The central nervous system (CNS) is considered an immunologically specialized site where leukocyte trafficking is restricted by the blood-brain barrier (BBB), a complex organization of tight junction-coupled endothelial cells, whose basement membranes are enveloped by glial foot processes. Leukocytes that traverse the microvasculature must exit the perivascular space through this glial limitans to gain entry into the CNS parenchyma.
In the autoimmune disease multiple sclerosis (MS) and in its animal model experimental autoimmune encephalomyelitis (EAE), the development of demyelinating lesions within the CNS is associated with the perivascular accumulation of mononuclear cells. Studies suggest that mononuclear cells within these infiltrates enter the CNS parenchyma and initiate inflammation that leads to demyelinating lesions. Although several adhesion molecules have been implicated in the interactions between mononuclear cells and CNS endothelium, the chemoattractant molecules responsible for leukocyte movement into and out of the perivascular space are unknown and are of considerable interest for the development of therapies that limit the development of inflammatory infiltrates in patients with MS.
CXCL12 is a secondary lymphoid chemokine that is constitutively expressed at multiple tissue sites including the CNS. Although CXCL12 has been detected within microvessel endothelial cells of the BBB and has been shown to exhibit a pro-inflammatory role in a variety of autoimmune diseases, its role in leukocyte infiltration of both the normal and inflamed CNS has not been established. Utilizing confocal microscopy, we observed that endothelial cell expression of the chemokine CXCL12 normally displays basolateral polarity at the BBB. Surprisingly, this polarization is lost during CNS autoimmune disease, suggesting CXCL12 may play a role in regulating BBB function.
We tested this hypothesis by administering AMD3100, a specific antagonist of the CXCL12 receptor, CXCR4, during the induction of EAE. In preliminary studies we observed that CXCR4 antagonism significantly enhances the migration of infiltrating leukocytes into the CNS parenchyma during EAE, leading to a striking loss of the typical, intense perivascular cuffs, and commensurate increase in the lesion size, with worsened associated demyelination and clinical severity. CXCL12 is most commonly postulated to play a pro-inflammatory role in the CNS but these dramatic results lead us to propose a novel, anti-inflammatory role for CXCL12.
We hypothesize that CXCL12 functions to localize CXCR4-expressing mononuclear cells to the perivascular space, thereby limiting the parenchymal infiltration of autoreactive, effector cells. These results also strongly suggest that the perivascular space itself constitutes a specialized, immunomodulatory site within the CNS. Consistent with this hypothesis, we observed that extensive mononuclear trafficking out of this site leads to significantly increased levels of pro-inflammatory mediators within the CNS parenchyma. Thus, premature migration of cells from the perivascular space might affect their exposure to counter-regulatory immune mechanisms that normally minimize immune activation during CNS autoimmune disease.
Using MS and EAE as model systems to study these fundamental questions in the immunobiology of the CNS, we will extend our preliminary findings to define how loss of localization to the perivascular space regulates immune activation of mononuclear and microglial cells through the following Specific Aims:
1. We will determine how CXCL12-mediated perivascular localization regulates mononuclear cell trafficking during CNS autoimmunity.
2. We will determine how perivascular localization regulates mononuclear cell activation during CNS autoimmunity.
An enhanced understanding of the molecular signals that govern immune cell recruitment and activation at the BBB will facilitate the development of targeted anti-inflammatory agents that mitigate the morbidity and mortality associated with CNS autoimmune diseases.