Regulatory T Cell-microglia Crosstalk Promotes White Matter Repair After Ischemic Stroke Through Osteopontin

Introduction:The precise mechanisms underlying the salutary effects of FOXP3~+regulatory T cells(Treg)on long-term brain tissue repair have remained elusive.Our previous work proved that after cerebral ischemic injury,Treg cells have a long-term brain tissue repair effect,which is partially depended on microglia cells.This study mainly focus on the specific mechanism by which Treg cells interact with microglia to promote white matter repair after ischemic injury.Methods:Treg cells infiltrated into ischemia brain and primary microglia stimulated by Treg cells,together with control groups were send to RNAseq.The possible mechanism of interaction between Treg cells and microglia was screened out using String database.In vivo experiments using transgenic mice together with in vitro experiments were used for the verification of the mechanismResults:The results of RNAseq suggest that Treg cells infiltrated into brain after ischemic stroke up-regulated the regulatory function towards other immune cells,and the microglia stimulated by Treg up-regulate those genes related to white matter repair.Protein-protein interaction analysis suggested the possibility of Treg cells interacting with the integrin subunit beta 1(Itgb1)on the surface of microglia by secreting osteopontin(OPN).In vitro experiments proved that stimulation of Treg cells can improve the ability of microglia to promote the differentiation and maturation of oligodendrocyte precursor cells(OPC).In vivo experiments have shown that Treg cell depletion can change the activation state of microglia/macrophages after ischemic stroke.After knocking out the OPN of Treg cells,the number of new generated oligodendrocytes(OL)in the brain tissue after ischemic injury is significantly reduced and the coverage of the myelin sheath at the edge of the injury is decreased.Recombinant OPN can enhance the ability of primary microglia to promote OPC differentiation.To verify whether OPN acts via the Itgb1 receptor on the surface of microglia,we used conditional knockout(c KO)mice to knock out the Itgb1 specifically in microglia.The results indicates that the regeneration of OL in the ischemic brain promoted by Treg cells is significantly reduced.The in vitro cell co-culture system confirmed that OPNko or Itgb1 antibody would reduce OPC differentiation which promoted by microglia.The IL2/IL2 Antibody complex(IL2/IL2Ab)can increase the number of peripheral blood and brain infiltrated Treg cells as well as the number of OPN~+Treg cells in the brain after ischemic injury and promote the remyelination of injured brain.Conclusion:Osteopontin(OPN)produced by Treg cells interacts with the integrin subunit beta(Itgb1)on the surface of microglia to enhance the white matter repair function of microglia after ischemia stroke.IL2/IL2Ab can promote the repair of brain white matter after stroke.