Axonal regeneration and lack of astrocytic gliosis in EphA4-deficient mice

Investigators : Dr Yona Goldshmit, Prof Mary Galea, Prof Perry Bartlett, Dr Ann Turnley

Injury to the central nervous system (CNS) usually results in very limited, if any, regeneration of lesioned axons, with subsequent permanent impairment of function. Although some CNS neurons appear to lose the intrinsic ability to regenerate neurites postnatally, many others, such as corticospinal tract (CST) neurons, appear able to regenerate, but are inhibited from doing so by the environment of the injury site. Major impediments to CNS regeneration are the presence of myelin inhibitors and astrocytic gliosis.

Other factors that may be implicated in inhibition of axonal regeneration, but which have received little attention in this regard, are cell surface molecules that are involved in axon guidance during development. One such family of molecules is the Eph receptor tyrosine kinase family, which, together with their ligands, the Ephrins, are implicated in the formation of a variety of neural structures, including the corticospinal tract and anterior commissure. As Eph/Ephrin signaling appears to regulate axon guidance through contact repulsion, inducing the collapse of neuronal growth cones, and members of this family are upregulated in the adult following neural injury, the aberrant expression or absence of Eph receptors could prove pivotal in determining the outcome of injury in the adult CNS. This possibility was investigated in this study by comparing neural regeneration following spinal cord hemisection in wildtype and EphA4-/- mice.

Adult mice lacking EphA4 (-/-) exhibit axonal regeneration and functional recovery after spinal cord hemisection. Anterograde and retrograde tracing showed that axons from multiple pathways, including corticospinal and rubrospinal tracts, crossed the lesion site. EphA4-/- mice recovered stride length, the ability to walk on and climb a grid and the ability to grasp with the affected hindpaw within 1-3 months of injury. EphA4 expression was upregulated on astrocytes at the lesion site in wildtype mice, while astrocytic gliosis and the glial scar were greatly reduced in lesioned EphA4-/- spinal cords.   EphA4-/- astrocytes failed to respond to the inflammatory cytokines IFN? or LIF in vitro . Neurons grown on wildtype astrocytes extended shorter neurites than on EphA4-/- astrocytes, but longer neurites when the astrocyte EphA4 was blocked by monomeric EphrinA5-Fc. Thus EphA4 regulates two important features of spinal cord injury, axonal inhibition and astrocytic gliosis.

Figure 1 . A montage of confocal images of EphA4-/- spinal cord (i) showed that the regenerating axons passed through the lesion site (indicated by dotted line and by H&E stained section in (iv) and extended caudally in a straight line with some "waviness" seen immediately post-lesion. In both panels rostral is to the right and caudal to the left, and the lesion site is indicated by dotted lines. Enlarged areas are indicated by boxed areas and arrows. Scale bars for panels (i) 250µm; panels (ii) 200µm, panels (iii to v) 50µm. Asterisk in panel (a i) indicates the midline .

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