Ann Rehabil Med.  2016 Aug;40(4):559-567. 10.5535/arm.2016.40.4.559.

The Modulation of Neurotrophin and Epigenetic Regulators: Implication for Astrocyte Proliferation and Neuronal Cell Apoptosis After Spinal Cord Injury

• Kim JH
• Kim SH
• Cho SR
• Lee JY
• Kim JH
• Baek A
• Jung HS

• KMID: 2356640
• DOI: http://doi.org/10.5535/arm.2016.40.4.559

Abstract

OBJECTIVE
To investigate alterations in the expression of the main regulators of neuronal survival and death related to astrocytes and neuronal cells in the brain in a mouse model of spinal cord injury (SCI).
METHODS
Eight-week-old male imprinting control region mice (n=36; 30-35 g) were used in this study and randomly assigned to two groups: the naïve control group (n=18) and SCI group (n=18). The mice in both groups were randomly allocated to the following three time points: 3 days, 1 week, and 2 weeks (n=6 each). The expression levels of regulators such as brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), nerve growth factor (NGF), histone deacetylase 1 (HDAC1), and methyl-CpG-binding protein 2 (MeCP 2) in the brain were evaluated following thoracic contusive SCI. In addition, the number of neuronal cells in the motor cortex (M1 and M2 areas) and the number of astrocytes in the hippocampus were determined by immunohistochemistry.
RESULTS
BDNF expression was significantly elevated at 2 weeks after injury (p=0.024). The GDNF level was significantly elevated at 3 days (p=0.042). The expression of HDAC1 was significantly elevated at 1 week (p=0.026). Following SCI, compared with the control the number of NeuN-positive cells in the M1 and M2 areas gradually and consistently decreased at 2 weeks after injury. In contrast, the number of astrocytes was significantly increased at 1 week (p=0.029).
CONCLUSION
These results demonstrate that the upregulation of BDNF, GDNF and HDAC1 might play on important role in brain reorganization after SCI.

Keyword

Spinal cord injuries; Genes; Regulator; Astrocytes; Neurons

MeSH Terms

Animals
Apoptosis*
Astrocytes*
Brain
Brain-Derived Neurotrophic Factor
Epigenomics*
Glial Cell Line-Derived Neurotrophic Factor
Hippocampus
Histone Deacetylase 1
Humans
Immunohistochemistry
Male
Methyl-CpG-Binding Protein 2
Mice
Motor Cortex
Nerve Growth Factor
Neurons*
Spinal Cord Injuries*
Spinal Cord*
Up-Regulation
Brain-Derived Neurotrophic Factor
Glial Cell Line-Derived Neurotrophic Factor
Histone Deacetylase 1
Methyl-CpG-Binding Protein 2
Nerve Growth Factor

Figure

• 

  Fig. 1 Neurotrophic factor expression by Western blotting analysis. (A) Representative expressions of BDNF, NGF, and GDNF in the brain after contusion of the spinal cord in mice. (B) BDNF significantly elevated compared with those of control group at 2 weeks after SCI (*p<0.05). (C) Representative expressions of NGF in brain after contusion of the spinal cord in mice. NGF elevated compared with those of control group at 1 and 2 weeks and decreased at 3 days; however, the differences were not significant. (D) Representative GDNF expression in the brain after contusion of the spinal cord in mice. GDNF expression was significantly elevated compared with the control group at 3 days (*p<0.05). Data are plotted as mean±standard error of the mean. BDNF, brain-derived neurotrophic factor; NGF, nerve growth factor; GDNF, glial cell line-derived neurotrophic factor; SCI, spinal cord injury; GAPDH, glyceraldehyde 3-phosphate dehydrogenase. *p<0.05 indicates values significantly different from those of SCI and the control group.

• 

  Fig. 2 HDAC1 and MeCP2 expression by Western blotting analysis. (A) Representative expressions of HDAC1 and MeCP2 in the brain after contusion of the spinal cord in mice. (B) The difference in the HDAC1 level between the SCI and control groups was significant at 1 week after SCI (*p<0.05). (C) The difference in the MeCP2 level between the SCI and control groups after SCI was not significant. Data are plotted as mean±standard error of the mean. HDAC1, histone deacetylase 1; MeCP2, methyl-CpG-binding protein 2. *p<0.05 indicates values significantly different from those of the SCI and control groups.

• 

  Fig. 3 Spinal cord injury (SCI) reduces the number of surviving neurons in the M1 and M2 areas. SCI resulted in significant neuronal cell loss in the M1 and M2 areas at 1 and 2 weeks after injury. (A) Schematic representation of a coronal atlas section of rodent brain corresponding to bregma 1.10 mm. (B) Representative findings of NeuN+ staining in the coronal sections passing through the rostral and caudal parts of the M1 and M2 areas at 1 and 2 weeks after SCI. (C) Higher magnification images of NeuN+ cells in the M1 and M2 areas. (D) The number of NeuN+ cells in the M1 and M2 areas. The total number of NeuN+ cells of the lesioned M1 and M2 areas were significantly lower in the SCI group (*p<0.05). Scale bar represents 500 µm in (B) and 50 µm in (C). *p<0.05 indicates values significantly different from those of the SCI and control groups.

• 

  Fig. 4 Spinal cord injury (SCI) increases the number of surviving astrocytes in the hippocampus. (A) Representative findings of GFAP staining in the coronal sections passing through the rostral and caudal parts of the hippocampus at 1 and 2 weeks after SCI. (B) The number of GFAP+ cells in the hippocampus. The total number of GFAP+ cells was significantly greater in the SCI group at 1 week compared with the control group (*p<0.05). Scale bar represents 100 µm in (A). *p<0.05 indicates values significantly different from those of the SCI and control groups.

Reference

1. Bregman BS, Goldberger ME. Infant lesion effect. II: Sparing and recovery of function after spinal cord damage in newborn and adult cats. Brain Res. 1983; 285:119–135. PMID: 6616260.

2. Yu SH, Cho DC, Kim KT, Nam KH, Cho HJ, Sung JK. The neuroprotective effect of treatment of valproic Acid in acute spinal cord injury. J Korean Neurosurg Soc. 2012; 51:191–198. PMID: 22737297.

3. Burns SP, Golding DG, Rolle WA Jr, Graziani V, Ditunno JF Jr. Recovery of ambulation in motor-incomplete tetraplegia. Arch Phys Med Rehabil. 1997; 78:1169–1172. PMID: 9365343.

4. Raineteau O, Schwab ME. Plasticity of motor systems after incomplete spinal cord injury. Nat Rev Neurosci. 2001; 2:263–273. PMID: 11283749.

5. Jain N, Catania KC, Kaas JH. Deactivation and reactivation of somatosensory cortex after dorsal spinal cord injury. Nature. 1997; 386:495–498. PMID: 9087408.

6. Florence SL, Taub HB, Kaas JH. Large-scale sprouting of cortical connections after peripheral injury in adult macaque monkeys. Science. 1998; 282:1117–1121. PMID: 9804549.

7. Bruehlmeier M, Dietz V, Leenders KL, Roelcke U, Missimer J, Curt A. How does the human brain deal with a spinal cord injury? Eur J Neurosci. 1998; 10:3918–3922. PMID: 9875370.

8. Curt A, Bruehlmeier M, Leenders KL, Roelcke U, Dietz V. Differential effect of spinal cord injury and functional impairment on human brain activation. J Neurotrauma. 2002; 19:43–51. PMID: 11852977.

9. Green JB, Sora E, Bialy Y, Ricamato A, Thatcher RW. Cortical sensorimotor reorganization after spinal cord injury: an electroencephalographic study. Neurology. 1998; 50:1115–1121. PMID: 9566404.

10. McKinley PA, Jenkins WM, Smith JL, Merzenich MM. Age-dependent capacity for somatosensory cortex reorganization in chronic spinal cats. Brain Res. 1987; 428:136–139. PMID: 3815108.

11. Boutillier AL, Trinh E, Loeffler JP. Selective E2F-dependent gene transcription is controlled by histone deacetylase activity during neuronal apoptosis. J Neurochem. 2003; 84:814–828. PMID: 12562525.

12. Legube G, Trouche D. Regulating histone acetyltransferases and deacetylases. EMBO Rep. 2003; 4:944–947. PMID: 14528264.

13. Kumar A, Loane DJ. Neuroinflammation after traumatic brain injury: opportunities for therapeutic intervention. Brain Behav Immun. 2012; 26:1191–1201. PMID: 22728326.

14. Wu J, Stoica BA, Luo T, Sabirzhanov B, Zhao Z, Guanciale K, et al. Isolated spinal cord contusion in rats induces chronic brain neuroinflammation, neurodegeneration, and cognitive impairment. Involvement of cell cycle activation. Cell Cycle. 2014; 13:2446–2458. PMID: 25483194.

15. Kim BG, Dai HN, McAtee M, Vicini S, Bregman BS. Remodeling of synaptic structures in the motor cortex following spinal cord injury. Exp Neurol. 2006; 198:401–415. PMID: 16443221.

16. Lee R, Kermani P, Teng KK, Hempstead BL. Regulation of cell survival by secreted proneurotrophins. Science. 2001; 294:1945–1948. PMID: 11729324.

17. Seidah NG, Chretien M. Proprotein and prohormone convertases: a family of subtilases generating diverse bioactive polypeptides. Brain Res. 1999; 848:45–62. PMID: 10701998.

18. Seidah NG, Benjannet S, Pareek S, Chretien M, Murphy RA. Cellular processing of the neurotrophin precursors of NT3 and BDNF by the mammalian proprotein convertases. FEBS Lett. 1996; 379:247–250. PMID: 8603699.

19. Ying SW, Futter M, Rosenblum K, Webber MJ, Hunt SP, Bliss TV, et al. Brain-derived neurotrophic factor induces long-term potentiation in intact adult hippocampus: requirement for ERK activation coupled to CREB and upregulation of Arc synthesis. J Neurosci. 2002; 22:1532–1540. PMID: 11880483.

20. Endo T, Spenger C, Tominaga T, Brene S, Olson L. Cortical sensory map rearrangement after spinal cord injury: fMRI responses linked to Nogo signalling. Brain. 2007; 130(Pt 11):2951–2961. PMID: 17913768.

21. Buss A, Brook GA, Kakulas B, Martin D, Franzen R, Schoenen J, et al. Gradual loss of myelin and formation of an astrocytic scar during Wallerian degeneration in the human spinal cord. Brain. 2004; 127(Pt 1):34–44. PMID: 14534158.

22. Wrigley PJ, Gustin SM, Macey PM, Nash PG, Gandevia SC, Macefield VG, et al. Anatomical changes in human motor cortex and motor pathways following complete thoracic spinal cord injury. Cereb Cortex. 2009; 19:224–232. PMID: 18483004.

23. Mowla SJ, Pareek S, Farhadi HF, Petrecca K, Fawcett JP, Seidah NG, et al. Differential sorting of nerve growth factor and brain-derived neurotrophic factor in hippocampal neurons. J Neurosci. 1999; 19:2069–2080. PMID: 10066260.

24. Sydekum E, Ghosh A, Gullo M, Baltes C, Schwab M, Rudin M. Rapid functional reorganization of the forelimb cortical representation after thoracic spinal cord injury in adult rats. Neuroimage. 2014; 87:72–79. PMID: 24185021.