT treated or untreated mice
  • In mice that developed GvHD, p38 MAPK phosphorylation was increased, indicating activation of the MAPK/NF-κB pathway
  • TNF knockout (KO) mice exhibited reduced number of CD3 +T cells in the CNS
  • NF-κB-mediated TAK1 activation was driven by TNF in mice
  • Microglial TAK1 KO mice exhibited reduced of CD3 +cell in the CNS, reduced microglial MHC-II expression and improved neurocognitive function while maintaining peripheral GvHD presentation
  • Pharmaceutical inhibition of TAK1 in mice resulted in reduced:
    • CNS CD3 +T cells and Iba-1 +microglia
    • CNS Th1 and Th17 cells
    • MHC-II expression by microglia
    • TNF production by microglia
    • Endothelial vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) expression on CNS endothelial cells
  • In-vivopharmaceutical inhibition of TAK1 improved neurocognitive function in mice
  • The number of Iba-1 +microglia was increased in human patients who developed acute GvHD (aGvHD) following allo-HCT compared to those who did not
  • The number of TNF and Iba-1 double positive microglia was significantly increased in patients who developed aGvHD compared to those who did not

Conclusions

  • Microglia play a crucial role in the establishment of CNS GvHD; augmenting T-cell stimulation and migration into the CNS
  • The microglial TAK1/TNF/MHC-II axis is imperative in the initiation and maintenance of CNS GvHD
  • The data from this study provides the grounds for investigating TAK1 inhibition in phase I clinical trials for the treatment of CNS GvHD
  1. Hartrampf S. et al. The central nervous system is a target of acute graft versus host disease in mice. Blood. 2013 March 7; 121(10): 1906–10. DOI: 10.1182/blood-2012-09-456590
  2. Mathew R.N. et al. Graft-versus-host disease of the CNS is mediated by TNF upregulation in microglia. J Clin Invest. 2019 December 17. DOI: 10.1172/JCI130272
  3. Schetters S.T.T. et al. Neuroinflammation: Microglia and T Cells Get Ready to Tango. Front. Immunol. 25 January 2018.DOI: 10.3389/fimmu.2017.01905
  4. Bachiller S. et al. Microglia in Neurological Diseases: A Road Map to Brain-Disease Dependent-Inflammatory Response. Front Cell Neurosci. 2018 December 18. DOI: 10.3389/fncel.2018.00488