Cell and Tissue Research

, Volume 349, Issue 1, pp 229–248

Chemokines in CNS injury and repair

Authors

  • Anne Jaerve
    • Molecular Neurobiology Laboratory, Department of NeurologyMedical Faculty Heinrich Heine University
    • Molecular Neurobiology Laboratory, Department of NeurologyMedical Faculty Heinrich Heine University
Review

DOI: 10.1007/s00441-012-1427-3

Cite this article as:
Jaerve, A. & Müller, H.W. Cell Tissue Res (2012) 349: 229. doi:10.1007/s00441-012-1427-3

Abstract

Recruitment of inflammatory cells is known to drive the secondary damage cascades that are common to injuries of the central nervous system (CNS). Cell activation and infiltration to the injury site is orchestrated by changes in the expression of chemokines, the chemoattractive cytokines. Reducing the numbers of recruited inflammatory cells by the blocking of the action of chemokines has turned out be a promising approach to diminish neuroinflammation and to improve tissue preservation and neovascularization. In addition, several chemokines have been shown to be essential for stem/progenitor cell attraction, their survival, differentiation and cytokine production. Thus, chemokines might indirectly participate in remyelination, neovascularization and neuroprotection, which are important prerequisites for CNS repair after trauma. Moreover, CXCL12 promotes neurite outgrowth in the presence of growth inhibitory CNS myelin and enhances axonal sprouting after spinal cord injury (SCI). Here, we review current knowledge about the exciting functions of chemokines in CNS trauma, including SCI, traumatic brain injury and stroke. We identify common principles of chemokine action and discuss the potentials and challenges of therapeutic interventions with chemokines.

Keywords

ChemokinesCentral nervous system (CNS)CNS injuryCNS repairAxon sproutingStem cells

Introduction

A feature common to spinal cord injury (SCI), traumatic brain injury (TBI) and stroke is that both the primary insult and the secondary degeneration events that amplify the primary damage lead to devastating neurological dysfunction (Fig. 1). Post-traumatic infiltration of inflammatory cells has been associated with secondary tissue damage, cell death and demyelination of axons. The central mediators of cell activation and recruitment are chemokines, which are small (8-14 kDa) polypeptides that are produced in elevated amounts at the traumatic injury site by resident tissue cells, activated resident and recruited leucocytes, cytokine-activated endothelial cells and some neurons. Recently, injury-induced chemokine expression in the liver has been shown to be important for cell recruitment into injury sites of the central nervous system (CNS; Campbell et al. 2005). Chemokines represent one of the first completely known molecular superfamilies that has expanded via gene duplication (DeVries et al. 2006); it consists in about 50 members of mammals, chicken, zebrafish, sharks, jawless fish and Caenorhabditis elegans (Zlotnik et al. 2006). Chemokines are classified into four subfamilies according to the position of two conserved cystein residues in the mature protein: CXC, CC, XC and CX3C. Those that have changed after the segregation of human and mice and that are located in clusters on chromosomes are mostly involved in inflammation (Zlotnik et al. 2006), whereas more ancient and conserved chemokines with distinct chromosomal locations have functions in the coordination of cell migration during the development and homeostasis of the CNS (Table 1). The arising redundancy in chemokine signalling networks and functions is further complicated by the promiscuity of binding to only about 18 known receptors. Uniquely, CXCL12 binds to two different receptors: CXCR4 and CXCR7.
https://static-content.springer.com/image/art%3A10.1007%2Fs00441-012-1427-3/MediaObjects/441_2012_1427_Fig1_HTML.gif
Fig. 1

Role of chemokines in spinal cord injury (SCI), traumatic brain injury (TBI) and stroke. The blood-brain barrier, which separates the brain or spinal cord parenchyma from the vascular system consists in endothelial cells (depicted as a blood vessel), is disrupted following SCI and TBI and becomes dysfunctional after stroke (GM grey matter, WM white matter). Red and white blood cells infiltrate (in stroke after reperfusion) and interact with resident immune cells via cytokines, such as interleukin-1β (IL-1β) and tumor necrosis factor-α, which activate glial cells and promote their proliferation and the production of chemokines in order to induce inflammation. Microglial cells migrate towards CCL2 sources and produce both neurotoxic and neuro-supportive factors. Astrocytes and, to a lesser extent, endothelial cells produce CCL2, CCL3, CXCL1, CXCL2 and other chemokines to elicit the migration of immune cells. Neutrophils are recruited by CXC chemokines CXCL1, CXCL2 and CXCL8 (depending on the IL-1 receptor) and by CC chemokines CCL2 and CCL3, which, however, mostly recruit monocytes bearing CCR2. CXCL10 recruits T lymphocytes but also monocytes from the bone marrow, as does CXCL12 (which is upregulated at later time points). In TBI, neutrophils can enter the CNS via the choroid plexus, which secretes CXCL1. Recruited leucocytes themselves produce chemokines that further amplify the recruitment process. In addition to cell recruitment, chemokines promote regenerative processes. For example, neovascularization by infiltrating endothelial progenitor cells or tissue resident precursors is dependent on chemokines (CCL2, CCL16, CXCL12, CXCL1-8, except CXCL4). Neuroprotection by the recruitment of stem cells is mediated mostly by CXCL12. In addition, CXCL12 promotes axonal sprouting. Some chemokines promote oligodendrocyte proliferation and remyelination (CXCL12, CXCL1)

Table 1

Chemokines, receptors and inflammatory function (NK natural killer cell). Based on Zlotnik et al. (2006), David and Kroner (2011) and Keeley et al. (2010)

Name

Acronym

Receptor

Chromosomal location

Inflammatory (I), homeostatic (H),dual (D), or unknown (U) function

Type of recruited leucocyte

CXC chemokine family

 

 CXCL1

GROα,KC

CXCR2

4q13.3

I

Neutrophil, monocyte

CINC-1

CXCR1

 CXCL2

MIP-2, GROβ

CXCR2

4q13.3

I

Neutrophil, monocyte

 CXCL3

Groγ

CXCR2

4q13.3

I

Neutrophil, monocyte

DcipI

 CXCL4

PF4

CXCR3B

4q13.3

U

Neutrophil, monocyte, T cell

 CXCL5

Lix

CXCR2

4q13.3

I

Neutrophil

ENA-78

CXCR1

 CXCL6a

GCP-2

CXCR2

4q13.3

I

Neutrophil

CXCR1

 CXCL7

Ppbp, NAP-2

CXCR2

4q13.3

I

Neutrophil

 CXCL8a

IL-8

CXCR1

4q13.3

I

Neutrophil

CXCR2

 CXCL9

MIG

CXCR3

4q21.1

I

Monocyte, T cell, NK

CXCR3B

 CXCL10

IP-10

CXCR3A

4q21.1

I

Monocyte, T cell, NK

 CXCL11

I-TAC

CXCR3

4q21.1

I

Monocyte, T cell, NK

CXCR3B

CXCR7

 CXCL12

SDF-1α/β

CXCR4

10q11.1

H

Monocyte, T cell, B cell, dendritic cell

CXCR7

 CXCL13

BLC

CXCR5

4q21

H

Monocyte, B cell

BCA-1

 CXCL14

BRAK,

U

5q31.1

I

Neutrophil, monocyte, dendritic cell

Bolekine

CXCL15b

Weche, lungkine

CXCR2

U

I

Neutrophil

 CXCL16

 

CXCR6

17p13.2

I

T cell

 CXCL17

 

U

19q13.2

U

 

 

CC chemokine family

 

 CCL1

I-309, TCA-3

CCR8

17q11.2

I

Monocyte, T cell

 CCL2

MCP-1, JE

CCR2

17q11.2

I

Neutrophil, eosinophil, basophil, monocyte, T cell, dendritic cell, NK

 CCL3

MIP-1α, LD78α

CCR1

17q11.2

I

Eosinophil, monocyte,T cell, B cell, dendritic cell, NK

CCR5

 CCL4

MIP-1β

CCR1

17q12

I

Monocyte, T cell, dendritic cell, NK

CCR5

 CCL5

Rantes

CCR1

17q12

I

Eosinophil, basophil, monocyte, T cell, dendritic cell, NK

CCR3

CCR5

 CCL6b

 

U

 

I

 

 CCL7

MCP-3, MARC

CCR1

17q11.2

I

Eosinophil, basophil, monocyte, T cell, dendritic cell, NK

CCR2

CCR3

CCR5

 CCL8

MCP-2

CCR2

17q11.2

I

Eosinophil, basophil, monocyte, T cell, dendritic cell, NK

CCR5

CCR1

CCR3

 CCL9b

 

U

 

I

 

 CCL10b

 

U

 

I

 

 CCL11

Eotaxin

CCR3

17q11.2

I

Eosinophil, basophil, monocyte, T cell, dendritic cell

CCR5

 CCL12b

MCP-5

CCR2

 

I

Neutrophil, basophil, monocyte, T cell, dendritic cell, NK

 CCL13a

MCP-4

CCR1

17q11.2

I

Eosinophil, basophil, monocyte, T cell, dendritic cell, NK

CCR3

CCR2

CCR5

 CCL14a

HCC-1

CCR1

17q12

H

T cell, monocyte

CCR3

CCR5

 CCL15a

MIP-1γ, HCC-2

CCR1

17q12

H

Eosinophil, basophil, monocyte, T cell

MMRP2, CCF18

CCR3

 CCL16a

HCC-4, LEC

CCR1

17q12

H

Monocyte, T cell

CCR2

CCR5

CCR8

HRH4

 CCL17

TARC

CCR4

16q13

D

Monocyte, T cell, dendritic cell

CCR8

 CCL18a

PARC

U

17q12

H

Monocyte, T cell

 CCL19

MIP-3β, ELC

CCR7

9p13.3

H

Monocyte, T cell, dendritic cell

 CCL20

MIP-3α, LARC

CCR6

2q36.3

D

Monocyte, dendritic cell

 CCL21

SLC

CCR7

9p13.3

D

T cell, dendritic cell

 CCL22

MDC, ABCD-1

CCR4

16q13

D

Monocyte, T cell, dendritic cell

 CCL23a

MPIF-1, C10

CCR1

17q12

I

Monocyte, T cell

FPRL-1

 CCL24

Eotaxin-2

CCR3

7q11.23

I

Eosinophil, basophil, monocyte, T cell, dendritic cell

 CCL25

TECK

CCR9

19p13.2

H

Monocyte, T cell, dendritic cell

 CCL26

Eotaxin-3

CCR3

7q11.23

I

Eosinophil, basophil, monocyte, T cell

 CCL27

CTACK, ILC

CCR10

9p13.3

H

T cell

 CCL28

MEC

CCR3

5p12

U

T cell

CCR3

 

C chemokine family

 

 XCL1

Lymphotactin α

XCR1

1q24.2

D

T cell

 XCL2a

Lymphotactin β

XCR1

1q24.2

D

T cell

CX3C chemokine family

 CX3CL1

Fractalkine

CX3CR1

16q13

I

Monocyte, T cell, NK

aHuman only

bMouse only

Although the good or bad role of inflammation is still under debate (Yong and Rivest 2009), the immunomodulatory role of chemokines is now being translated into anti-inflammatory interventions to reduce the destruction arising from the infiltration of inflammatory cells and consequent secondary damage after injury (Donnelly et al. 2011). Such studies have also reported chemokine-supported enhanced neuroprotection, tissue preservation and axon sprouting (Glaser et al. 2004, 2006). We and others have recently shown that CXCL12 directly promotes neurite outgrowth on inhibitory myelin in cultured neurons and moreover, that it enhances the sprouting of axons after SCI (Chalasani et al. 2003; Opatz et al. 2009; Jaerve et al. 2011). Previously, a role for CXCL12 in recruiting stem and progenitor cells into the lesion site in the injured spinal cord has been demonstrated (Takeuchi et al. 2007). The reduction of secondary damage and the stimulation of regenerative processes such as axon sprouting make chemokines a promising target for the repair of SCI, TBI and stroke. For translation into clinical application, the off-target effects of chemokines remain a major limitation. An essential step is therefore to bypass the redundant and important homeostatic functions of chemokines via cell-specific targeting in order to block, for example, the migration of inflammatory cells but the migration and function of progenitor/stem cells is therefore not essential. In this review, we further discuss other aspects that are potentially relevant for the targeting of the chemokine system, such as neuropathic pain and ageing.

Receptors and signalling

Chemokines are usually secreted by the producing cell. As an exception, CXCL16 and CX3CL1 are first expressed as transmembrane proteins but, upon proteolytic cleavage, the extracellular domain is released as a soluble fragment (Ludwig and Weber 2007). Secreted chemokines undergo rapid enzymatic processing into moderately active (CXCL5) or more active (CXCL7, CXCL8, CCL4, CCL14, CCL15, CCL23) molecules or receptor-specific isoforms (CCL3L1, CCL5; Mortier et al. 2011). Further processing of chemokines can produce natural chemokine signalling antagonists or might lead to the total inactivation or degradation of chemokines (Mortier et al. 2011). Modulation of chemokine stability is one of the tools to manipulate the chemokine system. In this respect, modified CXCL12 is less accessible for degradation and thus is active for a longer period of time. An important protection from rapid degradation/inactivation of highly positively charged chemokines is their interaction with negatively charged glycosaminoglycans. This is also central for building up the chemokine gradient and the stabilization of structures that would not otherwise form in solution (Salanga and Handel 2011). For example, oligomeric forms seem to be required for migration in vivo, whereas the binding of monomeric CCL2, CCL4, CCL5 and CCL10 to receptors induces cell migration in vitro. In contrast, dimeric CXCL12 is unable to induce cell migration but the elevated intracellular Ca2+ levels and cell migration can be stopped by dimerization at high CXCL12 concentrations. Moreover, receptors of chemokines have been reported to homo- or heterodimerize but also might form heterodimers with other cell surface receptors (Wang and Norcross 2008).

Chemokine signalling is mediated by members of 7-transmembrane–spanning G-protein–coupled receptors (GPCRs). Upon ligand binding, these receptors (CXCR1-6, CCR1-10, XCR1 and CX3CR1) are internalized from the cell surface and a wide range of intracellular pathways/targets such as phosphatidylinositol-3 kinase, mitogen-activated protein kinases and protein kinase C, adenylcyclase, phospholipases, GTPases such as Rho and Rac and Cdc42 are activated (Stamatovic et al. 2005; Wain et al. 2002; Neves et al. 2002). G-protein-mediated phosphoinositide hydrolysis generates diacylglycerol and inositol 1,4,5-trisphosphate, which then activate protein kinase C allowing the mobilization of calcium to initiate cellular responses such as migration, maturation, proliferation, survival, gene transcription and cytokine production (Wu et al. 1993). The molecular mechanisms behind the various effects, which are often elicited simulatenously, e.g. migration and survival, are only now being studied. For example, similar signalling molecules appear to play different regulatory roles downstream of chemokine receptors. In dendritic cells, CXCL12-dependent survival but not chemotaxis is regulated by the FOXO downstream of Akt (Delgado-Martín et al. 2011). Furthermore, migration and ERK activation in response to CXCL12 in T cells is mediated by two distinct phosphatidylinositol-specific phospholipase C (PLC) isozymes (Kremer et al. 2011). For therapeutic modulation of chemokine signalling, the identification of downstream targets will be essential in order to modulate the different chemokine-induced functions selectively.

Apparently, the newly discovered receptor for CXCL12 and CXCL11, namely CXCR7, does not signal along classic G-protein-mediated pathways. In contrast to CXCR4, it is unable to induce Ca2+ elevation and, similar to other decoy chemokine receptors such as duffy antigen receptor for chemokines, D6 and CCX-CKR (Mantovani et al. 2006), it might function as a scavenger receptor for its cognate ligand to regulate the extracellular availability of CXCL12 (Naumann et al. 2010). Sanchez-Alcaniz et al. (2011) have recently shown that, upon interaction with CXCR4, CXCR7 on cortical interneurons prevents the normally rapid internalization of CXCR4 and the subsequent time- and energy-consuming biosynthesis of new molecules of the latter receptor. Thus, the responsiveness of CXCR4 to CXCL12 is retained and desensitization is prevented, even at high levels of this chemokine ligand in the developing cortex. The selective manipulation of receptors would therefore be one possibility for modulating chemokine action. Chemokine–receptor complexes are phosphorylated and endocytosed after signalling through clathrin-dependent pathways and, once internalized, might be either degraded or transported back to the cell membrane for re-expression (Rose et al. 2004).

Chemokine expression

Chemokine expression in the intact CNS

In an oversimplified view, either chemokines are inflammatory and expressed by cells of the immune system (leucocytes), glial, epithelial and endothelial cells and fibroblasts upon activation or they are homeostatic and expressed in specific locations in the absence of apparent activating stimuli.

Constitutive expression in the brain has been reported for CCL3 (Xia and Hyman 1999) and CCL2. The latter chemokine has been found in astrocytes, microglia, endothelial cells (Glabinski et al. 1996; Ma et al. 2002; Harkness et al. 2003) and in neurons including cortical motor neurons (Banisadr et al. 2005). CXCL12 is expressed in neurons of, for example, the subventricular zone, white matter and cerebellum, in blood vessels of the brain, along the dorsal corticospinal tract (dCST) and in meningeal cells of the spinal cord (Tysseling et al. 2011). CX3CL1 is expressed in healthy neurons (Harrison et al. 1998). CCL21 has been found to be transported in presynaptic processes to nerve terminals (de Jong et al. 2005). CCL19 and CCL21, on the other hand, are constitutively expressed in cerebrovascular endothelium and the choroid plexus (Lalor and Segal 2010). The corresponding receptors are also constitutively expressed in the CNS: CX3CR1 on microglia, CCR2 in neurons (brain, spinal cord; Gosselin et al. 2005), astrocytes, microglia, neural progenitor cells and microvascular endothelial cells (Banisadr et al. 2002, 2005; Coughlan et al. 2000; Gourmala et al. 1997; Stamatovic et al. 2005; Horuk et al. 1997). CXCR4 is constitutively expressed on cortical and hippocampal neurons, astrocytes, microglia and ependymal cells (Stumm et al. 2002; Banisadr et al. 2002) in the brain, on ependymal cells and, at low levels, on dCST axons (Tysseling et al. 2011; Opatz et al. 2009; Jaerve et al. 2011). CXCR7 expression has been observed in neurons of various brain regions (including cortical layers IV-V), astrocytes and endothelial cells (Schönemeier et al. 2008). Recently, we have also detected low levels of CXCR7 on dCST axons in the spinal cord (Opatz et al. 2009; Jaerve et al. 2011). The constitutive expression and homeostatic function of these chemokines and their receptors in brain and spinal cord have to be taken into account by targeting the chemokine system in the CNS.

Altered chemokine expression following CNS injury

In CNS injuries, chemokine and chemokine receptor expression is upregulated within minutes to hours depending on the severity of the injury, with expression being maintained for at least several days after trauma (Table 2). Chemokine gradients are argued to persist indefinitely or to reoccur periodically, in order to explain the observation of infiltrated for long periods, namely about 6 weeks for neutrophils and over 6 months for monocytes, in the lesioned spinal cord, despite their short lifespan of roughly 5 days and 2 months, respectively (Pillay et al. 2010; Hawthorne and Popovich 2011). The rapid and transient induction of several chemokines, for example, CCL2, CCL21 and CXCL10, in neurons has recently been shown to precede the induction of these chemokines in astrocytes, although the latter are considered the major source of chemokines after CNS injury (de Haas et al. 2007). This observation opens up some new perspectives with regard to targeting the chemokine system in the injured CNS.
Table 2

Chemokine expression in central nervous system (CNS) injury (SCI spinal cod injury, TBI traumatic brain injury, dpi days post injury, CSF cerebral spinal fluid, dCST dorsal corticospinal tract)

Chemokine

Injury

Peak

Source

Reference

CXCL1

SCI

3-12 h

Astrocytes (mRNA)

Pineau et al. 2009

6 h

mRNA (30× up)

McTigue et al. 1998

TBI

1 dpi

CSF

Helmy et al. 2011

6 h

Choroid plexus (mRNA, protein)

Szmydynger-Chodobska et al. 2009

Stroke

12 h, 3-6 h

Protein, serum

Yamasaki et al. 1995

24, 72 h

Monocyte, neutrophils (mRNA, >10× up)

Brait et al. 2010

CSF, not serum

Losy et al. 2005

Within 24 h to 7 dpi

Serum

Ormstad et al. 2011

CXCL2

SCI

12 h

Astrocytes (mRNA)

Pineau et al. 2009

24 h

mRNA

Ma et al. 2002

TBI

6 h

Choroid plexus (mRNA)

Szmydynger-Chodobska et al. 2009

Stroke

24, 72 h

Monocyte, neutrophils (mRNA >10× up)

Brait et al. 2010

CXCL3

SCI

15 min to 24 h

mRNA

Rice et al. 2007

TBI

6 h

Choroid plexus (mRNA)

Szmydynger-Chodobska et al. 2009

CXCL5

SCI

6-12 h

mRNA

McTigue et al. 1998

Stroke

Within 24 h

CSF, not serum

Zaremba et al. 2006a

CXCL8

SCI

1 to 4 h

mRNA

Spitzbarth et al. 2011

12 h

Protein

Hirose et al. 2000

Within 3 dpi

CSF

Kwon et al. 2010

TBI

24 h

mRNA

Stefini et al. 2008

1 dpi

CSF

Helmy et al. 2011

Stroke

24 h to 7 dpi

Serum; CSF

Grau et al. 2001; Tarkowski et al. 1997; Kostulas et al. 1998

1-7 dpi

Blood mononuclear cells (mRNA)

CXCL10

SCI

30 min to 24 h

mRNA

Rice et al. 2007

6 h

mRNA

Gonzalez et al. 2003

6 h

mRNA

McTigue et al. 1998

6 h, till 7 dpi

mRNA, cells near injury site

Lee et al. 2000

TBI

24 h to 7 dpi

mRNA (very low levels)

Stefini et al. 2008

2 dpi

CSF, higher than plasma

Helmy et al. 2011

Stroke

3 h, 6 h and 10-15 dpi

mRNA

Wang et al. 1998

CXCL12

SCI

7 dpi

mRNA

Takeuchi et al. 2007

In intact and injured; 14 dpi

dCST, meninges

Tysseling et al. 2011

Infiltrating monocyte (protein)

Stroke

7 dpi

Astrocytes, endothelial cells, neurons (protein)

Hill et al. 2004

CCL2

SCI

Starting <5 min; 4 dpi

Mainly astrocytes, also microglia, oligodendrocytes, neurons, blood vessels (mRNA);monocyte/microglia

Pineau et al. 2009

7 dpi (reduced)

mRNA

Jones et al. 2005

24 h

mRNA

Ma et al. 2002

15 min to 24 h

mRNA

Rice et al. 2007

Within 72 h

CSF

Kwon et al. 2010

1 h

Cells near injury site (mRNA)

Lee et al. 2000

TBI

14 h and 8 dpi; 1 dpi

mRNA; CSF higher than plasma

Stefini et al. 2008; Helmy et al. 2011

6-12 h

mRNA; endothelial cells,monocyte, some astrocytes (protein)

Berman et al. 1996

Stroke

2-3 dpi

Neurons, astrocytes (protein)

Che et al. 2001

6 h-2 dpi; 4 dpi

Astrocytes;monocyte/microglia (mRNA)

Gourmala et al. 1997

2 dpi

Protein

Yamagami et al. 1999

24 h

CSF, but not serum

Losy and Zaremba 2001; Zaremba et al. 2006b

CCL3

SCI

15 min to 24 h

mRNA

Rice et al. 2007

Within 1 h

Grey matter of spinal cord (microglia), at 4 dpi cellular infiltrate (mRNA)

Bartholdi and Schwab 1997; Lee et al. 2000

TBI

3.5 h and 8 dpi; 1 dpi

mRNA; CSF, higher than serum

Stefini et al. 2008; Helmy et al. 2011

Stroke

6 h

mRNA, astrocytes (protein)

Kim et al. 1995

4-6 h

Microglia, astrocytes (mRNA)

Takami et al. 1997

8 to 72 h

Monocyte/microglia (protein)

Cowell et al. 2002

CCL4

SCI

Within 1 h

Grey matter of spinal cord (microglia)

Bartholdi and Schwab 1997

4 dpi

Cellular infiltrate (mRNA)

TBI

3.5 h

mRNA

Stefini et al. 2008

1 dpi

CSF higher than plasma

Helmy et al. 2011

CCL5

SCI

90 min to 3 h

mRNA

Rice et al. 2007

7dpi to 21 dpi

mRNA

Jones et al. 2005

24 h

mRNA, microvascular endothelial cells

Benton et al. 2008

TBI

6 dpi

mRNA (very low levels)

Stefini et al. 2008

1 dpi

CSF

Helmy et al. 2011

Stroke

24 h

Blood-derived cells, resident cells (protein)

Terao et al. 2008

CCL7

SCI

24 h

mRNA

Ma et al. 2002

TBI

Within 5 dpi

CSF higher than plasma

Helmy et al. 2011

Stroke

12 h

mRNA

Wang et al. 1999

CCL8

SCI

12 to 24 h

mRNA

McTigue et al. 1998

CCL12

SCI

12, 24 h

mRNA

McTigue et al. 1998

CCL20

SCI

6-12 h

mRNA

McTigue et al. 1998

Stroke

24 h

mRNA, protein

Terao et al. 2009

CCL21

SCI

4 weeks

Dorsal horn neurons, thalamus (protein)

Zhao et al. 2007

Stroke

6 h to 4 dpi

Cortical neurons and not glia (mRNA)

Biber et al. 2001

CX3CL1

SCI

7 dpi (downregulated)

mRNA, protein

Donnelly et al. 2011

TBI

24 h

CSF, not serum, no changes in mouse model (mRNA, protein)

Rancan et al. 2004

The chemokine profiles after brain and spinal cord traumas show some similarities but also differences, which might be important if a therapeutic approach is transferred from one CNS injury type to another. Common is the high upregulation of CCL2. In head trauma and in cortical stab wound injury, CCL2, produced by activated astrocytes (Glabinski et al. 1996), macrophages and endothelial cells (Berman et al. 1996), is the most strongly transcribed chemokine in the brain (Stefini et al. 2008) and is also found in extracellular fluid (Helmy et al. 2011). The protein, which is expressed at a high basal level, is further elevated ∼90-fold (to 1333 pg/mg) following TBI (Dalgard et al. 2012). However, when only fold changes are considered, then the ∼236-fold induction of CXCL1 (to 170 pg/mg) is even stronger. Likewise, in SCI, CCL2 mRNA is upregulated 18-fold at 12 h, whereby CXCL1 is elevated as much as 30-fold at 6 h and CCL8 35-fold at 12 h (McTigue et al. 1998). Notably, the time course of CXCL1 and CCL2 regulation in injured spinal cord at 6 h and 12 h, respectively, is similar, whereas in injured brain, CXCL1 peaks much earlier (6 h) than CCL2 (12 h; Dalgard et al. 2012). CXCL10 mRNA is upregulated approximately seven-fold at 6 h after SCI (McTigue et al. 1998) and in brain extracellular fluid following TBI (Helmy et al. 2011). However, in another study of TBI, CXCL10 mRNA is almost absent (Stefini et al. 2008). CCL20 mRNA is elevated eleven-fold at 6-12 h and CCL3 five-fold at 24 h following SCI (McTigue et al. 1998). In contrast to SCI, CCL20 peaks only at a later time point at 24 h in TBI (∼55-fold, to 55 pg/mg; Dalgard et al. 2012).

The monitoring of chemokine levels could enhance the precision of the assessment of injury severity, a major challenge in the management of SCI. Levels of CCL2 and CXCL8 in cerebrospinal fluid (CSF) at 24 h post-SCI accurately predict the American Spinal Injury Association (ASIA) grade for acutely injured patients and the prognosis of segmental motor recovery at 6 months can be more precisely predicted than with the patients' baseline ASIA grade (Kwon et al. 2010). Interestingly, elevated CCL2 levels in blood samples of ischaemic stroke patients correlate well with the clinical outcome as early as a few hours after the onset of symptoms (Worthmann et al. 2010).

The expression of several chemokines can be regulated by levels of oxygen and cytokines. For example, CCL2 production is induced by changes in oxygen levels in rat peripheral blood mononuclear cells (Reale et al. 2003). Treatment with lipopolysaccharide, interferon, interleukin-1 beta (IL-1β), colony-stimulating factor-1, transforming growth factor-β and tumour necrosis factor-α (TNFα) can induce the expression of several chemokines, as has been shown for CCL2 in various cell types in vivo and in vitro (Harkness et al. 2003; Hurwitz et al. 1995; Thibeault et al. 2001).

Chemokines and their receptors as therapeutic targets in CNS injury

CXCL1-CXCL8: CXCR2 ligands

Chemokines CXCL1 to CXCL8 recruit neutrophils, which express the CXCR2 receptor, to the site of CNS injury. Neutrophil infiltration is generally associated with the propagation of secondary tissue damage. In agreement with this, the recruitment of neutrophils to the injury site, tissue damage and neuronal apoptosis are all significantly decreased in CXCR2-deficient mice suffering from closed head injury but no significant functional effects have been observed (Semple et al. 2010). Cutaneous wound healing in CXCR2 knockout mice is, however, impaired as neutrophil and monocyte recruitment is defective, epithelialization (wound closure) is delayed and neovascularization is decreased (Devalaraja et al. 2000). This indicates that the blocking of the CXCR2-mediated recruitment of neutrophils has significant drawbacks as a therapeutic target. Moreover, the depletion of Gr-1+ neutrophils by the administration of anti-Gr-1 antibody reduces wound healing and worsens the functional recovery of SCI (Stirling et al. 2009) implying that neutrophils can also exert beneficial effects. The controversial role of neutrophils has thus prompted studies to explore the mechanisms regulating the production of chemokines, especially CXCL1 and CXCL2, which attract neutrophils into the injured CNS. The IL-1 receptor/MyD88 in astrocytes (Pineau et al. 2009) and the inhibitor of kappa B (IκB) kinase-β in neutrophils (Kang et al. 2011) have been shown to be critical for the expression of CXCL1 following SCI. Astrocyte-specific (Pineau et al. 2009) and myeloid cell-specific (Kang et al. 2011) knockout of these signals via the abolishment of the downstream activation of nuclear factor kappa B leads to reduced CXCL1 expression and thus compromises the entry of neutrophils and the recruitment of type I “inflammatory” monocytes to sites of SCI leading to improved conditions for recovery. On the other hand, CXCL1/CXCR2 signalling has been shown to be beneficial in remyelination and oligodendrocyte precursor cell (OPC) biology. However, the underlying mechanisms remain controversial. Conditional overexpression of CXCL1 in astrocytes in a mouse model of multiple sclerosis (MS) leads to positive responses regarding neuroprotection and remyelination (Omari et al. 2009). This observation is further supported by the finding that the neutralization of the CXCL1 receptor, CXCR2, delayed clinical recovery in a mouse model for viral-induced demyelination through the induction of OPC apoptosis (Hosking et al. 2010). In contrast, in an EAE (experimental allergic encephalomyelitis) model Kerstetter et al. (2009) have shown that the inhibition of CXCR2 signalling promotes functional recovery by enhancing the differentiation of myelin-producing cells. Omari et al. (2006) have proposed that OPCs, which constitutively express CXCR2 are recruited to demyelination/lesion areas by CXCL1. The latter chemokine is released by reactive astrocytes and is known to halt OPC differentiation in vivo (Tsai et al. 2002). However, this concept has not been supported by cuprizone-induced demyelination in CXCR2-deficient mice, as no impaired OPC recruitment has been observed (Lindner et al. 2008). The controversial role of CXCL1/CXCR2 remains to be investigated further. However, it draws attention to the finding that chemokines have potentially distinct effects on different types of cells (here neutrophils or OPC), clearly pointing to the need of cell-type-specific targeting.

CXCL4, as a strong chemoattractant for neutrophils and fibroblasts, has been proposed to play a role in inflammation and wound repair (Eisman et al. 1990). It efficiently neutralizes heparin-like molecules on the endothelial surface of blood vessels, thereby inhibiting local antithrombin III activity and promoting coagulation. Under neurodegenerative conditions in entorhinal cortex slice cultures treated with N-methyl-D-aspartate (NMDA), the brain microglial cells have been shown to migrate towards CXCL4 via mechanisms involving the CXCR3b receptor. Furthermore, CXCL4 attenuates lipopolysaccharide-induced microglial phagocytosis and nitric oxide production in microglia and BV-2 cells (de Jong et al. 2008).

CXCL8 is the most intensely investigated pro-inflammatory human analogue of rat CXCL1 recruiting neutrophils. The blocking of its action has been shown to be beneficial in CNS injury. A CXCL8-neutralizing antibody is able to reduce brain oedema and cerebral infarct size (Matsumoto et al. 1997). Inhibition of CXCL8 receptor by reparixin causes the same beneficial effect in rats suffering from permanent and transient cerebral ischaemia resulting in the attenuation of neurological deficits (Villa et al. 2007). In the clamp SCI, the therapeutic steroid analogue lazaroid U-74389 G, which inhibits lipid peroxidation but does not have the side effects of steroid therapy (Kavanagh and Kam 2001), has been shown to reduce the production of systemic and spinal CXCL8 (Kunihara et al. 2000). CXCL8 has been demonstrated to increase the expression of matrix metalloproteinases, cell cycle and pro-apoptotic proteins and cell death in cultured neurons (Thirumangalakudi et al. 2007). On the other hand, some neuroprotective effects of CXCL8 cannot be excluded. For example, this chemokine has been shown to recruit human neural precursor cells across brain endothelial cells (Rainey-Barger et al. 2010). Furthermore, CXCL8, CXCL2 or CXCL1 are reported to protect hippocampal neurons from beta-amyloid (1-42)-induced cell death (Watson and Fan 2005).

CXCL9, CXCL10 and CXCL11: CXCR3 ligands

CXCR3 ligands are potent chemoattractants for T lymphocytes and natural killer cells, and their expression can be induced by interferon-gamma (Shurin et al. 2007). The location of CXCR3 on the surface of endothelial cells suggests its role in maintaining the cytokine gradient in the region of CNS inflammation (Ghersa et al. 2002). Blocking CXCR3 might thus be beneficial in neuroinflammation.

Administration of a neutralizing antibody against CXCL10 in spinal cord-injured mice reduces neuroinflammation (Gonzalez et al. 2003) and tissue damage (Glaser et al. 2004). A decreased infiltration of monocytes (lacking CXCR3) has been observed that corresponds to the reduced infiltration of pathogenic Th1 T cells in SCI and in MS and EAE (Balashov et al. 1999; Sørensen et al. 1999; Jones et al. 2005). Th1 T cells are known to attract monocytes by secreting CCL5 (Fransen et al. 2000). In a follow-up study, Glaser et al. (2006) have also found that this anti-CXCL10 treatment remarkably enhances the sprouting of corticospinal axons and increases their number caudal to the injury site. This finding could help to explain the previous observation that locomotor recovery in SCI is significantly improved under anti-inflammatory conditions (Gonzalez et al. 2003). CXCL10 is known as an anti-angiogenetic factor (Strieter et al. 1995a, b, 1995c; Angiolillo et al. 1995; Luster et al. 1995; Bodnar et al. 2009) and its neutralization almost doubles the number of newly formed blood vessels in regenerating spinal cord (Glaser et al. 2004) indicating the important modulatory role of chemokines in neovascularization after CNS injury.

In addition to its known role in T cell recruitment, CXCL10 is expressed in neurons in response to brain injury and has been shown to activate and direct microglia to the lesion site. In CXCR3 knock-out mice, microglial migration into areas of axonal degeneration is impaired and, moreover, denervated dendrites are not lost following lesion. Unfortunately, functional consequences following brain or spinal cord lesions have not been studied in these mouse mutants (Rappert et al. 2004).

The expression of other ligands of CXCR3, CXCL9 and CXCL11 seems not to be significantly regulated by CNS pathologies and might not be essential for the initial migration of CXCR3-bearing cells (McColl et al. 2004).

CXCL12: ligand for CXCR4 and CXCR7

CXCL12/SDF-1 has extraordinary features among the chemokine family of proteins. CXCL12 plays an extremely important role not only in CNS development but also in CNS homeostasis (neurogenesis, neuromodulation) and CNS injury. We have shown that the infusion of this ligand into the lesion site after dorsal hemisection in rat spinal cord promotes the sprouting of the dCST rostral to the lesion site. Fibres grow into the grey and white matter of the spinal cord (Opatz et al. 2009). In addition to dCST, the rostral sprouting of serotonergic (5-HT) and tyrosine hydroxylase (TH)-positive fibres increase in both young and aged rats (Jaerve et al. 2011). The mechanism of action for dCST sprouting probably involves signalling via CXCR4 and/or CXCR7, as both receptors can be detected on CST axons. However, direct evidence for this hypothesis remains to be obtained. The sprouting of 5-HT and TH-positive fibres, on the other hand, could be attributable to indirect mechanisms, as these fibres do not express detectable levels of CXCL12 receptors.

CXCL12 appears to be one of the most potent coordinators of stem and progenitor cell migration; it recruits mesenchymal stem cells (Li et al. 2009), intracerebrally transplanted bone-marrow-derived mesenchymal stem cells (Wang et al. 2008), monocytes (Hill et al. 2004), endothelial cell progenitors, which are important participants of neovascularization (Bogoslovsky et al. 2011) and neuroblasts (Cui et al. 2007; Imitola et al. 2004; Robin et al. 2006; Fan et al. 2010) through signalling via CXCR4. In addition, the upregulation of CXCL12 and CXCR4 in murine olfactory ensheathing cells has been shown to promote axonal regeneration in vitro and in vivo (Shyu et al. 2008a). In stroke, CXCL12 has been demonstrated to moblize bone-marrow-derived cells stem cells, promote angiogenesis and protect neurons (Shyu et al. 2008b). At 1 week after SCI in mice, elevated levels of CXCL12 correlate well with the accumulation of neuronal progenitor cells injected at the injury site (Takeuchi et al. 2007). Ependymal cells in the spinal cord, which constitute the endogenous stem cell pool, express high levels of CXCR4 (Tysseling et al. 2011; Jaerve et al. 2011) and are, therefore, probably guided to the lesion site by its ligand. CXCL12 is also involved in neurotransmission and modulates neuronal activity, as GABAergic inputs of immature neurons at the infarct boundary are enhanced by this chemokine (Bhattacharyya et al. 2008). Moreover, survival-supporting effects for neural progenitor cells mediated via the second CXCL12 receptor, CXCR7, have been reported (Bakondi et al. 2011). Signalling by binding to CXCR4 (Patel et al. 2010) and to CXCR7 (Gottle et al. 2010) has been implicated in OPC differentiation and remyelination. Probably, signalling via both receptors can activate parallel and protective pathways in ischaemic stroke. The developmental regulation of neuronal migration and axonal pathfinding by CXCL12/CXCR4 also suggest a role of this ligand-receptor interaction in regeneration in the adult brain (Stumm and Höllt 2007). Surprisingly, CXCL12 reduces the senescence of an endothelial progenitor subpopulation through telomerase activation and telomere elongation (Zheng et al. 2010). The findings described here imply that numerous cell biological aspects of this interesting chemokine remain to be discovered in the future.

CXCL13, CXCL14 and CXCL16

CXCL13 levels are often elevated in the inflamed CNS and shown to recruit B cells in animal models of infectious and inflammatory demyelinating disease (Rainey-Barger et al. 2010). However, B cell depletion can lead to progressive multifocal leucoencephalopathy highlighting the importance of B cell surveillance of the CNS. As reported above for CXCL8, enhanced levels of CXCL13 have been found in active demyelination areas and presumably assist systemically injected neural precursor cells across the brain endothelium and favour functional recovery in animal models of MS (Weiss et al. 2010; Bagaeva et al. 2006). CXCL14 is also involved in myelination, is up-regulated in the sciatic nerve of a mouse model of Charcot-Marie-Tooth disease type 1A and alters myelin gene expression in cultured Schwann cells (Barbaria et al. 2009).

Chemokine CXCL16 levels were reported to be dramatically elevated in EAE, being up to 10-fold higher in the spinal cord as compared with the brain (Kim et al. 2009). This rise could be attributable to the invasion of inflammatory cells into the spinal cord (Matloubian et al. 2000; Fukumoto et al. 2004; Ludwig et al. 2005), cells that, in addition to the astrocytes and endothelial cells, produce CXCL16. Interestingly, when attracted by CXCL16, the infiltration of neutrophil and monocytes into injured muscle promotes muscle regeneration and suppresses fibrosis (Zhang et al. 2009). The role of this chemokine in CNS injury might be worthy of more detailed investigation in the future.

MCP family: CCL2, CCL7, CCL8, CCL12 and CCL13

Each member of the monocyte chemotactic protein (MCP) family (designated as MCP-1-5, respectively) attracts a different subset of leucocytes after binding with various affinities to several receptors (Gouwy et al. 2004). Although CCL2, CCL7 and CCL8 signal through the CCR2 receptor, CCL2 is the most potent in activating signal transduction pathways that lead to monocyte transmigration (Sozzani et al. 1994) and is probably the most studied chemokine.

CCL2 and its receptor CCR2 are considered pro-inflammatory. There is strong evidence for its detrimental effect. Over-expression of CCL2 increases brain infarct volume (Chen et al. 2003b) and exacerbates responses to brain injury (Muessel et al. 2002) and mouse mutants deficient in the genes for CCL2 or CCR2 show decreased inflammatory infiltration and infarct size (Hughes et al. 2002; Dimitrijevic et al. 2007). Similar results have been obtained with chemokine receptor-antagonist treatment (Minami and Satoh 2003) or with anti-CCL2 antibody, which decreases the permeability of the blood-brain barrier (BBB) after ischaemia/reperfusion (Xia and Sui 2009). In CCR2-deficient mice, the number of infiltrating monocytes in the lesion epicentre, myelin degradation and the expression of CCR1 and CCR5 are decreased at 1 week after SCI (Ma et al. 2002). In agreement with this is the observation that CCL2 mainly attracts the monocyte “inflammatory population” and not Gr1-/Ly6ClowCCR2-CX3CR1high cells, which have been shown to be neuroprotective (Woollard and Geissmann 2010; Weber et al. 2008).

Nonetheless, the targeting of CCL2 should take into account its constitutive expression and homeostatic role in neurons throughout the rat brain. For example, in vivo, CCL2 induces the excitability of dopaminergic neurons, dopamine release and locomotor activity in rats (Guyon et al. 2009). Moreover, whereas CCL2 first worsens the effect of stroke, it has been suggested later to promote neurogenesis by recruiting bone-marrow stromal cells to the ischaemic brain (Wang et al. 2002) and newly formed neuroblasts from neurogenic regions (the subventricular zone and the posterior peri-ventricular region) to damaged cerebral areas after focal ischaemia (Yan et al. 2007). The suppression of the rapid and transient induction of CCL2 in neurons after ischaemia and axonal injury might be beneficial in order to block neuronal apoptosis (Hinojosa et al. 2011).

CCL3, CCL4 and CCL5

CCL3 recruits monocytes and microglial cells in the injured brain (Cowell et al. 2002; Deshmane et al. 2009) via CCR1 and CCR5 (Cheng et al. 2001). CCL3 is also a potent neutrophil chemoattractant in mice and humans (Reichel et al. 2009) in the presence of other pro-inflammatory molecules, such as TNF-α or insulin (Montecucco et al. 2008a). Intra-cerebro-ventricular injection of mouse CCL3 increases the infarct volume (Takami et al. 1997).

On the other hand, CCL3 levels show an inverse correlation with functional deficits in stroke (Zaremba et al. 2006b). CCL3 has been proposed as a potential anti-apoptotic agent in stroke (Hau et al. 2008). Indeed, CCL3 has been demonstrated to promote the migration of cells from human umbilical cord blood into ischaemic regions (Jiang et al. 2008). CCL3 together with its receptor CCR1 probably plays an important role in the subpial white matter neural stem- and progenitor-cell niche after SCI, as increased immunoreactivity for CCL3 (and CCL2, CXCL12) colocalizes with astroglial and oligodendroglial markers and with nestin, 3CB2 and brain lipid-binding protein in cells morphologically resembling radial glia (Knerlich-Lukoschus et al. 2010).

CCL4 is involved in monocyte migration (Montecucco et al. 2008b). It increases reactive oxygen species production and the adhesion of THP-1 cells (human acute monocytic leukaemia cell line) to human umbilical vein endothelial cells in vitro (Tatara et al. 2009). Elevated serum levels have been shown to predict stroke and cardiovascular events independently in hypertensive patients (Tatara et al. 2009).

The detrimental or beneficial role of CCL5 and its receptors CCR1, CCR3 and CCR5 (Braunersreuther et al. 2007) in the damaged CNS remains controversial. In CCL5 knockout mice or in wild-type mice receiving CCL5-deficient bone marrow, the infarct volumes and BBB permeability are smaller than those in controls (Terao et al. 2008). On the other hand, CCL5 has been reported to increase cerebral damage through the secondary induction of other potent pro-inflammatory cytokines such as IL-6 (Shahrara et al. 2006). Not only CCL5 but also CCR5 are upregulated in astrocytes, microglia, endothelial cells and neurons following CNS injury (Spleiss et al. 1998; Tripathy et al. 2010a, b) and have been proposed to mediate neuronal protection and survival (Rostène et al. 2007). In vitro evidence further suggests a benefical role of CCL5 in mediating neuronal protection and survival. For example, the treatment of primary cortical neuronal cultures with CCL5 increases neuronal survival and protects against the neurotoxic agent thrombin and sodium nitroprusside (Tripathy et al. 2010a, b). CCL5 also protects mixed cultures of neurons and astrocytes from excitotoxic NMDA-induced apoptosis (Eugenin et al. 2003). This agrees with the finding that numerous CCL5-responsive genes in cultured neurons are involved in the regulation of neuronal survival and differentiation (Valerio et al. 2004).

Moreover, mice carrying mutations of CCR5, the receptor for chemokines CCL3, CCL4, CCL5, CCL8, CCL11, CCL14 and CCL16, show larger cerebral infarct size, with increased neuronal death and neutrophil infiltration associated with behavioural deficits (Sorce et al. 2010). The mechanism by which CCR5 reduces secondary damage after CNS injury is not known, although both direct effects on neurons and avoidance of excessive microglial activation have been considered.

CCL19 and CCL21: ligands of CCR7; CCL20: ligand of CCR6

CCL19/CCR7 has been implicated in the immune surveillance of the CNS by lymphocytes (Lalor and Segal 2010; Shannon et al. 2010). CCL21 is rapidly upregulated in cortical neurons upon ischaemic injury and glutamate-mediated excitotoxicity, is transported in secretory granules along axons to presynaptic structures and, in this way, activates remote microglia expressing CXCR3 (Biber et al. 2001, 2011; for the role of CXCR3 in CNS inflammatory reactions, see above).

CCL20–CCR6 interactions attract inflammatory monocytes and activate microglia and thus probably contribute to neuroinflammation following brain injury (Terao et al. 2009). Indeed, hypothermia has been shown to be effective in suppressing these effects, as is neutralization with MAB540 antibody to reduce the ischaemic area (Terao et al. 2009).

CX3CL1: ligand of CX3CR1

CX3CL1 is expressed on neurons, whereas CX3CR1 is found on microglia and the interaction between them is thought to maintain microglia in a resting state, thereby suppressing the neurotoxic and phagocytic activity of these cells in the CNS (Cardona et al. 2006). The blockade of CX3CR1 signalling leads to better anatomical and functional recovery from SCI in mice (Donnelly et al. 2011). The production of inflammatory cytokines by spinal microglia and monocyte-derived monocytes is reduced, as is the number of pro-inflammatory monocytes in the injury site. Knocking out the CX3CR1 gene attenuates hyperalgesia and allodynia in a modality-dependent fashion (Staniland et al. 2010). In focal cerebral ischaemia (Denes et al. 2008) or cerebral ischaemia-reperfusion models of stroke (Soriano et al. 2002), mice lacking the fractalkine receptor are less susceptible to damage. However, in Parkinson’s disease and amyotrophic lateral sclerosis models, the lack of CX3CR1 exacerbates neuronal loss (Cardona et al. 2006), indicating that the result depends on the neurological condition.

Perspectives

Only a limited number of therapeutics targeting the chemokine system has been approved for clinical use, among which is the CXCR4 antagonist mozobil for stimulating the mobilization of haematopoietic bone-marrow stem cells to the bloodstream and the inhibitor of CCR5 maraviroc for the prevention of infection by human immunodeficiency virus (HIV). Numerous potential therapeutics are under trial for HIV, MS and rheumatoid arthritis (but none for traumatic CNS injuries or stroke) and no anti-inflammatory agents have been tested in TBI (Proudfoot et al. 2010; Mirabelli-Badenier et al. 2011).

Neuroinflammation

Preclinical therapeutic appoaches include small molecule inhibitors or neutralizing antibodies and modified chemokines (Chevigne et al. 2011). For example, a broad spectrum inhibitor of CC and CXC chemokines, NR58–3.14.3, a retroinverso-analogue of a 12-mer peptide, reduces the inflammatory response and the lesion size and, consequently, improves neurological function in a rat model of cerebral ischaemia (Beech et al. 2001).

An example of a potent naturally available broad chemokine antagonist is a chemokine analogue peptide encoded by Kaposi-sarcoma-associated Herpes virus, namely viral monocyte inflammatory protein-II (vMIP-II), which protects the brain against focal cerebral ischaemia in mice (Takami et al. 2001). Infusion of vMIPII suppresses gliotic reactions and reduces neuronal damage and apoptosis. Moreover, vMIPII largely prevents inflammatory cellular infiltration following SCI (Ghirnikar et al. 2000, 2001).

Rather than applying such broad-range agents, most current research efforts are directed to defining and identifying distinct cell-type-specific regulation and function and responsible downstream signalling as potentially more attractive targets to circumvent off-target effects, such as the inhibition of T cell recruitment but not function. Chemokines have also been recognized and appreciated selectively to recruit subpopulations of cells with different properties. The heterogeneity of monocytes with pro-inflammatory or regeneration-promoting characteristics and the interchange between their functional states (David and Kroner 2011) might explain some controversial results obtained with anti-inflammatory strategies blocking the action of chemokines (e.g. CCL3, CCL5). In the study of Donnelly et al. (2011), blockade of CX3CR1 achieved a better anatomical and functional recovery from SCI in mice (see above), because recruitment or maturation of monocyte-derived macrophages of a neurotoxic phenotype (Ly6Clo/iNOS+/MHCII+/CD11c-) was reduced, as was their ability to produce inflammatory cytokines and oxidative metabolites; instead, more regeneration-supporting CCR2+/Ly6Chi/MHCII-/CD11c+ monocytes were recruited.

Various types of monocytes also produce and release complex patterns of different chemokines. For example, M1 macrophages, which can induce neuron death in vitro, produce CXCL9, CXCL10, CXCL11, CXCL13, CCL8, CCL15, CCL19 and CCL20. M2 subpopulations, M2a and M2c, with anti-inflammatory and reparative properties, produce CCL13, CCL14, CCL17, CCL18, CCL22, CCL23, CCL24 and CCL26 and CCL16, CCL18 and CXCL13, respectively. The M2b type, which produces beneficial IL-10 but also pro-inflammatory molecules IL-12, TNFα, IL-1β and IL-6, synthesizes CCL1, CCL20, CXCL1, CXCL2 and CXCL3 (David and Kroner 2011). Information about the expression of the corresponding chemokine receptors by the different macrophage subtypes would be valuable in order to block their action.

Tissue repair

Chemokine functions other than in neuroinflammation are of potential interest for therapeutic intervention (Table 3). CNS repair processes are promoted by increased proliferation and migration of neural and other progenitor cells as mediated through chemokines such as CXCL12/CXCR4, CCL3/CCR1, CXCL8 and CXCL13. Neovascularization could be supported by the recruitment of endothelial progenitor cells that are attracted by CXCL12. Angiogenetic properties have also reported for CCL2, CCL11, CCL16 and CXCL1-8, whereas CXCL9-CXCL11 are angiostatic (Keeley et al. 2010).
Table 3

Chemokines or their receptors as targets in CNS injury (EAE experimental allergic encephalomyelitis, TBI traumatic brain injury)

Function

Chemokine

Intervention

Injury

Reference

Axon sprouting

CXCL12

Infusion

SCI

Opatz et al. 2009; Jaerve et al. 2011

CXCL10

Neutralization

SCI

Glaser et al. 2006

Remyelination

CXCL12

Stimulation

EAE

Gottle et al. 2010

CXCL1

Cond. over-expression

EAE

Omari et al. 2009

Demyelination

CXCR3

Deletion

EAE

Müller et al. 2007

Stem/progenitor cell recruitment

CXCL12

Stimulation

Stroke

Cui et al. 2007; Imitola et al. 2004; Robin et al. 2006; Fan et al. 2010; Wang et al. 2008; Li et al. 2009

CCL2

Stimulation

Stroke

Yan et al. 2007

CCL3

Stimulation

Stroke

Hughes et al. 2002

Jiang et al. 2008

CXCL8

Stimulation

EAE

Weiss et al. 2010

CXCL13

Neovascularization

CXCL10

Neutralization

SCI

Glaser et al. 2004

Reduced tissue damage

CXCL10

Neutralization

SCI

Glaser et al. 2004

CX3CL1

Deletion

SCI

Donnelly et al. 2011

CXCL8

Neutralization

Stroke

Matsumoto et al. 1997

CXCR2

Deletion

TBI

Semple et al. 2010

CXCR2

Inhibiton

Stroke

Villa et al. 2007

CCL2

Deletion

Stroke

Hughes et al. 2002

Neutralization

Xia and Sui 2009

CCR2

Deletion

Stroke

Dimitrijevic et al. 2007

Inhibition

Minami and Satoh 2003

CCL20

Neutralization

Stroke

Terao et al. 2009

CX3CR1

Deletion

Stroke

Soriano et al. 2002

Denes et al. 2008

We have shown recently that the infusion of CXCL12 enhances axonal sprouting following SCI (Opatz et al. 2009; Jaerve et al. 2011). CXCL12 is able to induce neurite arborization by elevating cAMP levels (Chalasani et al. 2003; Pujol et al. 2005), which activates the expression of regeneration-supporting genes such as Arginase I, which is involved in the synthesis of polyamines (Cai et al. 2002). However, the exact mechanism of CXCL12-induced axon sprouting in SCI remains to be determined.

Remyelination by oligodendrocytes is also regulated by some chemokines. CXCL12 has been shown to induce oligodendrocyte proliferation, maturation and remyelination (Gottle et al. 2010). Moreover, CXCL1/CXCR2 signalling might play a role in oligodendrocyte proliferation and recruitment to demyelinated areas. Whereas CXCL14 alters myelin gene expression in cultured Schwann cells (Barbaria et al. 2009), its role in the CNS is still unknown.

Neuropathic pain

Some chemokines have also been implicated as potential candidates for neuropathic pain following peripheral nerve injury (Kiguchi et al. 2010) and thus might also be potential targets following CNS injury. CCL2, CCL3 and CXCL12 and their receptors are involved in nociceptive processing in the dorsal horn and in the brain (Knerlich-Lukoschus et al. 2010; Van Steenwinckel et al. 2011). Thus, antagonists of the CCL2 receptor are promising agents for treating neuropathic pain, as pain-transmission effects can be prevented by the administration of INCB3344 or ERK inhibitor (PD98059). As one mechanism for neuropathic pain, the upregulation of P2X4 expression in microglia following stimulation through neuronally derived CCL21 has been demonstrated (Biber et al. 2011). CX3CL1 obviously also plays a role, as the deficiency of its receptor attenuates hyperalgesia and allodynia in a modality-dependent fashion (Staniland et al. 2010).

Several chemokines are apparently involved in a range of different supportive and/or adverse cellular processes such as axon sprouting vs neuropathic pain, or neural precursor/stem cell attraction vs tumorigenic potential. The complexity of the heterogeneous functions of chemokines could potentially limit their clinical application, unless the underlying cell signalling cascades and appropriate targets for selective pharmacological intervention have been identified. However, even if the latter are known, additional factors such as aging might interfere with the therapeutic efficacy of chemokines.

Impact of ageing on chemokine function

Evidence has been presented that age has an impact on chemokine induction and its functional effect following CNS trauma. For example, in old rats with spinal cord injuries, the down-modulation of inflammatory responses is reported to be disturbed resulting in persistent neutrophil infiltrates (Genovese et al. 2006). This could be attributable to enhanced neutrophil attractor CXCL1 levels in old animals. Indeed, the expression of CXCL1, which has been found to be produced mainly by activated microglia (see above), is significantly higher in adult (10 weeks) compared with young (4 weeks) mice at 3 h post-SCI and correlates well with a higher infiltration of neutrophils (Kumamaru et al. 2011). In TBI, exclusive induction of CXCL1 upon IL-1β treatment in young but not adult rat brain has been attributed to the marked BBB breakdown and neutrophil response in young compared with adult rats (Campbell et al. 2002). Treatment with anti-CXCL1 antibody attenuates these effects in young rats.

Circulating levels of cytokines are influenced by aging. With increased age, reduced levels of CCL2 (Kim et al. 2011; Tripathy et al. 2010a, b; Reale et al. 2003) and CX3CL1 (Wynne et al. 2010) have been reported, whereas the levels of most other chemokines have been shown to be increased. CCL2 (Seidler et al. 2010; Inadera et al. 1999; Antonelli et al. 2006), CCL3, CCL4, CCL5 (Felzien et al. 2001; Chen et al. 2003a), CXCL1 (Fimmel et al. 2007), CXCL8 (Baune et al. 2008), CXCL9 (Njemini et al. 2007; Shurin et al. 2007), CXCL10 (Antonelli et al. 2006; Shurin et al. 2007) and CXCL11 (Shurin et al. 2007) are elevated at a higher age. Reduced CX3CL1 levels in aged brain are associated with microglial activation during aging and the suppression of neurogenesis (Bachstetter et al. 2009). Administration of CX3CL1 reverses the decline in age-associated neurogenesis. In contrast, increased serum levels of CXCL8 are likely to cause poor memory and motor performance (Baune et al. 2008). Interestingly, reduced levels of CXCL12 among other hypoxia-responsive cytokines are responsible for impaired wound healing in aged animals, as neovascularization is less efficient (Loh et al. 2009). Moreover, we have recently been able to demonstrate that, despite reduced axon sprouting in aged rats, intrathecal infusion of CXCL12 is still able to significantly increase axonal sprouting activity after SCI (Jaerve et al. 2011).

Concluding remarks

A huge amount of information is available concerning the expression, regulation, signalling and action of chemokines. Several therapeutic paradigms with chemokines in experimental CNS trauma models of SCI, TBI and stroke have gained significant improvements in axon growth and tissue preservation. The finding that chemokines act in a similar manner in these pathologies might facilitate the development of therapies in CNS repair. For example, the chemoattraction function of chemokines is an interesting feature for therapeutic intervention, because the whole neuroinflammation scenario can be manipulated, starting with the suppression or changes in the time course of infiltration of inflammatory cells or their various invading subtypes. Moreover, precursor/stem cells that promote regenerative processes can be called to the scene. Importantly, at least one of the chemokines (CXCL12) can directly stimulate axon sprouting of injured CNS neurons. The success of chemokines in clinical transfer, however, depends on the elucidation of their cell-subtype-specific action and cellular signalling pathways in CNS injuries in order to identify selective drug targets.

Copyright information

© Springer-Verlag 2012