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Molecular Neurobiology

, Volume 49, Issue 1, pp 276–287 | Cite as

Autophagy Reduces Neuronal Damage and Promotes Locomotor Recovery via Inhibition of Apoptosis After Spinal Cord Injury in Rats

  • Peifu Tang
  • Hongping Hou
  • Licheng Zhang
  • Xia Lan
  • Zhi Mao
  • Daohong Liu
  • Chunqing He
  • Hailong Du
  • Lihai ZhangEmail author
Article

Abstract

Autophagy is an intracellular catabolic mechanism that maintains the balance of proteins, lipids and aging organelles. 3-Methyladenine (3-MA) is a selective inhibitor of autophagy, whereas rapamycin, an antifungal agent, is a specific inducer of autophagy, inhibiting the protein mammalian target of rapamycin. In the present study, we examined the role of autophagy, inhibited by 3-MA and enhanced by rapamycin, in a model of acute spinal cord injury in rats. We found that rapamycin could significantly increase the expression of microtubule-associated protein 1 light chain 3 (LC3) and Beclin1 at the injury site. At the same time, the number of neurons and astrocytes with LC3 positive in the spinal cord was upregulated with time. In addition, administration of rapamycin produced an increase in the Basso, Beattie and Bresnahan scores of injured rats, indicating high recovery of locomotor function. Furthermore, expression of the proteins Bcl-2 and Bax was upregulated and downregulated, respectively. By contrast, the results for rats treated with 3-MA, which inhibits autophagy, were the opposite of those seen with the rapamycin-treated rats. These results show that induction of autophagy can produce neuroprotective effects in acute spinal cord injury in rats via inhibition of apoptosis.

Keywords

Spinal cord injury Autophagy 3-Methyladenine Rapamycin p62 Apoptosis 

Abbreviations

3-MA

3-Methyladenine

ANOVA

Analysis of variance

BBB

Basso, Beattie and Bresnahan

LC3

Microtubule-associated protein 1 light chain 3

mTOR

Mammalian target of rapamycin

PI3K

Phosphatidylinositol-3-kinase

SCI

Spinal cord injury

Notes

Acknowledgments

This research was supported by Natural Science Foundation of China (No. 30973068) and General projects of the Twelve-Fifth Scientific Plan in Army Medical Science and Technology (No. CWS11J101).

Conflict of interest

The authors declare that they have no competing interests.

References

  1. 1.
    Kabuta T, Furuta A, Aoki S, Furuta K, Wada K (2008) Aberrant interaction between Parkinson disease-associated mutant UCH-L1 and the lysosomal receptor for chaperone-mediated autophagy. J Biol Chem 283:23731–23738CrossRefPubMedGoogle Scholar
  2. 2.
    Mizushima N, Yamamoto A, Matsui M, Yoshimori T, Ohsumi Y (2004) In vivo analysis of autophagy in response to nutrient starvation using transgenic mice expressing a fluorescent autophagosome marker. Mol Biol Cell 15:1101–1111CrossRefPubMedCentralPubMedGoogle Scholar
  3. 3.
    Mizushima N (2004) Methods for monitoring autophagy. Int J Biochem Cell Biol 36:2491–2502CrossRefPubMedGoogle Scholar
  4. 4.
    Mizushima N, Yoshimori T, Levine B (2010) Methods in mammalian autophagy research. Cell 140:313–326CrossRefPubMedCentralPubMedGoogle Scholar
  5. 5.
    Klionsky DJ, Abeliovich H, Agostinis P, Agrawal DK, Aliev G, Askew DS et al (2008) Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy 4:151–175PubMedCentralPubMedGoogle Scholar
  6. 6.
    Nixon RA, Cataldo AM, Mathews PM (2000) The endosomal–lysosomal system of neurons in Alzheimer’s disease pathogenesis: a review. Neurochem Res 25(9–10):1161–1172CrossRefPubMedGoogle Scholar
  7. 7.
    Ross CA, Poirier MA (2005) Opinion: what is the role of protein aggregation in neurodegeneration? Nat Rev Mol Cell Biol 6:891–898CrossRefPubMedGoogle Scholar
  8. 8.
    Pickford F, Masliah E, Britschgi M, Lucin K, Narasimhan R, Jaeger PA et al (2008) The autophagy-related protein beclin 1 shows reduced expression in early Alzheimer disease and regulates amyloid beta accumulation in mice. J Clin Invest 118:2190–2199PubMedCentralPubMedGoogle Scholar
  9. 9.
    Nixon RA (2007) Autophagy, amyloidogenesis and Alzheimer disease. J Cell Sci 120:4081–4091CrossRefPubMedGoogle Scholar
  10. 10.
    Hayashi S, Sato N, Yamamoto A, Ikegame Y, Nakashima S, Ogihara T et al (2009) Alzheimer disease-associated peptide, amyloid beta40, inhibits vascular regeneration with induction of endothelial autophagy. Arterioscler Thromb Vasc Biol 29:1909–1915CrossRefPubMedGoogle Scholar
  11. 11.
    Michiorri S, Gelmetti V, Giarda E, Lombardi F, Romano F, Marongiu R et al (2010) The Parkinson-associated protein PINK1 interacts with Beclin1 and promotes autophagy. Cell Death Differ 17:962–974CrossRefPubMedGoogle Scholar
  12. 12.
    Chu CT (2010) A pivotal role for PINK1 and autophagy in mitochondrial quality control: implications for Parkinson disease. Hum Mol Genet 19:R28–R37CrossRefPubMedGoogle Scholar
  13. 13.
    Yamamoto A, Cremona ML, Rothman JE (2006) Autophagy-mediated clearance of huntingtin aggregates triggered by the insulin-signaling pathway. J Cell Biol 172:719–731CrossRefPubMedGoogle Scholar
  14. 14.
    Cao L, Xu J, Lin Y, Zhao X, Liu X, Chi Z (2009) Autophagy is upregulated in rats with status epilepticus and partly inhibited by vitamin E. Biochem Biophys Res Commun 379:949–953CrossRefPubMedGoogle Scholar
  15. 15.
    Caro LH, Plomp PJ, Wolvetang EJ, Kerkhof C, Meijer AJ (1988) 3-Methyladenine, an inhibitor of autophagy, has multiple effects on metabolism. Eur J Biochem 175:325–329CrossRefPubMedGoogle Scholar
  16. 16.
    Takatsuka C, Inoue Y, Matsuoka K, Moriyasu Y (2004) 3-Methyladenine inhibits autophagy in tobacco culture cells under sucrose starvation conditions. Plant Cell Physiol 45:265–274CrossRefPubMedGoogle Scholar
  17. 17.
    Ito S, Koshikawa N, Mochizuki S, Takenaga K (2007) 3-Methyladenine suppresses cell migration and invasion of HT1080 fibrosarcoma cells through inhibiting phosphoinositide 3-kinases independently of autophagy inhibition. Int J Oncol 31:261–268PubMedGoogle Scholar
  18. 18.
    McFarland AJ, Anoopkumar-Dukie S, Perkins AV, Davey AK, Grant GD (2011) Inhibition of autophagy by 3-methyladenine protects 1321N1 astrocytoma cells against pyocyanin- and 1-hydroxyphenazine-induced toxicity. Arch Toxicol 86(2):275–284CrossRefPubMedGoogle Scholar
  19. 19.
    Sehgal SN, Baker H, Vezina C (1975) Rapamycin (AY-22,989), a new antifungal antibiotic. II. Fermentation, isolation and characterization. J Antibiot (Tokyo) 28:727–732CrossRefGoogle Scholar
  20. 20.
    Erlich S, Alexandrovich A, Shohami E, Pinkas-Kramarski R (2007) Rapamycin is a neuroprotective treatment for traumatic brain injury. Neurobiol Dis 26:86–93CrossRefPubMedGoogle Scholar
  21. 21.
    Rami A, Langhagen A, Steiger S (2008) Focal cerebral ischemia induces upregulation of Beclin 1 and autophagy-like cell death. Neurobiol Dis 29:132–141CrossRefPubMedGoogle Scholar
  22. 22.
    Puyal J, Vaslin A, Mottier V, Clarke PG (2009) Postischemic treatment of neonatal cerebral ischemia should target autophagy. Ann Neurol 66:378–389CrossRefPubMedGoogle Scholar
  23. 23.
    Carloni S, Buonocore G, Balduini W (2008) Protective role of autophagy in neonatal hypoxia–ischemia induced brain injury. Neurobiol Dis 32:329–339CrossRefPubMedGoogle Scholar
  24. 24.
    Diskin T, Tal-Or P, Erlich S, Mizrachy L, Alexandrovich A, Shohami E et al (2005) Closed head injury induces upregulation of Beclin 1 at the cortical site of injury. J Neurotrauma 22:750–762CrossRefPubMedGoogle Scholar
  25. 25.
    Erlich S, Shohami E, Pinkas-Kramarski R (2006) Neurodegeneration induces upregulation of Beclin 1. Autophagy 2:49–51PubMedGoogle Scholar
  26. 26.
    Smith CM, Chen Y, Sullivan ML, Kochanek PM, Clark RS (2011) Autophagy in acute brain injury: feast, famine, or folly? Neurobiol Dis 43:52–59CrossRefPubMedCentralPubMedGoogle Scholar
  27. 27.
    Rangaraju S, Verrier JD, Madorsky I, Nicks J, Dunn WJ, Notterpek L (2010) Rapamycin activates autophagy and improves myelination in explant cultures from neuropathic mice. J Neurosci 30:11388–11397CrossRefPubMedCentralPubMedGoogle Scholar
  28. 28.
    Basso DM, Beattie MS, Bresnahan JC (1996) Graded histological and locomotor outcomes after spinal cord contusion using the NYU weight-drop device versus transection. Exp Neurol 139:244–256CrossRefPubMedGoogle Scholar
  29. 29.
    Basso DM, Beattie MS, Bresnahan JC, Anderson DK, Faden AI, Gruner JA et al (1996) MASCIS evaluation of open field locomotor scores: effects of experience and teamwork on reliability. Multicenter Animal Spinal Cord Injury Study. J Neurotrauma 13:343–359CrossRefPubMedGoogle Scholar
  30. 30.
    Mathew R, Karp CM, Beaudoin B, Vuong N, Chen G, Chen HY et al (2009) Autophagy suppresses tumorigenesis through elimination of p62. Cell 137:1062–1075CrossRefPubMedCentralPubMedGoogle Scholar
  31. 31.
    Bjorkoy G, Lamark T, Brech A, Outzen H, Perander M, Overvatn A et al (2005) p62/SQSTM1 forms protein aggregates degraded by autophagy and has a protective effect on huntingtin-induced cell death. J Cell Biol 171:603–614CrossRefPubMedGoogle Scholar
  32. 32.
    Pankiv S, Clausen TH, Lamark T, Brech A, Bruun JA, Outzen H et al (2007) p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregates by autophagy. J Biol Chem 282:24131–24145CrossRefPubMedGoogle Scholar
  33. 33.
    Yang J, Liu X, Bhalla K, Kim CN, Ibrado AM, Cai J et al (1997) Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science 275:1129–1132CrossRefPubMedGoogle Scholar
  34. 34.
    Wei MC, Zong WX, Cheng EH, Lindsten T, Panoutsakopoulou V, Ross AJ et al (2001) Proapoptotic BAX and BAK: a requisite gateway to mitochondrial dysfunction and death. Science 292:727–730CrossRefPubMedCentralPubMedGoogle Scholar
  35. 35.
    Gupta S, Knowlton AA (2005) HSP60, Bax, apoptosis and the heart. J Cell Mol Med 9:51–58CrossRefPubMedGoogle Scholar
  36. 36.
    Jurgensmeier JM, Xie Z, Deveraux Q, Ellerby L, Bredesen D, Reed JC (1998) Bax directly induces release of cytochrome c from isolated mitochondria. Proc Natl Acad Sci U S A 95:4997–5002CrossRefPubMedCentralPubMedGoogle Scholar
  37. 37.
    Huang J, Klionsky DJ (2007) Autophagy and human disease. Cell Cycle 6:1837–1849CrossRefPubMedGoogle Scholar
  38. 38.
    Levine B, Kroemer G (2008) Autophagy in the pathogenesis of disease. Cell 132:27–42CrossRefPubMedCentralPubMedGoogle Scholar
  39. 39.
    Zhivotovsky B, Orrenius S (2010) Cell cycle and cell death in disease: past, present and future. J Intern Med 268:395–409CrossRefPubMedGoogle Scholar
  40. 40.
    Barnett A, Brewer GJ (2011) Autophagy in aging and Alzheimer’s disease: pathologic or protective? J Alzheimers Dis 25:385–394PubMedCentralPubMedGoogle Scholar
  41. 41.
    Sasaki S (2011) Autophagy in spinal cord motor neurons in sporadic amyotrophic lateral sclerosis. J Neuropathol Exp Neurol 70:349–359CrossRefPubMedGoogle Scholar
  42. 42.
    Brown KN, Chen S, Han Z, Lu CH, Tan X, Zhang XJ et al (2011) Clonal production and organization of inhibitory interneurons in the neocortex. Science 334:480–486CrossRefPubMedCentralPubMedGoogle Scholar
  43. 43.
    Korn MJ, Koppel SJ, Cramer KS (2011) Astrocyte-secreted factors modulate a gradient of primary dendritic arbors in nucleus laminaris of the avian auditory brainstem. PLoS One 6:e27383CrossRefPubMedCentralPubMedGoogle Scholar
  44. 44.
    Barreto GE, Sun X, Xu L, Giffard RG (2011) Astrocyte proliferation following stroke in the mouse depends on distance from the infarct. PLoS One 6:e27881CrossRefPubMedCentralPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Peifu Tang
    • 1
  • Hongping Hou
    • 1
    • 2
  • Licheng Zhang
    • 1
  • Xia Lan
    • 1
  • Zhi Mao
    • 1
  • Daohong Liu
    • 1
  • Chunqing He
    • 1
  • Hailong Du
    • 1
  • Lihai Zhang
    • 1
    Email author
  1. 1.Department of OrthopaedicsChinese PLA General HospitalBeijingPeople’s Republic of China
  2. 2.Medical CollegeNankai UniversityTianjinPeople’s Republic of China

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