However, the increased difference in migratory rates of Treg and non-Treg in the presence of a MBMEC layer hints to Treg-specific interactions with the endothelial Opaganib supplier cell layer, either due to direct cell–cell contact or due to a constitutive secretion

of soluble factors by the endothelial cells. CCL20 as a soluble stimulus secreted by the MBMEC layer can be excluded since its expression is only found in epithelial cells of the choroid plexus and astrocytes during EAE relapse 20, 21 but not in brain endothelium. More likely, Treg seem to have an advantage in forming stable cell–cell contacts with the brain endothelium, consistent with their higher expression of LFA-1 and CD49d, as they intensively accumulated in or on top of the endothelial cell monolayer compared to their non-regulatory counterparts. The preferential migration of Treg through a porous membrane in the presence of the chemoattractant CCL20 was expected by their CCR6 cell surface expression see more and was maintained when T cells migrated across an in vitro model of the BBB. In the non-regulatory fraction,

particularly the Th17 cells should be attracted by the CCL20 gradient as they are known to express high amounts of CCR6 compared to other effector cell types 22. This finding further supports the current notion that CCR6 expressing, autoreactive effector Th17 cells may be able to gain entry to the yet non-inflamed CNS, facilitated through CCL20 secretion by epithelial cells of the choroid plexus or brain resident glia cells 21, 23, 24, and induce the subsequent immune responses by producing CCL20 among other inflammatory stimuli 22. In consequence, this might

lead to inflammation of the BBB endothelium allowing further, CCL20 independent lymphocyte infiltration into the CNS parenchyma. Treg, exhibiting a stronger migratory response to CCL20 than conventional CD4+ T cells, should therefore Aldehyde dehydrogenase have a higher prevalence in the brain tissue compared to their effector counterparts under healthy conditions, consistent with our in vivo finding. Human Treg have been reported to be present in the CNS in certain neurological disorders, such as gliomas 25, 26. Under conditions of experimental autoimmune neuroinflammation as in EAE, Treg accumulate in the murine CNS 4, 10, most notably in the remission phases 11, counterbalancing encephalitogenic CNS responses. As mentioned above, data on the presence and function of Treg in the human CNS are sparse 12–14, 18. To translate our findings into human pathophysiology, we used an in vitro model of the human BBB to mimic lymphocyte diapedesis in vivo. In contrast to HD, MS patient-derived Treg failed to outmatch their non-regulatory counterparts in crossing the BBB under basal, non-inflammatory conditions.