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| Status |
Public on Jun 28, 2024 |
| Title |
Augmenting fibronectin levels in injured adult CNS promotes axon regeneration in vivo [bulk RNA-seq] |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
In an attempt to repair injured central nervous system (CNS) nerves/tracts, immune cells are recruited into the injury site, but endogenous response in adult mammals is insufficient for promoting regeneration of severed axons. Here, we found that a portion of retinal ganglion cell (RGC) CNS projection neurons that survive after optic nerve crush (ONC) injury are enriched for and upregulate fibronectin (Fn)-interacting integrins Itga5 and ItgaV, and that Fn promotes long-term survival and long-distance axon regeneration of a portion of axotomized adult RGCs in culture. We then show that, Fn is developmentally downregulated in the axonal tracts of optic nerve and spinal cord, but injury-activated macrophages/microglia upregulate Fn while axon regeneration-promoting zymosan augments their recruitment (and thereby increases Fn levels) in the injured optic nerve. Finally, we found that Fn’s RGD motif, established to interact with Itga5 and ItgaV, promotes long-term survival and long-distance axon regeneration of adult RGCs after ONC in vivo, with some axons reaching the optic chiasm when co-treated with Rpl7a gene therapy. Thus, experimentally augmenting Fn levels in the injured CNS is a promising approach for therapeutic neuroprotection and axon regeneration of at least a portion of neurons.
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| Overall design |
Adult uninjured or ONC-injured RGCs transduced with AAV2 expressing mCherry were Thy1-immunopanned from single cell retinal suspension (from 10 retinas per condition) and FACS’ed for mCherry+ cells. Approximately, 5,000 RGCs (per condition) were collected by FACS. RNA was isolated immediately using the Zymo QuickRNA microprep kit. Total RNA with RNA Integrity Number (RIN) ≥ 9 (by Bioanalyzer 2100 using the Nano 6000 kit, Agilent) was extracted using Direct-zol RNA MiniPrep kit (R2050, Zymo Research). cDNA libraries were prepared using polyA-selected RNA (TruSeq RNA Library Prep Kit, Illumina), paired reads sequenced 100 bp from each end on HiSeq 2000 Sequencer (Illumina), passed QC filters, mapped to the mm10 genome and transcriptome by Hisat2, gene expression was normalized as fragments per kilobase of transcript per million mapped reads (FPKM), and analyzed by Cufflinks/CuffDiff.
Please note: This series contains raw and processed data for two genes which is atypical for GEO studies.
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| Contributor(s) |
Lukomska A, Frost MP, Rheaume BA, Trakhtenberg EF |
| Citation(s) |
38944331 |
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| Submission date |
Jun 13, 2024 |
| Last update date |
Jul 01, 2024 |
| Contact name |
Ephraim F Trakhtenberg |
| E-mail(s) |
trakhtenberg@uchc.edu
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| Organization name |
University of Connecticut School of Medicine
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| Department |
Neuroscience
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| Street address |
263 Farmington Ave. RM L4005
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| City |
Farmington |
| State/province |
CT |
| ZIP/Postal code |
06030 |
| Country |
USA |
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| Platforms (1) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
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| Samples (2) |
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| This SubSeries is part of SuperSeries: |
| GSE269816 |
Augmenting fibronectin levels in injured adult CNS promotes axon regeneration in vivo |
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| Relations |
| BioProject |
PRJNA1124236 |
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