30 April 2011. Once again, BACE1 is the baddie. But in this instance, it is in nerve regeneration, not Alzheimer’s disease, where β-site amyloid precursor protease (APP) cleaving enzyme 1 (BACE1) seems to get in the way. BACE1 knockout mice regenerate crushed peripheral nerves faster, according to an April 13 paper in the Journal of Neuroscience. Although the mechanism at play is unclear, macrophages without the enzyme do a better job of clearing away debris from broken nerves, setting the stage for new growth. The study authors, at Johns Hopkins University in Baltimore, Maryland, also report that small molecule BACE1 inhibitors similarly promoted axon growth.
First author Mohamed Farah led the study with senior author John Griffin. Griffin succumbed April 16 to bladder cancer. He was the “mastermind of the whole project,” Farah said. “Jack Griffin was a beloved and much admired mentor,” he added in an e-mail to ARF. Griffin spent most of his four-decade career at Johns Hopkins School of Medicine, where he headed the department of neurology and founded the Brain Science Institute. Both the Brain Science Institute and the Baltimore Sun ran obituaries.
The researchers were inspired to examine BACE1’s potential role in nerve regeneration by reports that the enzyme cleaves neuregulin, which is involved in myelination during development (see ARF conference story on Willem et al., 2006; Michailov et al., 2004; Taveggia et al., 2005). Later studies showed that N-APP, another BACE1 cleavage product, triggers axon pruning and neural degeneration during development (see ARF related news story on Nikolaev et al., 2009).
Farah first examined the repair of sciatic nerve injury in BACE1 knockout mice, in which axons are hypomyelinated (Hu et al., 2006). Fixing a crushed or severed nerve is a multistep process. First, the damaged axons retract from the injury site. Macrophages clean up debris, particularly myelin. Finally, new axons grow into the site. Young rodents are able to re-innervate after crush injury to peripheral nerves, but humans are rarely so lucky: Distances are long, and the axons regrow so slowly that Schwann cells and target tissues atrophy and die before the new axons reach them (reviewed in Höke, 2006).
Within days of sciatic nerve crush, axons disintegrate in a process called Wallerian degeneration. Farah observed no difference in this process between wild-type and BACE1 knockout mice. But in the knockout animals, leftover fragments disappeared faster. Five days after the crush, knockout mice had less degraded myelin basic protein at the injury site. Their macrophages rapidly infiltrated the crushed nerves and accumulated cholesterol droplets, giving them a foamy appearance that indicates digestion of myelin. Two weeks after the nerve crush, the macrophages had mostly completed their task and left the site injury of BACE1 knockouts, whereas the macrophages were still present in the injury of wild-type mice at that time point.
The quick clearance of myelin was no great surprise; in part, it could have been because the BACE1 knockout mice have less myelin to begin with. Farah experimented on the macrophages in vitro to better understand the cleanup process. BACE1 knockout macrophages, he found, take up more small beads than do wild-type macrophages. Thus, the absence of BACE1 in these cells revs up their phagocytic appetite. The researchers suggest that macrophages must contain a BACE1 substrate, although they are not sure what that might be. The protein triggering receptor expressed on macrophage 2 (TREM2) is one attractive candidate, since it can improve the rate of myelin phagocytosis (Takahashi et al., 2007).
Having cleared a path, the BACE1 knockout mice then grew more and bigger axons, and did so faster. Axonal sprouts had grown farther in knockout than wild-type mice just three days after nerve crush. Five days into recovery, knockouts had more axonal sprouts in the distal stump, and more of them were greater than a micron in diameter, than in wild-type mice. Finally, foot and leg muscles were re-innervated faster in BACE1 mice.
This could be a result of neurons themselves growing faster, or simply a downstream result of the macrophages’ quick clearance of old fragments. To test the effects of BACE1 knockout in neurons alone, Farah and colleagues prepared explants of dorsal root ganglia (DRG). By four days of culture, axons in the knockout DRG were one and a half times as long as those in wild-type DRG, suggesting the loss of BACE1 directly affects axonal regrowth within the neuron. Perhaps, Farah speculated, N-APP acts as a “brake” to stop axon growth.
“A BACE1 inhibitor could be useful for conditions in which peripheral axons regenerate,” Farah said. He examined the effects of two chemical BACE1 inhibitors and found they promoted myelin clearance, regeneration, and re-innervation—though less than a BACE1 knockout did. Given within a month of injury, he suggested, such an inhibitor might promote regeneration. Then, removing the inhibitor would allow the axons to become normally myelinated.
Alzheimer’s researchers have been working on BACE1 inhibitors for a decade, with a few early-stage clinical candidates to show for the effort (for a recent update, see ARF AD/PD 2011 conference story). Many of the earlier candidates fail to cross the blood-brain barrier, but that would be unnecessary to treat peripheral nerve injury, Farah noted. More challenging would be treating spinal cord injury, which Farah is currently studying.
In addition, he suggested BACE1 inhibition might be helpful for people with neuropathy caused by diabetes or chemotherapy. As a proof of principle, Farah is currently crossing BACE1 knockout mice with mice that model the neuropathy amyotrophic lateral sclerosis. However, he noted the treatment would only regrow nerves that still have a cell body.
Therapeutic possibilities aside, the study offers a more general message about myelin and axon regeneration, wrote Rudolf Martini of the University of Würzburg in Germany, in an e-mail to ARF: “The study indirectly supports the concept of myelin removal as an important prerequisite for axon regrowth. This process is extremely delayed in the central nervous system, and may at least partially explain the chronically poor outcome of spinal cord injury and other traumata of the CNS.” (See full comment below.)—Amber Dance.
Farah MH, Pan BH, Hoffman PN, Ferraris D, Tsukamoto T, Nguyen T, Wong PC, Price DL, Slusher BS, Griffin JW. Reduced BACE1 activity enhances clearance of myelin debris and regeneration of axons in the injured peripheral nerve system. J Neurosci. 2011 Apr 13;31(15):5744-54. Abstract