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| Status |
Public on Nov 08, 2022 |
| Title |
Comparative analysis of single-cell and single-nucleus RNA-sequencing in a rabbit model of retinal detachment-related proliferative vitreoretinopathy |
| Organism |
Oryctolagus cuniculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Proliferative vitreoretinopathy (PVR) is the most common cause of failure of retinal reattachment surgery and the molecular changes leading to this aberrant wound healing process is currently unknown. We aimed to study PVR pathogenesis using single-cell transcriptomics to dissect cellular heterogeneity in a rabbit PVR model. PVR was induced unilaterally in Dutch Belted rabbits. At different timepoints following PVR induction, retinas were dissociated into either cells or nuclei suspension and processed for single-cell or single-nucleus RNA sequencing (scRNA-seq or snRNA-seq). ScRNA-Seq and snRNA-Seq were conducted on retinas at 4 hours and 14 days after disease induction. While the capture rate of unique molecular identifiers (UMI) and genes were greater in scRNA-seq samples, overall gene expression profiles of individual cell types were highly correlated between scRNA-seq and snRNA-seq. A major disparity between the two sequencing modalities is the cell type capture rate, however, with glial cell types over-represented in scRNA-seq, and inner retinal neurons were enriched by snRNA-seq. Furthermore, fibrotic Müller glia were over-represented in snRNA-seq samples, while reactive Müller glia were in scRNA-seq samples. Trajectory analyses were similar between the two methods, allowing for the combined analysis of the scRNA-seq and snRNA-seq datasets.These findings highlight limitations of both scRNA-seq and snRNA-seq analysis and imply that use of both techniques can more accurately identify transcriptional networks critical for aberrant fibrogenesis in PVR.
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| Overall design |
Retinas from control or PVR induced rabbits were dissociated into a single cell suspension or lysed into a single nuclei solution and run on the 10x Genomics Chromium Single Cell System platform using ver3.1 chemistry. Libraries were sequenced on Illumina NovaSeq 6000 and processed through the Cell Ranger pipeline using the OryCun2.0 reference genome.
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| Contributor(s) |
Santiago CP, Gimmen M, Lu Y, McNally MM, Duncan LH, Creamer TJ, Orzolek LD, Blackshaw S, Singh MS |
| Citation(s) |
38153753 |
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https://doi.org/10.1016/j.xops.2023.100335
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| Submission date |
Nov 04, 2022 |
| Last update date |
Jan 03, 2024 |
| Contact name |
Clayton Pio Santiago |
| E-mail(s) |
csanti10@jhmi.edu
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| Organization name |
Johns Hopkins University
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| Department |
Neuroscience
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| Street address |
733 North Broadway
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| City |
Baltimore |
| State/province |
Maryland |
| ZIP/Postal code |
21205 |
| Country |
USA |
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| Platforms (1) |
| GPL26786 |
Illumina NovaSeq 6000 (Oryctolagus cuniculus) |
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| Samples (10)
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| Relations |
| BioProject |
PRJNA898403 |
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