Skip to main content
NCBI home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2009 Feb 23.
Published in final edited form as: Restor Neurol Neurosci. 2008;26(2-3):175–182.

Axon regeneration after spinal cord injury: Insight from genetically modified mouse models

Jae K Lee 1, Binhai Zheng 1,*
PMCID: PMC2646464  NIHMSID: NIHMS93458  PMID: 18820409

Abstract

The use of genetically modified mice to study axon regeneration after spinal cord injury has served as a useful in vivo model for both loss-of-function and gain-of-function analysis of candidate proteins. This review discusses the impact of genetically modified mice on axon regeneration after spinal cord injury in the context of axon growth inhibition by myelin, the glial scar, and chemorepellent molecules. We also discuss the use of mice which transgenically express fluorescent proteins in specific axons for increasing our understanding of how spinal cord axons behave after injury.

1. Introduction

During the past decade, there has been an intensive effort to understand why axons fail to regenerate in the adult mammalian central nervous system (CNS) in the hope of finding an effective therapeutic strategy to promote regeneration after CNS pathology such as spinal cord injury (SCI). We now understand that in addition to the limited endogenous ability of CNS neurons to regenerate, there are numerous inhibitory molecules in the CNS environment that may prevent regeneration. The use of genetically modified mouse models has served as a useful in vivo proof-of-principle to test promising in vitro and pharmacological profiles of various potentially inhibitory molecules. The results from mutant mouse models have provided significant insights into the role of these inhibitory molecules, and have often contradicted long-standing theories on axon regeneration in the CNS. Genetically modified mice include knockout mice for loss of function analysis as well as transgenic mice that carry exogenous genes for gain of function analysis and for genetic tract tracing. This review considers the impact that both types of mouse models have had on the advancement of axon regeneration after spinal cord injury.

2. Myelin inhibition

In 1981, Aguayo and colleagues demonstrated that CNS axons can grow through a peripheral nerve bridge, suggesting that CNS neurons are capable of regenerating axons but the environment in the CNS differs from that in the peripheral nervous system (PNS) in their capacity to support regeneration (David and Aguayo, 1981). In the late 80’s and early 90’s, Schwab and colleagues showed that there are inhibitory molecules present in CNS myelin (Savio and Schwab, 1989) and developed a monoclonal antibody, called IN-1, that binds to the inhibitory molecule(s) (Caroni and Schwab, 1988). Rats treated with IN-1 after a dorsal hemisection model of SCI showed significant regeneration of corticospinal axons (Schnell and Schwab, 1990). This was the first study to show significant regeneration of adult mammalian CNS axons in a spinal cord injury model by using a pharmacological agent, and this promising result opened the door to the field of axon regeneration. This inhibitory molecule, termed Nogo, was later identified as a member of the Reticulon family (GrandPre et al., 2000).

Upon identification of Nogo, three independent labs generated Nogo knockout mice to be tested in a model of spinal cord injury. This intensive synchronized endeavor was published as three companion papers each reporting unexpectedly varying results. Contrary to the expected role of Nogo, Tessier-Lavigne and colleagues failed to observe any significant regeneration of CST fibers after a dorsal hemisection in Nogo deficient mice (Zheng et al., 2003). Strittmatter and colleagues, on the other hand, reported robust regeneration of CST fibers in a Nogo mutant (Kim et al., 2003), while Schwab and colleagues demonstrated a tendency for increased regeneration caudal to the lesion that failed to reach statistical significance (Simonen et al., 2003). A detailed discussion of these contradictory results is beyond the scope of this review and has been published previously (Teng and Tang, 2005; Zheng et al., 2006). Recent studies have suggested background mouse strain differences (Dimou et al., 2006) as well as methodological issues (Steward et al., 2007) as possible explanations for these discrepancies, but at the very least, these genetic studies raise questions about whether blocking Nogo alone can promote significant axon regeneration after SCI.

In addition to Nogo, myelin associated glycoprotein (MAG) and oligodendrocyte myelin glycoprotein (OMgp) are the two other canonical myelin inhibitors. Although both MAG and OMgp show neurite growth inhibiting effects in vitro (Mukhopadhyay et al., 1994; McKerracher et al., 1994; Wang et al., 2002; Kottis et al., 2002), MAG knockout mice failed to show any significant axon regeneration in vivo (Bartsch et al., 1995), while OMgp knockout mice have yet to be analyzed for their regenerative potential. Surprisingly, both MAG and OMgp can bind to the Nogo receptor (NgR1), and this presumed redundancy of all three myelin inhibitors converging at a single receptor made NgR1 an attractive candidate target to disrupt in order to promote axon regeneration. Intrathecal infusion of a function-blocking NgR1 fragment (Li et al., 2004) or its expression as a transgene driven by the glial fibrillary acidic protein (GFAP) gene promoter (Li et al., 2005) resulted in sprouting of raphespinal and CST fibers caudal to the lesion. However, analysis of NgR1 knockout mice failed to show any enhanced regeneration of CST fibers (Kim et al., 2004; Zheng et al., 2005). Instead, there was regeneration of raphespinal and rubrospinal descending fibers associated with improved behavior after a complete transection (Kim et al., 2004). It is still unclear why the CST would regenerate in at least some Nogo KO mice (Kim et al., 2003; Dimou et al., 2006), but not in any NgR1 KO mice. One possibility is an even greater level of functional redundancy at the receptor level due to the presence of other Nogo receptors, such as NgR2 and NgR3 (Venkatesh et al., 2005). The effect of redundancy on axon regeneration at both the receptor as well as the ligand level is an issue that needs to be investigated further in future studies in order to elucidate the mechanism of myelin inhibition.

Genetic analysis of myelin inhibition using mutant mice has, at the very least, raised questions about the role of the three canonical myelin inhibitors in axon regeneration. The role of Nogo in axon regeneration still remains controversial, especially since none of the above mutant studies have passed the rigors of reproducibility by other independent laboratories. Once the reproducibility of the regenerative phenotype in these mice is established, future studies will need to focus on understanding the mechanism behind myelin inhibitors to ensure the implementation of the most effective therapeutic strategy.

3. Glial scar

After injury to the CNS, a glial scar typically forms around the injury site. This glial scar is composed of reactive astrocytes extending their hypertrophied processes to form both a physical barrier, by forming an interweaving network of processes, and a chemical barrier, by releasing inhibitory factors such as chondroitin sulfate proteoglycans (CSPGs) (for review, see Silver and Miller, 2004). The glial scar has traditionally been regarded inhibitory to axon regeneration mainly from indirect evidence such as labeled axons stopping at the region of astrogliosis (Davies et al., 1997) and detection of inhibitory factors such as CSPGs in this region.

The role of CSPGs in axon regeneration has received much attention from multiple labs ever since it was shown that breakdown of CSPGs using chondroitinase ABC (ChABC) promotes regeneration of sensory and CST axons as well as improves behavioral recovery after SCI in rats (Bradbury et al., 2002). However, transgenic mice expressing ChABC under the GFAP promoter failed to show any CST regeneration through the injury site after dorsal hemisection of the spinal cord (Cafferty et al., 2007). This was surprising considering that in theory the transgenic model would be a more efficient system of delivering ChABC within the lesion site whereas an intrathecal delivery method acts predominantly at the surface of the spinal cord. Aside from the expression level of the ChABC transgene, possible explanations of these differences will await further dissecting the mechanism of inhibition by CSPGs as well as sites of receptor activation. Furthermore, an inducible knockout model of glycosaminoglycan side chain synthesis of CSPGs may provide a more thorough analysis of the role of CSPGs in inhibiting axon regeneration.

The use of genetically modified mice has allowed a more direct investigation of the role of the glial scar as a physical barrier after spinal cord injury. The first studies to address this issue targeted the gene for GFAP, which is the intermediate filament whose increased expression is associated with reactive astrogliosis. The first study found that GFAP knockout mice do not exhibit increased sprouting or long distance regeneration of CST axons through the injury site after dorsal hemi-section of the spinal cord (Wang et al., 1997). A later study reported that double knockout mice for GFAP and vimentin, another astrocyte-associated intermediate filament, displayed increased sprouting of 5-HT axons into the denervated side caudal to a lateral hemi-section, but regeneration of axons across the lesion site was not reported (Menet et al., 2003).

A more direct way of investigating the role of the glial scar was employed by the use of transgenic mice expressing herpes simplex virus-thymidinekinase (HSVTK) under the GFAP promoter (Faulkner et al., 2004). Treatment of these transgenic mice with ganciclovir, which kills HSVTK-expressing cells, results in ablation of dividing, reactive astrocytes. Targeted ablation of these transgenic astrocytes after spinal cord injury showed very surprising results: there was increased tissue loss and lesion area, as well as greater motor deficits compared to controls. These results were corroborated by a more recent study that went further to investigate the signaling mechanism involved. Taking advantage of a downstream signal to reduce astrogliosis, deletion of Stat3 in nestin-positive (which is activated in astrocytes after SCI) cells resulted in increased infiltration of inflammatory cells, neuronal disruption, demyelination and motor deficits compared to control mice (Okada et al., 2006). In contrast, SCI in Nes-Socs3 mice (conditional Socs3 knockout in nestin-positive cells), which show increased activation of Stat3 after SCI, resulted in beneficial effects including reduced infiltration of inflammatory cells, lesion size, and demyelination. In addition, Nes-Socs3 mice attained remarkable behavioral recovery compared to controls: control mice, on the average, exhibited extensive movement of the hindlimb joints while dragging their lower bodies, but the Nes-Socs3 mutants had consistent weight-bearing plantar stepping with some degree of forelimb-hindlimb coordination. If verified, this would be one of the most significant improvements in locomotor recovery in any experimental SCI study reported thus far.

The use of mouse genetics has clearly demonstrated that although the glial scar may be a source of molecules inhibitory to axon regeneration, its formation plays a critical role in limiting the extent of the injury. Based on the data by Okada and colleagues (Okada et al., 2006), it could be argued that the benefits of the wound healing property of the glial scar outweighs the detrimental effect of inhibiting axon regeneration since no reports of axon regeneration has been associated with such a significant improvement in open field locomotion. A promising strategy might then be to neutralize specific components of the glial scar that inhibit axon regeneration without significantly altering its wound healing properties.

4. Chemorepulsive cues

During development, repulsive and attractive molecules are important for the proper wiring of the CNS (Tessier-Lavigne and Goodman, 1996). For reasons not fully understood, many repulsive signals are expressed into adulthood and some even exhibit increased expression at the lesion site after SCI (De Winter et al., 2002). The presence of chemorepulsive cues, such as semaphorins, at the injury site raises the possibility that it would be inhibitory to axon regeneration. This hypothesis is supported by recent pharmacological studies targeting two different chemorepulsive molecules. Intrathecal delivery of an antibody against repulsive guidance molecule (RGMa) promoted axonal growth of CST fibers and improved functional recovery after a dorsal hemisection in the rat (Hata et al., 2006). Likewise, a study with a Sema3A antagonist in a rat model of spinal cord injury indicates that blocking Sema3A promotes axonal sprouting into the lesion site, increases myelination by infiltration Schwann cells, and improves locomotor recovery (Kaneko et al., 2006). The promising results of these pharmacological studies have yet to be verified in their respective mutant mouse models.

There has only been one reported study investigating the role of axon guidance cues in spinal cord injury using mutant mice. Goldschmit and colleagues (Goldshmit et al., 2004) reported remarkable axon regeneration after a lateral hemisection in EphA4 knockout mice. In fact, approximately 70% of the labeled axons in the mutant mice crossed the lesion site, which would make this the strongest regenerative phenotype of any study reported so far. Interestingly, EphA4 knockout mice also displayed less reactive astrocytes at the injury site, and astrocytes cultured from EphA4 knockout mice showed migratory defects after a scratch wound assay in vitro (Goldshmit et al., 2004). Taken together, the regenerative phenotype and the recovery of hindpaw grip-strength observed in EphA4 knockout mice could be a direct effect of disrupting Ephrin signaling on the axons, and/or an indirect effect of disrupting the glial scar formation. If the latter is true, this would be in conflict with the phenotypes displayed by GFAP-HSV-TK transgenic mice and the Nes-Stat3 as well as Nes-Socs3 conditional knockout mice, which demonstrated that disrupting the glial scar formation has a detrimental effect on recovery after spinal cord injury (Faulkner et al., 2004; Okada et al., 2006). Future studies need to address the tissue-specific effect of EphA4 in axon regeneration in order to understand the mechanism of the significant regenerative phenotype observed in these mutant mice.

Currently, there are not enough published studies (using pharmacological or genetic methods) to make a generalized conclusion on the role of chemorepellent cues on axon regeneration after spinal cord injury. One lesson to be learned from using mutant mice in this area is the importance of careful baseline phenotype characterization since many of these mutants have axon guidance defects. For example, EphA4 mutants have a midline crossing defect where descending axons (such as the CST) that normally innervate one side of the spinal cord now also cross (or re-cross) over to the other side at the level of the spinal cord (Dottori et al., 1998). Although Goldshmit et al addressed the possibility of re-crossed fibers contributing to the regeneration of anterogradely labeled axons, this re-crossing phenotype invalidates the retrograde tracing method used in this mutant to identify the origin of the regenerating axons. Furthermore, the EphA4 mice have a gross motor defect in that they possess a hopping gait, which may be partially attributed to a wiring defect in the local central pattern generator (Kullander et al., 2003). This precludes the EphA4 mutants from any locomotor (and perhaps any other behavioral) assays because the anatomical defect not only alters the baseline behavior, but also confounds the interpretation of any behavioral recovery after SCI: is any improved recovery due to regeneration or just differences in anatomy? In order to circumvent these developmental abnormalities, future studies on chemorepellent cues will need to use acute gene deletion using inducible knockouts.

5. Genetic tracing and in vivo imaging

Much of the controversy in the axon regeneration field comes from the fact that regeneration itself still remains a very ill defined term. The most conservative definition of regeneration would be the regrowth of a cut axon. As simple as this may sound, it is an extremely difficult process to study experimentally. The current standard operating procedure in studying regeneration after SCI is to perform a partial or a complete lesion of the spinal cord and several weeks later perform an end-point analysis of either tracer-labeled or immunohistochemically labeled axons in tissue sections. Using this experimental paradigm, it is extremely difficult, if not impossible, to be absolutely sure that the “regenerated” axons labeled in the tissue section were cut by the initial lesion. The “regenerated” axons could be uncut axons initially located outside the lesion site but over time has traversed through or around the lesion site (i.e. sprouting of uninjured fibers, Fig. 1D). By the current standard, this type of axon, as long as it extends far into the other side of the lesion, would be considered to have regenerated. But can an axon that was never cut in the first place be truly considered to have regenerated? Alternatively, an axon that is cut by the lesion but regrows around the lesion site because of its inability to extend through it might be classified as local sprouting due to the inability to distinguish between cut and spared axons.

Fig. 1.

Fig. 1

Open in a new tab

Spinal cord injury results in lesion as well as sparing of axons (A), which may lead to a spectrum of axon growth behavior. Axon regeneration may occur if cut axons re-grow through or around the injury site (B). This re-growth may occur at either the terminal end or at a more distant point on the cut axon. Axon sprouting may occur through growth of spared axons that terminate rostral and/or caudal to the lesion (D). Spinal cord injury most likely results in both phenomena, thereby complicating the proper interpretation of many axon regeneration studies. Green represents regenerating axons, while blue represents sprouting axons.

The use of transgenic mice whose axons are genetically labeled with fluorescent markers has allowed us to overcome this experimental obstacle. Although axons can also be labeled with surgically or virally delivered fluorescent tracers, genetically traced axons are superior in many ways such as continuous production of fluorescent protein, no immunological response, and cell-type specificity. Kirchensteiner et al. (2005) was the first to capture images of axonal response to spinal cord injury through in vivo microscopy. By using the GFP-S line (Feng et al., 2000) in which very few ascending sensory axons are labeled with green fluorescent protein (GFP), Kerchensteiner et al. were able to capture in vivo images of axonal response after the labeled axon was cut with a needle. Through repetitive imaging of the same cut axon, they demonstrated an early phase of axonal degeneration (termed acute axonal degeneration), followed by Wallerian degeneration. Between 6–24 hours after lesion, many axons started to regrow at the transected tip or at the node of Ranvier at approximately 4 μm/hour. However, these regenerating axons grew laterally or in the opposite direction and did not appear to go through or past the lesion site. This phenomenon would most likely have been misinterpreted as sprouting rather than regeneration using conventional end-point histological analysis since these axons did not grow significantly beyond the lesion site.

More recently, Davalos et al. (2008) have developed a protocol for stable in vivo imaging of the mouse spinal cord from the YFP-H (Feng et al., 2000) and Cx3cr1GFP/+ (Jung et al., 2000) lines (available from the Jackson Laboratory) using two-photon microscopy to capture images of densely populated regions of microglia, axons and blood vessels. The use of two-photon microscopy enables reduced phototoxicity, deeper tissue penetration, greater spatial resolution and better spectral separation as compared to conventional wide-field epifluorescent microscopy that had been used previously (Kerschensteiner et al., 2005; Misgeld et al., 2007). Furthermore, the two-photon laser can be used to microablate single axons as used previously to demonstrate axon regeneration in C. elegans (Yanik et al., 2004; Wu et al., 2007). These technical improvements will allow us to better understand the mechanisms of axon degeneration/regeneration as well as the underlying cellular interaction through the use of transgenic mice in which multiple cell types are labeled with different fluorophores.

The use of repetitive imaging of axons has enormous potential in elucidating the molecular mechanisms of axon degeneration/regeneration. From just one study (Kerschensteiner et al., 2005), we have already learned that axons (ascending sensory axons in this case) may have some ability to regenerate, but such regeneration may be unfruitful due to a lack of the proper directional cues. Therefore, just because axons do not appear to cross the lesion site it does not necessarily mean that the experimental treatment does not promote regeneration; it may simply need the proper guidance cue(s). Through the use of two-photon microscopy, future in vivo imaging studies can investigate how axon degeneration/regeneration is affected by axonal interaction with other cell types such as astrocytes, oligodendrocytes and/or microglia.

6. Conclusion

We have yet to find a mutant mouse model with an incontrovertible enhanced axon regeneration phenotype after SCI. Nevertheless, the power of mouse genetics has significantly altered the landscape of studies of axon regeneration after spinal cord injury. For example, mouse studies question the major expectations for myelin inhibitors. In fact, we have yet to identify a single myelin molecule that presents a major obstacle to axon regeneration in vivo. In addition, the glial scar can no longer be considered simply as detrimental, since it seems to have beneficial effects in the healing of the injury site. Therefore, although the glial scar may still be inhibitory to axon regeneration, we need to be cautious in overcoming this barrier because it may do more harm than good. Lastly, a regenerating axon may lack the proper directional cue to reach its proper target, which must be carefully considered when designing experimental treatments to promote regeneration.

Another important lesson learned from mutant mouse studies has been the significant influence of mouse strain differences on functional recovery and regeneration after spinal cord injury. Axon sprouting/regeneration as well as behavioral recovery are lower in C57BL/6 strain compared to other pure or mixed strains (Ma et al., 2004; Dimou et al., 2006; Basso et al., 2006). When Nogo-A is deleted in a 129X1/SvJ background, it has two to four times more regenerating CST fibers as compared to a C57BL/6 background (Dimou et al., 2006). Therefore, controlling for strain differences will be an important consideration in the use of mutant mice for axon regeneration, especially when using double (or a greater level of combination of) knockouts since it will be more difficult to obtain littermate controls.

A major drawback in the field has been the use of mutant mice with germline mutations rather than inducible knockout technology. The use of germline mutations can lead to compensatory processes, such as up-regulation of Nogo transcripts after deletion of NgR1 (Kim et al., 2004; Zheng et al., 2005), and/or developmental defects, such as the axon guidance defect in EphA4 mice discussed above, both of which can confound the interpretation of the data. A better experimental paradigm would be to use inducible knockout mice in which the gene of interest is acutely deleted in the adult stage, thereby significantly reducing the likelihood of compensatory gene expression changes or developmental defects. There is also an additional advantage in having control over the exact timing of the gene deletion. Gene deletion can be induced after the injury, which will have significant impact on determining the therapeutic window of the axon regeneration promoting treatment. Future studies will need to shift towards the use of inducible gene knockout technology in assessing the role of candidate molecules in axon regeneration after spinal cord injury.

Although genetically modified mice can be a very powerful tool to study axon regeneration in vivo, it should be utilized with careful consideration of its strengths and weaknesses. As with other experimental approaches, the strongest evidence of axon regeneration will need to come from a combination of supporting experiments and will need to be reproducible by independent laboratories. Therefore, exchange of genetically modified mice should be highly encouraged in the field so that other independent laboratories can corroborate any claim of an axon regeneration phenotype before the gene or protein of interest is considered a potential therapeutic target.

Acknowledgments

Research in the authors’ laboratory has been supported by grants from the Roman Reed Spinal Cord Injury Research Fund of California, the Christopher and Dana Reeve Foundation, the International Spinal Research Trust, the Dana Foundation, the California Institute of Regenerative Medicine and NIH/NINDS (NS054734) to B.Z. J.K.L. is currently supported by a NRSA Postdoctoral Fellowship (F32NS056697) from NINDS.

References

  1. Bartsch U, Bandtlow CE, Schnell L, Bartsch S, Spillmann AA, Rubin BP, et al. Lack of evidence that myelin-associated glycoprotein is a major inhibitor of axonal regeneration in the CNS. Neuron. 1995;15:1375–1381. doi: 10.1016/0896-6273(95)90015-2. [DOI] [PubMed] [Google Scholar]
  2. Basso DM, Fisher LC, Anderson AJ, Jakeman LB, McTigue DM, Popovich PG. Basso Mouse Scale for locomotion detects differences in recovery after spinal cord injury in five common mouse strains. J Neurotrauma. 2006;23:635–659. doi: 10.1089/neu.2006.23.635. [DOI] [PubMed] [Google Scholar]
  3. Bradbury EJ, Moon LD, Popat RJ, King VR, Bennett GS, Patel PN, et al. Chondroitinase ABC promotes functional recovery after spinal cord injury. Nature. 2002;416:636–640. doi: 10.1038/416636a. [DOI] [PubMed] [Google Scholar]
  4. Cafferty WB, Yang SH, Duffy PJ, Li S, Strittmatter SM. Functional axonal regeneration through astrocytic scar genetically modified to digest chondroitin sulfate proteoglycans. J Neurosci. 2007;27:2176–2185. doi: 10.1523/JNEUROSCI.5176-06.2007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Caroni P, Schwab ME. Antibody against myelin-associated inhibitor of neurite growth neutralizes nonpermissive substrate properties of CNS white matter. Neuron. 1988;1:85–96. doi: 10.1016/0896-6273(88)90212-7. [DOI] [PubMed] [Google Scholar]
  6. Davalos D, Lee JK, Smith WB, Brinkman B, Ellisman MH, Zheng B, et al. Stable in vivo imaging of densely populated glia, axons and blood vessels in the mouse spinal cord using two-photon microscopy. J Neurosci Methods. 2008;169:1–7. doi: 10.1016/j.jneumeth.2007.11.011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. David S, Aguayo AJ. Axonal elongation into peripheral nervous system “bridges” after central nervous system injury in adult rats. Science. 1981;214:931–933. doi: 10.1126/science.6171034. [DOI] [PubMed] [Google Scholar]
  8. Davies SJ, Fitch MT, Memberg SP, Hall AK, Raisman G, Silver J. Regeneration of adult axons in white matter tracts of the central nervous system. Nature. 1997;390:680–683. doi: 10.1038/37776. [DOI] [PubMed] [Google Scholar]
  9. De Winter F, Oudega M, Lankhorst AJ, Hamers FP, Blits B, Ruitenberg MJ, et al. Injury-induced class 3 semaphorin expression in the rat spinal cord. Exp Neurol. 2002;175:61–75. doi: 10.1006/exnr.2002.7884. [DOI] [PubMed] [Google Scholar]
  10. Dimou L, Schnell L, Montani L, Duncan C, Simonen M, Schneider R, et al. Nogo-A-deficient mice reveal strain-dependent differences in axonal regeneration. J Neurosci. 2006;26:5591–5603. doi: 10.1523/JNEUROSCI.1103-06.2006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dottori M, Hartley L, Galea M, Paxinos G, Polizzotto M, Kilpatrick T, et al. EphA4 (Sek1) receptor tyrosine kinase is required for the development of the corticospinal tract. Proc Natl Acad Sci USA. 1998;95:13248–13253. doi: 10.1073/pnas.95.22.13248. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Faulkner JR, Herrmann JE, Woo MJ, Tansey KE, Doan NB, Sofroniew MV. Reactive astrocytes protect tissue and preserve function after spinal cord injury. J Neurosci. 2004;24:2143–2155. doi: 10.1523/JNEUROSCI.3547-03.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Feng G, Mellor RH, Bernstein M, Keller-Peck C, Nguyen QT, Wallace M, et al. Imaging neuronal subsets in transgenic mice expressing multiple spectral variants of GFP. Neuron. 2000;28:41–51. doi: 10.1016/s0896-6273(00)00084-2. [DOI] [PubMed] [Google Scholar]
  14. Goldshmit Y, Galea MP, Wise G, Bartlett PF, Turnley AM. Axonal regeneration and lack of astrocytic gliosis in EphA4-deficient mice. J Neurosci. 2004;24:10064–10073. doi: 10.1523/JNEUROSCI.2981-04.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. GrandPre T, Nakamura F, Vartanian T, Strittmatter SM. Identification of the Nogo inhibitor of axon regeneration as a Reticulon protein. Nature. 2000;403:439–444. doi: 10.1038/35000226. [DOI] [PubMed] [Google Scholar]
  16. Hata K, Fujitani M, Yasuda Y, Doya H, Saito T, Yamagishi S, et al. RGMa inhibition promotes axonal growth and recovery after spinal cord injury. J Cell Biol. 2006;173:47–58. doi: 10.1083/jcb.200508143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jung S, Aliberti J, Graemmel P, Sunshine MJ, Kreutzberg GW, Sher A, et al. Analysis of fractalkine receptor CX(3)CR1 function by targeted deletion and green fluorescent protein reporter gene insertion. Mol Cell Biol. 2000;20:4106–4114. doi: 10.1128/mcb.20.11.4106-4114.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kaneko S, Iwanami A, Nakamura M, Kishino A, Kikuchi K, Shibata S, et al. A selective Sema3A inhibitor enhances regenerative responses and functional recovery of the injured spinal cord. Nat Med. 2006;12:1380–1389. doi: 10.1038/nm1505. [DOI] [PubMed] [Google Scholar]
  19. Kerschensteiner M, Schwab ME, Lichtman JW, Misgeld T. In vivo imaging of axonal degeneration and regeneration in the injured spinal cord. Nat Med. 2005;11:572–577. doi: 10.1038/nm1229. [DOI] [PubMed] [Google Scholar]
  20. Kim JE, Li S, GrandPre T, Qiu D, Strittmatter SM. Axon regeneration in young adult mice lacking Nogo-A/B. Neuron. 2003;38:187–199. doi: 10.1016/s0896-6273(03)00147-8. [DOI] [PubMed] [Google Scholar]
  21. Kim JE, Liu BP, Park JH, Strittmatter SM. Nogo-66 receptor prevents raphespinal and rubrospinal axon regeneration and limits functional recovery from spinal cord injury. Neuron. 2004;44:439–451. doi: 10.1016/j.neuron.2004.10.015. [DOI] [PubMed] [Google Scholar]
  22. Kottis V, Thibault P, Mikol D, Xiao ZC, Zhang R, Dergham P, et al. Oligodendrocyte-myelin glycoprotein (OMgp) is an inhibitor of neurite outgrowth. J Neurochem. 2002;82:1566–1569. doi: 10.1046/j.1471-4159.2002.01146.x. [DOI] [PubMed] [Google Scholar]
  23. Kullander K, Butt SJ, Lebret JM, Lundfald L, Restrepo CE, Rydstrom A, et al. Role of EphA4 and EphrinB3 in local neuronal circuits that control walking. Science. 2003;299:1889–1892. doi: 10.1126/science.1079641. [DOI] [PubMed] [Google Scholar]
  24. Li S, Liu BP, Budel S, Li M, Ji B, Walus L, et al. Blockade of Nogo-66, myelin-associated glycoprotein, and oligodendrocyte myelin glycoprotein by soluble Nogo-66 receptor promotes axonal sprouting and recovery after spinal injury. J Neurosci. 2004;24:10511–10520. doi: 10.1523/JNEUROSCI.2828-04.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Li S, Kim JE, Budel S, Hampton TG, Strittmatter SM. Transgenic inhibition of Nogo-66 receptor function allows axonal sprouting and improved locomotion after spinal injury. Mol Cell Neurosci. 2005;29:26–39. doi: 10.1016/j.mcn.2004.12.008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Ma M, Wei P, Wei T, Ransohoff RM, Jakeman LB. Enhanced axonal growth into a spinal cord contusion injury site in a strain of mouse (129X1/SvJ) with a diminished inflammatory response. J Comp Neurol. 2004;474:469–486. doi: 10.1002/cne.20149. [DOI] [PubMed] [Google Scholar]
  27. McKerracher L, David S, Jackson DL, Kottis V, Dunn RJ, Braun PE. Identification of myelin-associated glycoprotein as a major myelin-derived inhibitor of neurite growth. Neuron. 1994;13:805–811. doi: 10.1016/0896-6273(94)90247-x. [DOI] [PubMed] [Google Scholar]
  28. Menet V, Prieto M, Privat A, Gimenez y Ribotta M. Axonal plasticity and functional recovery after spinal cord injury in mice deficient in both glial fibrillary acidic protein and vimentin genes. Proc Natl Acad Sci USA. 2003;100:8999–9004. doi: 10.1073/pnas.1533187100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Misgeld T, Nikic I, Kerschensteiner M. In vivo imaging of single axons in the mouse spinal cord. Nat Protoc. 2007;2:263–268. doi: 10.1038/nprot.2007.24. [DOI] [PubMed] [Google Scholar]
  30. Mukhopadhyay G, Doherty P, Walsh FS, Crocker PR, Filbin MT. A novel role for myelin-associated glycoprotein as an inhibitor of axonal regeneration. Neuron. 1994;13:757–767. doi: 10.1016/0896-6273(94)90042-6. [DOI] [PubMed] [Google Scholar]
  31. Okada S, Nakamura M, Katoh H, Miyao T, Shimazaki T, Ishii K, et al. Conditional ablation of Stat3 or Socs3 discloses a dual role for reactive astrocytes after spinal cord injury. Nat Med. 2006;12:829–834. doi: 10.1038/nm1425. [DOI] [PubMed] [Google Scholar]
  32. Savio T, Schwab ME. Rat CNS white matter, but not gray matter, is nonpermissive for neuronal cell adhesion and fiber outgrowth. J Neurosci. 1989;9:1126–1133. doi: 10.1523/JNEUROSCI.09-04-01126.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Schnell L, Schwab ME. Axonal regeneration in the rat spinal cord produced by an antibody against myelin-associated neurite growth inhibitors. Nature. 1990;343:269–272. doi: 10.1038/343269a0. [DOI] [PubMed] [Google Scholar]
  34. Silver J, Miller JH. Regeneration beyond the glial scar. Nat Rev Neurosci. 2004;5:146–156. doi: 10.1038/nrn1326. [DOI] [PubMed] [Google Scholar]
  35. Simonen M, Pedersen V, Weinmann O, Schnell L, Buss A, Ledermann B, et al. Systemic deletion of the myelin-associated outgrowth inhibitor Nogo-A improves regenerative and plastic responses after spinal cord injury. Neuron. 2003;38:201–211. doi: 10.1016/s0896-6273(03)00226-5. [DOI] [PubMed] [Google Scholar]
  36. Steward O, Zheng B, Banos K, Yee KM. Response to: Kim et al., "axon regeneration in young adult mice lacking Nogo-A/B." Neuron 38, 187–199. Neuron. 2007;54:191–195. doi: 10.1016/j.neuron.2007.04.004. [DOI] [PubMed] [Google Scholar]
  37. Teng FY, Tang BL. Why do Nogo/Nogo-66 receptor gene knockouts result in inferior regeneration compared to treatment with neutralizing agents? J Neurochem. 2005;94:865–874. doi: 10.1111/j.1471-4159.2005.03238.x. [DOI] [PubMed] [Google Scholar]
  38. Tessier-Lavigne M, Goodman CS. The molecular biology of axon guidance. Science. 1996;274:1123–1133. doi: 10.1126/science.274.5290.1123. [DOI] [PubMed] [Google Scholar]
  39. Venkatesh K, Chivatakarn O, Lee H, Joshi PS, Kantor DB, Newman BA, et al. The Nogo-66 receptor homolog NgR2 is a sialic acid-dependent receptor selective for myelin-associated glycoprotein. J Neurosci. 2005;25:808–822. doi: 10.1523/JNEUROSCI.4464-04.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wang KC, Koprivica V, Kim JA, Sivasankaran R, Guo Y, Neve RL, et al. Oligodendrocyte-myelin glycoprotein is a Nogo receptor ligand that inhibits neurite outgrowth. Nature. 2002;417:941–944. doi: 10.1038/nature00867. [DOI] [PubMed] [Google Scholar]
  41. Wang X, Messing A, David S. Axonal and nonneuronal cell responses to spinal cord injury in mice lacking glial fibrillary acidic protein. Exp Neurol. 1997;148:568–576. doi: 10.1006/exnr.1997.6702. [DOI] [PubMed] [Google Scholar]
  42. Wu Z, Ghosh-Roy A, Yanik MF, Zhang JZ, Jin Y, Chisholm AD. Caenorhabditis elegans neuronal regeneration is influenced by life stage, ephrin signaling, and synaptic branching. Proc Natl Acad Sci USA. 2007;104:15132–15137. doi: 10.1073/pnas.0707001104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Yanik MF, Cinar H, Cinar HN, Chisholm AD, Jin Y, Ben-Yakar A. Neurosurgery: functional regeneration after laser axotomy. Nature. 2004;432:822. doi: 10.1038/432822a. [DOI] [PubMed] [Google Scholar]
  44. Zheng B, Atwal J, Ho C, Case L, He XL, Garcia KC, et al. Genetic deletion of the Nogo receptor does not reduce neurite inhibition in vitro or promote corticospinal tract regeneration in vivo. Proc Natl Acad Sci USA. 2005;102:1205–1210. doi: 10.1073/pnas.0409026102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Zheng B, Ho C, Li S, Keirstead H, Steward O, Tessier-Lavigne M. Lack of enhanced spinal regeneration in Nogo-deficient mice. Neuron. 2003;38:213–224. doi: 10.1016/s0896-6273(03)00225-3. [DOI] [PubMed] [Google Scholar]
  46. Zheng B, Lee JK, Xie F. Genetic mouse models for studying inhibitors of spinal axon regeneration. Trends Neurosci. 2006;29:640–646. doi: 10.1016/j.tins.2006.09.005. [DOI] [PubMed] [Google Scholar]

RESOURCES