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| Status |
Public on Oct 28, 2021 |
| Title |
Spatial transcriptomics of healing and non-healing diabetic foot ulcers |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
Other
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| Summary |
Diabetic foot ulcers (DFUs) are a devastating complication of diabetes. To better understand the molecular mechanisms and cell types implicated in DFU healing, we used NanoString’s GeoMx Digital Spatial profiling platform on DFU tissue sections and compared gene expression of areas within the same ulcer as well as between patients who in 12 weeks following surgery healed their DFU (Healers, N=2) vs those who did not (Non-Healers, N=2).
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| Overall design |
The spatial transcriptome profiling was performed using NanoString’s GeoMx Digital Spatial profiling platform on unfixed frozen 5 μm tissue sections. Samples were processed as follows: 1) 10% neutral buffered formalin fixation overnight, 2) target retrieval (1X Tris EDTA, pH 9.0 for 20 min), 3) proteinase K digestion (1 µg/mL for 15 min), 4) post-fixation (10% NBF, Tris-glycine stop buffer), 5) in-situ hybridization overnight with the GeoMx Cancer Transcriptome Atlas probe panel (1800-plex), 6) stringent washes (50:50 formamide/4X SSC), and 7) fluorescent antibody/marker (aSMA, Clone: 1A4, Abcam; CD45, Clone: 2B11+PD7/26, Novus; PanCK, Clone: AE1/AE3, Novus) incubation, 1 hour at room temperature. Sections were then loaded onto the GeoMx® Digital Spatial Profiler. For profiling, circular regions of interest (ROIs), approximately 500 μm in diameter, located within the ulcers or in neighboring non-ulcerated tissue were selected to include high concentrations of CD45+ immune cells in close proximity to vessels (aSMA+ structures). After ROI selection, the GeoMx instrument illuminated each ROI separately with UV light to cleave, aspirate, and deposit the oligonucleotides from the hybridized ISH probes for downstream sequencing into a 96-well plate.
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| Contributor(s) |
Theocharidis G, Veves A |
| Citation(s) |
35013299 |
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| Submission date |
Feb 03, 2021 |
| Last update date |
Jan 12, 2022 |
| Contact name |
Aristidis Veves |
| E-mail(s) |
[email protected]
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| Phone |
6176627075
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| Organization name |
Beth Israel Deaconess Medical Center
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| Lab |
Rongxiang Xu, MD, Center for Regenerative Therapeutics
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| Street address |
330 Brookline Ave
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
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| Platforms (1) |
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| Samples (23)
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| Relations |
| BioProject |
PRJNA699294 |
| SRA |
SRP304590 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE166120_Alpha_CTA_v2.0.pkc.txt.gz |
676.0 Kb |
(ftp)(http) |
TXT |
| GSE166120_RAW.tar |
730.0 Kb |
(http)(custom) |
TAR (of DCC) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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