2 Demyelinating Disease 2.1 Multiple Sclerosis
3 Role of Chemokines
The role of chemokines in these stages of evolution of brain disease must now be considered.
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Chemokine biology has been described elsewhere and the reader is referred to excellent reviews on the topic (Cyster, 1999; Rossi and Zlotnik, 2000). The CC and CXC families contain by far the most of the known chemokines and so predominate in discussion of chemokines in inflammatory and demyelinating disease. The CC and CXC chemokines act through G-protein coupled 7-transmembrane CCR and CXCR receptors, respectively, and chemokine involvement in disease and develop-mental processes is divined both through analysis of receptor expression and of chemokines themselves.
The by-now classic concept underlying the role of chemokines in cellular entry to tissues includes that CC or CXC chemokines are presented on endothelium to attract specific cell types to targeted locations. These chemokines act on cells in blood via specific receptors and this induces upregulation and activation-dependent conformational change of adhesion ligands, leading to arrest and migration at the site of chemokine expression and adhesion receptor upregulation. Response to chemokines is concentration-dependent and a gradient effect ‘guides’ cells towards the source of their production (reviewed in (von Andrian and Mackay, 2000) ).
To understand how this operates in the context of entry of immune cells to the CNS, we must first define those chemokines that are candidate players in these immune processes. Then their induction and presentation to leukocytes at the blood–brain barrier will be discussed, particularly in experimental models for this, including transgenic and knockout animals.
3.1 Chemokines that Drive T Cell Entry
The migration of T cells in inflammatory responses is predominantly guided by receptors for CC chemokines. Mature activated T cells express CCR1, CCR2 (pri-marily a receptor for MCP-1 or CCL2), CCR3, CCR4 and CCR5 (reviewed in (Moser et al., 2004) ). It is noteworthy that the chemokine receptors expressed by T cells that participate in tissue responses tend to be those that are ‘promiscuous’in their ligand binding, allowing such T cells to respond to a broad range of potential chemoattractants. Many of the CC receptors we have listed bind CCL5/RANTES, though CCR3 and CCR5 are the receptors most commonly associated to CCL5 response. CCR4 binds CCL3/MIP-1α and CCL2/MCP-1, as well as CCL5/
RANTES, and is more associated to the Th2 subset (see below).
The precise pattern of receptor expression can be used to sub-classify T cells and has functional correlates. Th1 and Th2 CD4 + T cells can be distinguished by their reciprocal expression of CXCR3 (Th1) and CCR3 (Th2) (reviewed in (Baggiolini, 1998; Moser et al., 2004) ). Th1 T cells, most strongly associated with inflammation, express CXCR3, which binds the ELR-CXC chemokines CXCL10/IP-10, CXCL9/I-TAC and CXCL11/MIG. Th1 cells also express CCR1 and CCR2. The chemokines CCL2, CCL5 and CXCL10 are of particular interest because of their regulation (eg. IP-10 = interferon-regulated protein-10), and because they are upregu-lated in the inflamed CNS (Engelhardt and Ransohoff, 2005). The Th2-associated CCR3 binds CCL11/eotaxin, CCL8/MCP-2, CCL7/MCP-3 and CCL13/MCP-4.
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The chemokines listed so far fall into a general category of inflammatory chemokines. The alternate category is homeostatic, whose members act on rest-ing leukocytes or at initiation of responses (Moser et al., 2004). These have a potentially important role to play in autoimmunity. Two homeostatic chemokines deserve consideration in demyelinating disease. Naive T cells in circulation respond to CXCL12, also known as SDF (stromal cell-derived factor), via CXCR4. The functionally-related chemokine CXCL13/BCA-1 or BLC is best-known for effects in the lymph node where it directs CXCR5 + B and T cell migration to enter follicles. However, CXCL12 and CXCL13 have also been detected in MS CSF and may act on T cells in CNS (Krumbholz et al., 2006).
3.2 Chemokines in B Cell Entry to CNS
There are essentially no B cells in the normal CNS and relatively few even in MS.
The B cells that are found in the CNS in MS include plasmablasts (CD19 + CD138+) and plasma cells (CD19-, CD138+), as well as memory phenotype B cells (eg. CD27+) and cells with a centroblast-like phenotype (CD19 + , CD38high, CD77 +, Ki67+) (Meinl et al., 2006). The latter are associated with germinal center-like follicular aggregates (Serafini et al., 2004), that are especially prominent in pro-gressive disease. Nevertheless, because oligoclonal Ig bands are a diagnostic for MS, it is obvious that B cell entry and activation occur early. The mechanism underlying B cell migration to the CNS has not been as thoroughly examined as that for T cells, but processes analogous to those guiding T cell entry operate.
TNFα and TNF-family members (BAFF, APRIL) are identified as B cell factors which promote B cell survival in the CNS. It remains to be established whether they play an analogous role in induction of B cell-tropic chemokines, as TNFα does for T cells. The chemokines CXCL12 and CXCL13 are detected at elevated levels in EAE and MS CSF (Columba-Cabezas et al., 2003; Magliozzi et al., 2004;
Krumbholz et al., 2005), and their receptors CXCR5 and CXCR4 respectively are expressed on plasmablasts and on B cells. These chemokine are best-known for directing B cell migration within lymph nodes and entry to follicles, and their expression in MS CNS is consistent with such activity. The inflammatory chemok-ine CXCL10 or IP-10, whose expression is strongly elevated in MS and EAE, also acts on B cells, via the CXCR3 receptor, and is implicated in entry of plasmablasts to the CNS (Meinl et al., 2006).
3.3 Chemokines that Drive Macrophage and Neutrophil Entry
Monocytes and macrophages express a wide range of chemokine receptors. The very naming of macrophage chemoattractant protein’s (MCP’s) and macrophage inflammatory proteins (MIP’s) identifies a role for CCL2/MCP-1, CCL3/MIP-1α, CCL4/MIP-1β, CCL9/MIP-1γ, to list a few of these CC chemokines, in regulating
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macrophage migration. Macrophages also express receptors for CCL5/RANTES, and for the CXC chemokine CXCL10. Other CXC chemokines are associated with neutrophil trafficking. Predominant among these are CXCL1, CXCL2 (or KC) and CXCL8/IL-8, all acting through the CXCR2 receptor (reviewed in Rossi and Zlotnik, 2000).
3.4 Chemokines in MS
Studies on chemokine expression in MS patients have revealed that the composition of chemokines expressed depends on the subtypes of MS. By measuring chemokine expression in PBMCs via real-time PCR and applying a multivariate statistical analysis it was possible to distinguish healthy individuals versus MS patients as well as primary progressive versus RR-MS, respectively (Furlan et al., 2005).
Interestingly chemokines (CCL2/MCP-1,CCL3/MIP-1α, CCL4/MIP-1β, CCL8/
MCP-2 and CCL7/MCP-3) and their receptors (CCR2, CCR3, and CCR5) driving T cell and macrophage infiltration, were shown to be upregulated in CSF and lesions of RR-MS patients during relapse (McManus et al., 1998a; Simpson et al., 1998, 2000a; Bartosik-Psujek and Stelmasiak, 2005).
At anatomical sites like vascular endothelium, perivascular space and paren-chyma, chemokines are differently expressed, (reviewed in (Muller et al., 2004) ) indicating their putative functions to guide leukocytes through different possible migration routes to CNS as e.g. (1) from blood to CSF across the choroid plexus, (2) from blood to subarachnoid space and (3) from blood to parenchymal perivas-cular space (Ransohoff et al., 2003). The expression of CCL21/SLC on choroid plexus epithelium and the detection of CCR7 positive T cells in CSF of MS patients speaks for a direct entry of T cells to CFS from systemic circulation (Kivisakk et al., 2004), a process which is dependent on P-selectin expression (Kivisakk et al., 2003). Furthermore Simpson et al. (1998) reported selective expression of CCL5/
RANTES in blood vessel endothelium, perivascular cells and glial limitants indi-cating that CCL5/RANTES is functional during transmigration from blood to parenchyma via the perivascular space. In vitro studies analyzing transmigration of ex vivo leukocytes of MS patients across human brain-derived endothelial cells (HBECs) revealed that endothelial cells produce CCL2/MCP-1 which drives leu-kocyte movement (Prat et al., 2002; Seguin et al., 2003). CCL2/MCP-1 was shown to be present in CSF of MS patients, however at reduced levels during relapse of the disease (Mahad et al., 2002; Bartosik-Psujek and Stelmasiak, 2005). This apparent contradiction between HBECs in vitro and chemokine expression studies in MS respectively was solved by the study of Mahad et al. showing that leukocytes consume CCL2/MCP-1 during migration across the blood-brain barrier (BBB) (Mahad et al., 2006). CCL2/MCP-1 is produced by parenchymal glial cells in active lesions of MS patients as well (Simpson et al., 1998). While expression of CCL2/MCP-1 by endothelial cells may be responsible for leukocyte transmigration across BBB, its expression within the parenchyma may result in augmenting local
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inflammation. CCL2/MCP-1 represents therefore an example that the same chemokine fulfils different functions depending at which anatomical site it is expressed.
It is known that chemokines not only contribute in triggering leukocyte movements. This counts among others for the homeostatic chemokines CXCL12/
SDF-1 and CXCL13/BCA-1. Functions of CXCL12/SDF-1 have been studied using New Zealand Black/New Zealand White mice, expressing a self-reactive repertoire resulting in lupus associated nephritis. Administration of anti-CXCL12/SDF-1 antibodies repressed lymphocyte activation and autoantibody production in these mice (Balabanian et al., 2003). Furthermore CXCL12/SDF-1 was shown to partially inhibit spontaneous apoptosis of adenotonsillar memory T cells (Pajusto et al., 2004). Considering the homeostatic functions and the pres-ence of CXCL12/SDF-1 in active MS lesions (Calderon et al., 2006; Krumbholz et al., 2006) it may be assumed that this chemokine is not only responsible to attract leukocytes to CNS but more importantly regulates the subsequent autoim-mune reaction. Coricone et al. detected CD19+ CD38highCD77+ Bcl2− B cells in CSF of MS patients together with CXCL12/SDF-1 and CXCL13/BCA-1 expres-sion on the outer layer of capillaries. As these B cells are normally exclusively present in secondary lymph nodes and CXCL12/SDF-1 together with CXCL13/
BCA-1 is known to be key mediator of lymphoneogenesis the authors concluded that B cell differentiation occurs within the CNS (Corcione et al., 2004). This is supported by the detection of lymphoid follicle-like structures containing B cells and CXCL13/BCA positive dendritic cells within cerebral meninges of second-ary progressive MS patients (Serafini et al., 2004) and in mice with EAE (Magliozzi et al., 2004). Furthermore lymphoid chemokines like CCL19/MIP-3α and CCL21/SLC have been described in EAE (Alt et al., 2002; Columba-Cabezas et al., 2003) and in MS (Pashenkov et al., 2003). Coeval expression of their receptor CCR7 was demonstrated on T cells infiltrating the parenchyma during EAE (Alt et al., 2002). However CCR7 expression on T cells within the paren-chyma could not be confirmed in MS patients (Kivisakk et al., 2004). Therefore, the mechanisms of the local induction and maintenance of autoimmunity through the formation of lymphoid follicle-like structures within the CNS needs to be further analyzed.