Folate regulation of axonal regeneration in the rodent central nervous system through DNA methylation
Bermans J. Iskandar, … , Thomas D. Cook, Kirk J. Hogan
Bermans J. Iskandar, … , Thomas D. Cook, Kirk J. Hogan
Published April 26, 2010
Citation Information: J Clin Invest. 2010;120(5):1603-1616. https://doi.org/10.1172/JCI40000.
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Research Article

Folate regulation of axonal regeneration in the rodent central nervous system through DNA methylation

Abstract

The folate pathway plays a crucial role in the regeneration and repair of the adult CNS after injury. Here, we have shown in rodents that such repair occurs at least in part through DNA methylation. In animals with combined spinal cord and sciatic nerve injury, folate-mediated CNS axon regeneration was found to depend on injury-related induction of the high-affinity folate receptor 1 (Folr1). The activity of folate was dependent on its activation by the enzyme dihydrofolate reductase (Dhfr) and a functional methylation cycle. The effect of folate on the regeneration of afferent spinal neurons was biphasic and dose dependent and correlated closely over its dose range with global and gene-specific DNA methylation and with expression of both the folate receptor Folr1 and the de novo DNA methyltransferases. These data implicate an epigenetic mechanism in CNS repair. Folic acid and possibly other nontoxic dietary methyl donors may therefore be useful in clinical interventions to promote brain and spinal cord healing. If indeed the benefit of folate is mediated by epigenetic mechanisms that promote endogenous axonal regeneration, this provides possible avenues for new pharmacologic approaches to treating CNS injuries.

Authors

Bermans J. Iskandar, Elias Rizk, Brenton Meier, Nithya Hariharan, Teodoro Bottiglieri, Richard H. Finnell, David F. Jarrard, Ruma V. Banerjee, J.H. Pate Skene, Aaron Nelson, Nirav Patel, Carmen Gherasim, Kathleen Simon, Thomas D. Cook, Kirk J. Hogan

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Figure 7

Combined spinal cord and peripheral nerve injury suppresses the protein levels of the de novo methyltransferases Dnmt3a and Dnmt3b, but not the maintenance methyltransferase Dnmt1, and FA treatment restores the levels in a biphasic fashion that parallels the biphasic response to FA treatment seen with spinal regeneration and Folr1 expression.

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Combined spinal cord and peripheral nerve injury suppresses the protein ...
(A–D) Combined spinal cord and peripheral nerve injury suppresses Dnmt3a and Dnmt3b protein levels in mouse spinal cords, with no effect on Dnmt1. Furthermore, FA treatment restores these levels in both Dnmt3a and Dnmt3b. When increasing doses of FA are given i.p., a biphasic response is observed in the de novo methyltransferases Dnmt3a and Dnmt3b, with maximal protein levels occurring at 80 μg/kg. (F) Folic acid supplementation was previously shown to affect rat CNS regeneration following injury in a dose-dependent manner that corresponds tightly to the de novo Dnmt response. These results, obtained from our laboratory and previously reported by Iskandar et al. (7), are confirmed here in a different group of animals. One-way ANOVA with Bonferroni’s correction; n (FA dose) = 9 (0 μg/kg); 9 (40 μg/kg); 5 (80 μg/kg); 9 (400 μg/kg); 9 (800 μg/kg). Mean ± SEM; P < 0.05 for all comparisons except 0 versus 400, 0 versus 800, and 40 versus 400. Conversely, treatment with MTX significantly decreases Dnmt3a and Dnmt3b protein levels, while treatment with N2O nearly eliminates Dnmt3a and Dnmt3b protein expression. Neither injury nor folate, MTX, nor N2O affect the levels of the maintenance methyltransferase Dnmt1 (A, E). Furthermore, although Folr1 expression increases after injury, its response to increasing doses of FA is similarly biphasic. (A) Western blots. All lanes were run on the same gel. Noncontiguous lanes are separated by a thin white line in the figure. (B–D, E) Plots of the band densities relative to the corresponding actin bands. One-way ANOVA with Bonferroni’s correction; n = 3 in each group; mean ± SEM; *P < 0.05.