How aging impacts axon regeneration after CNS injury is not known. We assessed the impact of age on axon regeneration induced by Pten deletion in corticospinal and rubrospinal neurons, two neuronal populations with distinct innate regenerative abilities. As in young mice, Pten deletion in older mice remains effective in preventing axotomy-induced decline in neuron-intrinsic growth state, as assessed by mTOR activity, neuronal soma size, and axonal growth proximal to a spinal cord injury. However, axonal regeneration distal to injury is greatly diminished, accompanied by increased expression of astroglial and inflammatory markers at the injury site. Thus, the mammalian CNS undergoes an age-dependent decline in axon regeneration, as revealed when neuron-intrinsic growth state is elevated. These results have important implications for developing strategies to promote axonal repair after CNS injuries or diseases, which increasingly affect middle-aged to aging populations.

The myelin-derived neurite growth inhibitor Nogo has been proposed to play a major role in blocking axon regeneration in the CNS after injuries. However, past studies have produced mixed results regarding the regenerative phenotype of various Nogo-deficient mouse lines after experimental spinal cord injury. Two lines did not display enhanced corticospinal tract (CST) regeneration, and one displayed modest regeneration. A fourth line, a Nogo-A,B gene-trap mutant, was instead reported to exhibit extensive CST regeneration, but the results were later found to be inadvertently confounded with an axon labeling artifact. Of the four Nogo mutant lines studied so far, three continue to express some isoform(s) of Nogo, leaving open the question whether any remaining Nogo protein contributes to the modest regenerative phenotype reported in some. The remaining Nogo mutant line studied was confounded by the unexplained rescue of embryonic lethality associated with this mutation. To gain a better understanding of the contribution of Nogo as an inhibitor of regeneration of CNS axons, and particularly CST axons, we reanalyzed the Nogo-A,B gene-trap mutant line and analyzed a novel, fully viable Nogo deletion mutant line that is null for all known isoforms of Nogo. Our analyses failed to reveal any enhanced CST regeneration after experimental spinal cord injury in either line. These results indicate that Nogo alone does not account for lack of CST regeneration and have implications for current therapeutic development for spinal cord injury in humans by targeting Nogo.

Axons in the adult CNS have poor ability to grow after injury, impeding functional recovery in patients of spinal cord injury. This has been attributed to both a developmental decline in neuron-intrinsic growth ability and the presence of extrinsic growth inhibitors. We previously showed that genetic deletion of Nogo, an extrinsic inhibitor, promoted axonal sprouting from uninjured corticospinal tract (CST) neurons but not regeneration from injured CST neurons, whereas genetic deletion of PTEN, an intrinsic inhibitor, promoted both CST sprouting and regeneration. Here we test the hypothesis that combining an elevation of neuron-intrinsic growth ability and a reduction of extrinsic growth inhibition by genetic codeletion of PTEN and Nogo may further improve injury-induced axonal growth. In an apparent paradox, additionally deleting Nogo further enhanced CST regeneration but not sprouting in PTEN-deleted mice. Enhanced CST regeneration and sprouting in PTEN and PTEN/Nogo-deleted mice were associated with no or only temporary improvement in functional recovery. Our data illustrate that neuron-intrinsic and -extrinsic factors regulate axon regeneration and sprouting in complex ways and provide proof-of-principle evidence that targeting both can further improve regeneration. Neuron-intrinsic growth ability is an important determinant of neuronal responsiveness to changes in extrinsic growth inhibition, such that an elevated intrinsic growth state is a prerequisite for reducing extrinsic inhibition to take effect on CST regeneration. Meanwhile, additional strategies are required to unleash the full potential for functional recovery with enhanced axon regeneration and/or sprouting.