Abstract
Nogo-A, an axonal growth inhibitory protein known to be mostly present in CNS myelin, was upregulated in retinal ganglion cells (RGCs) after optic nerve injury in adult mice. Nogo-A increased concomitantly with the endoplasmic reticulum stress (ER stress) marker C/EBP homologous protein (CHOP), but CHOP immunostaining and the apoptosis marker annexin V did not co-localize with Nogo-A in individual RGC cell bodies, suggesting that injury-induced Nogo-A upregulation is not involved in axotomy-induced cell death. Silencing Nogo-A with an adeno-associated virus serotype 2 containing a short hairpin RNA (AAV2.shRNA-Nogo-A) or Nogo-A gene ablation in knock-out (KO) animals had little effect on the lesion-induced cell stress or death. On the other hand, Nogo-A overexpression mediated by AAV2.Nogo-A exacerbated RGC cell death after injury. Strikingly, however, injury-induced sprouting of the cut axons and the expression of growth-associated molecules were markedly reduced by AAV2.shRNA-Nogo-A. The axonal growth in the optic nerve activated by the intraocular injection of the inflammatory molecule Pam3Cys tended to be lower in Nogo-A KO mice than in WT mice. Nogo-A overexpression in RGCs in vivo or in the neuronal cell line F11 in vitro promoted regeneration, demonstrating a positive, cell-autonomous role for neuronal Nogo-A in the modulation of axonal regeneration.
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Abbreviations
- AAV:
-
adeno-associated virus
- ATF:
-
activating transcription factor
- BDNF:
-
brain-derived neurotrophic factor
- Bip/GRP78:
-
78 kDa glucose-regulated protein
- CHOP/GADD153:
-
C/EBP homologous protein/growth arrest- and DNA damage-inducible gene 153
- CNTF:
-
ciliary neurotrophic factor
- CTb-594:
-
cholera toxin β subunit-alexa594
- DRG:
-
dorsal root ganglion
- eIF2α:
-
eukaryotic translation initiation factor 2 α
- FGF2:
-
fibroblast growth factor 2
- FL:
-
fiber layer
- GAP-43:
-
growth-associated protein 43
- GAPDH:
-
glyceraldehyde 3-phosphate dehydrogenase
- GCL:
-
ganglion cell layer
- GFAP:
-
glial fibrillary acidic protein
- GFP:
-
green fluorescent protein
- GS:
-
glutamine synthetase
- ILM:
-
inner limiting membrane
- INL:
-
inner nuclear layer
- IPL:
-
inner plexiform layer
- Jak3:
-
janus kinase 3
- LIF:
-
leukemia inhibitory factor
- LINGO1:
-
leucine rich repeat and Ig domain containing 1
- MAG:
-
myelin-associated glycoprotein
- mTOR:
-
mammalian target of rapamycin
- NgR1:
-
Nogo66 receptor 1
- OLM:
-
outer limiting membrane
- ONL:
-
outer nuclear layer
- OPL:
-
outer plexiform layer
- OS:
-
outer segment
- Pam3Cys:
-
(S)-(2,3-bis(palmitoyloxy)-(2RS)-propyl)-N-palmitoyl-(R)-Cys-(S)-Ser(S)-Lys(4)-OH^trihydrochloride
- PTEN:
-
phosphatase and tensin homolog
- qRT-PCR:
-
quantitative real-time polymerase chain reaction
- RGCs:
-
retinal ganglion cells
- RTN:
-
reticulon
- shRNA:
-
small hairpin ribonucleic acid
- SOCS3:
-
suppressor of cytokine signaling 3
- Sprr1A:
-
small proline-rich protein 1A
- Stat3:
-
signal transducer and activator of transcription 3
- TNF-α:
-
tumor necrosis factor α
- TSC1:
-
tuberous sclerosis complex 1
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Acknowledgements
We want to thank Dr. Jody L Martin for the preparation of the AAV.shRNA-Nogo-A and the INSERM viral vector core facility in Nantes, France, for the production of AAV2.Nogo-A and AAV2.GFP. This work was supported by Swiss National Science Foundation (SNF) Grant no. 31-122527/1 and the SNF National Center of Competence in Research ‘Neural Plasticity and Repair’.
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Pernet, V., Joly, S., Dalkara, D. et al. Neuronal Nogo-A upregulation does not contribute to ER stress-associated apoptosis but participates in the regenerative response in the axotomized adult retina. Cell Death Differ 19, 1096–1108 (2012). https://doi.org/10.1038/cdd.2011.191
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DOI: https://doi.org/10.1038/cdd.2011.191
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