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01.12.2014 | Research article | Ausgabe 1/2014 Open Access

Molecular Neurodegeneration 1/2014

Optic nerve crush induces spatial and temporal gene expression patterns in retina and optic nerve of BALB/cJ mice

Zeitschrift:
Molecular Neurodegeneration > Ausgabe 1/2014
Autoren:
Tasneem P Sharma, Colleen M McDowell, Yang Liu, Alex H Wagner, David Thole, Benjamin P Faga, Robert J Wordinger, Terry A Braun, Abbot F Clark
Wichtige Hinweise

Electronic supplementary material

The online version of this article (doi:10.​1186/​1750-1326-9-14) contains supplementary material, which is available to authorized users.

Competing interests

The authors declare that they have no competing interests.

Authors’ contributions

TPS performed the RNA extraction, tissue sample staining, bioinformatics Partek and DAVID analysis, qRT-PCR and correlation analysis of all optic nerve crush samples; including writing all sections of the manuscript. CM and YL both performed the optic nerve crush and subsequently CM did the Nissl stained retinal flat mount neuron survival analysis while YL prepared the immunohistochemistry sections. BF processed image data. DT performed differential expression analysis and AW analyzed the microarray data using the BETR analysis. AFC, RJW and TAB conceived the study, actively participated in the design and coordination of the study, reviewed all the data, and helped draft the manuscript. All authors read and approved the final manuscript.

Abstract

Background

Central nervous system (CNS) trauma and neurodegenerative disorders trigger a cascade of cellular and molecular events resulting in neuronal apoptosis and regenerative failure. The pathogenic mechanisms and gene expression changes associated with these detrimental events can be effectively studied using a rodent optic nerve crush (ONC) model. The purpose of this study was to use a mouse ONC model to: (a) evaluate changes in retina and optic nerve (ON) gene expression, (b) identify neurodegenerative pathogenic pathways and (c) discover potential new therapeutic targets.

Results

Only 54% of total neurons survived in the ganglion cell layer (GCL) 28 days post crush. Using Bayesian Estimation of Temporal Regulation (BETR) gene expression analysis, we identified significantly altered expression of 1,723 and 2,110 genes in the retina and ON, respectively. Meta-analysis of altered gene expression (≥1.5, ≤-1.5, p < 0.05) using Partek and DAVID demonstrated 28 up and 20 down-regulated retinal gene clusters and 57 up and 41 down-regulated optic nerve clusters. Regulated gene clusters included regenerative change, synaptic plasticity, axonogenesis, neuron projection, and neuron differentiation. Expression of selected genes (Vsnl1, Syt1, Synpr and Nrn1) from retinal and ON neuronal clusters were quantitatively and qualitatively examined for their relation to axonal neurodegeneration by immunohistochemistry and qRT-PCR.

Conclusion

A number of detrimental gene expression changes occur that contribute to trauma-induced neurodegeneration after injury to ON axons. Nrn1 (synaptic plasticity gene), Synpr and Syt1 (synaptic vesicle fusion genes), and Vsnl1 (neuron differentiation associated gene) were a few of the potentially unique genes identified that were down-regulated spatially and temporally in our rodent ONC model. Bioinformatic meta-analysis identified significant tissue-specific and time-dependent gene clusters associated with regenerative changes, synaptic plasticity, axonogenesis, neuron projection, and neuron differentiation. These ONC induced neuronal loss and regenerative failure associated clusters can be extrapolated to changes occurring in other forms of CNS trauma or in clinical neurodegenerative pathological settings. In conclusion, this study identified potential therapeutic targets to address two key mechanisms of CNS trauma and neurodegeneration: neuronal loss and regenerative failure.
Zusatzmaterial
Additional file 1: Figure S1: A, B: Optic nerve crush (ONC) significantly reduces neurons in the retinal ganglion cell layer (RGCL). (JPEG 380 KB)
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Additional file 2: Figure S2: A, B: Frequency distribution of genes altered following optic nerve crush. (JPEG 357 KB)
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Additional file 3: Table S1: A, B: BETR probabilities based retinal and ON genes distributed within frequency bins. (JPEG 617 KB)
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Additional file 4: Table S2: A, B: Microarray ratios and linear regression correlation values of selected target genes. (JPEG 456 KB)
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Additional file 5: Figure S3: A-I: Naïve control images and expression of Syt1, Synpr, Nrn1 and Nfl in the ON at 7 days post crush. (JPEG 1 MB)
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Additional file 6: Figure S4: A, B: Expression of tissue specific genes within normal retina and ON samples. (JPEG 245 KB)
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Additional file 7: Table S3: Primers for key genes validated from the retina and optic nerve datasets. (JPEG 399 KB)
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Additional file 8: Table S4: qRT-PCR cycles performed for confirming retina and optic nerve dataset gene expression levels. (JPEG 398 KB)
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Additional file 9: Table S5: Antibodies against key proteins validated by IHC. (JPEG 484 KB)
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Authors’ original file for figure 1
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Authors’ original file for figure 2
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Authors’ original file for figure 3
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Authors’ original file for figure 4
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Authors’ original file for figure 5
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Authors’ original file for figure 6
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