Targeting the BBB to treat CNS inflammation

点击翻译按钮获取中文摘要

Project 2 will determine how targeting molecules that regulate permeability and polarity at central nervous system (CNS) endothelial barriers impacts neuroinflammation, as assessed via novel neuroimaging modalities with murine models of multiple sclerosis (MS). We have shown that endothelium in MS lesions exhibits altered polarity, displaying the abluminal chemokine CXCL12 aberrantly along luminal surfaces. Abluminal CXCL12 normally serves to limit leukocyte entry whereas luminal expression of CXCL12 is associated with increased activation of its signaling receptor CXCR4 on infiltrating leukocytes. We showed that the scavenging CXCL12 receptor, CXCR7 (also expressed by CNS endothelium), is a critical regulator of leukocyte entry via internalization of CXCL12 from abluminal surfaces. In preliminary studies, we have also found that sphingosine 1-phosphate (S1P) signaling via S1P2 disturbs abluminal expression of CXCL12, which then disrupts immune privilege. Thus, mice with targeted deletion of S1P2 or administered a specific antagonist, JTE-013, show reduced migration of leukocytes into the CNS parenchyma during EAE. We hypothesize that alterations in membrane polarity of CXCL12 at CNS endothelial barriers promote leukocyte capture and migration into the CNS, contributing to the establishment of disease cycles in relapsing-remitting forms of CNS autoimmunity. We have identified several molecules that regulate this process, leading to reversal of apicobasal expression of the localizing cue, CXCL12. In this proposal, we will examine the roles of CXCR7 and S1P2 in lymphocyte trafficking and inflammation in CNS autoimmune disease using novel imaging modalities including two-photon intravital microscopy and longitudinally using Diffusion Basis Spectrum Imaging (DBSI), which distinguishes and quantitates cellularity and edema in live mice (see Project 1) and examination of autopsied human MS patient and control CNS tissues. We expect DBSI to be more sensitive than Gd enhancement to inflammation, including in the setting of less profound blood-brain barrier alterations. To determine the relationships between endothelial and immune cell (Th1 versus Th17) interactions and loss of BBB integrity, we will directly compare results utilizing intravital imaging to DBSI and Gd enhancement in the same animals. To address mechanisms by which endothelial cell polarity and localizing cues are induced by interactions with inflammatory cells, we will utilize in vitro and in situ approaches with human tissues. Thus, Aim 1 will determine whether CXCR7-mediated internalization of CXCL12 requires infiltration with Th1 versus Th17 cells during EAE. Aim 2 will determine whether S1P2- mediated reversal of endothelial cell polarity leads to increased T cell capture and entry during EAE, and Aim 3 will examine the relationship between CXCR7 and S1PR2 activation and T cell migration at the BBB in MS. The experimental design of Project 2 will define how DBSI determined cellularity and edema changes responding to various degrees of BBB abnormality and cell infiltration in EAE mice.