Neuroprotective Mechanisms of CXCL12 During CNS Autoimmunity

Robyn S. Klein, M.D., Ph.D.

Washington University School of Medicine

Funded in September, 2006: $200000 for 3 years


back to top

Challenging Current Thinking about Molecular Actions in Autoimmune Multiple Sclerosis

This animal and human research study will explore the possibility that specific molecular interactions direct immune lymphocytes to the blood-brain-barrier (BBB) and minimize brain tissue inflammation in multiple sclerosis (MS) and its animal counterpart, “EAE.” If so, this finding would be contrary to current thinking.

Autoimmune MS in humans and EAE in animals are characterized by immune-based inflammation and destruction of myelin, the insulator of nerve axons and conductor of electrical communication signals along the axons. How, though, do immune lymphocytes (white blood cells) manage to get through the BBB, which ordinarily only opens to immune cells once a harmful agent has been identified there? The standard view is that “attractant” molecules, called chemokines, are produced and displayed on the lining of brain blood vessels.  These chemokines then guide the disease-producing white blood cells into the brain, where they harm the myelin, the fatty sheath that insulates nerve cell axons (the communication cables). Dr. Klein's observations, however, suggest that, for certain chemokines, precisely the opposite occurs.  She has found evidence that a chemokine, called CXCL12, actually might help immobilize the white blood cells, rather than guide them into the brain.  She will now pursue these surprising findings in EAE, the animal model of MS.  Additionally, in collaboration with other researchers, she will also study autopsy brain specimens from patients with MS who have died and from patients who died of other causes. This will allow her to ascertain whether CXCL12 expression is altered during MS and if and how this relates to disease severity.

Significance:  Understanding the molecular events that allow immune system lymphocytes to enter the brain and produce tissue damage in autoimmune MS may lead to development of new therapies targeted to these molecular events.


back to top

Neuroprotective Mechanisms of CXCL12 During CNS Autoimmunity

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.


back to top
Robyn S. Klein, M.D., Ph.D.

Dr. Klein joined the Washington University School of Medicine in 2003. She received her M.D. and Ph.D. degrees from Albert Einstein College of Medicine. She then completed her internship and residency in Internal Medicine at the Brigham & Women's Hospital, Harvard University and her fellowship in Infectious Diseases and post-doctoral training in Immunology at the Massachusetts General Hospital, Harvard University. Her long-term research goal is to understand the molecular basis of inflammation-associated dysfunction and damage in the central nervous system (CNS) in viral and autoimmune encephalitides and to identify potential therapeutic targets for the treatment of neuroinflammatory diseases.

Work in the Klein laboratory focuses on two components of CNS inflammatory states: the mechanism of leukocyte recruitment into the CNS and the direct effects of inflammatory mediators on neurons. Common to both of these is the action of chemokines, which both recruit leukocytes into the CNS and signal through chemokine receptors present on neurons, affecting their function and survival. Her experimental approach involves the development of in vitro and in vivo models of CNS mononuclear cell recruitment and neuronal chemokine receptor signaling responses. Using these tools, she examines the role of chemokines and their receptors in both normal and inflamed CNS in order to determine how their actions relate to the wide range of pathology observed in CNS inflammatory diseases. Understanding the mechanism of mononuclear cell recruitment within the CNS and the neuronal injury induced by their secreted chemokines is crucial for the development of therapies that limit CNS damage in a variety of infectious and autoimmune diseases.

Dr. Klein is a member of the American Association of Immunologist, the American Society for Microbiology, and the Society for Neuroscience and is a founding member of the International Society for Neurovirology.