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| Status |
Public on Dec 11, 2020 |
| Title |
(bulk RNAseq) An extracellular vesicle-related gene expression signature identifies high-risk patients in medulloblastoma |
| Organisms |
Homo sapiens; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Background: Medulloblastoma (MB) is a malignant brain tumor in childhood. It comprises four subgroups with different clinical behaviors. The aim of this study was to characterize the transcriptomic landscape of MB, both at the level of individual tumors as well as in large patient cohorts. Methods: We used a combination of single-cell transcriptomics, cell culture models and biophysical methods such as nanoparticle tracking analysis (NTA) and electron microscopy (EM), to investigate intercellular communication in the MB tumor niche. Results: Tumor cells of the SHH-MB subgroup show a differentiation blockade. These cells undergo extensive metabolic reprogramming. The gene expression profiles of individual tumor cells show a partial convergence with those of tumor-associated glial and immune cells. One possible cause is the transfer of extracellular vesicles (EVs) between cells in the tumor niche. We were able to detect EVs in co-culture models of MB tumor cells and oligodendrocytes. We also identified a gene expression signature, EVS, which shows overlap with the proteome profile of large oncosomes from prostate cancer cells. This signature is also present in MB patient samples. A high EVS expression is one common characteristic of tumors that occur in high-risk patients from otherwise very different MB subgroups or subtypes. Conclusions: With EVS, our study uncovered a novel gene expression signature that has a high prognostic significance across MB subgroups.
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| Overall design |
For bulk mRNA sequencing, total RNA was isolated using RNeasy mini Kit (Qiagen) from three different cell lines: murine oligodendrocytic cell line Oli-neu (gift of Tanja Kuhlmann, Institute of Neuropathology, University Münster), murine SHH-MB tumor cells MB1, and human Group 3 medulloblastoma cell line D341 (purchased from ATCC via LGC Standards GmBH, Wesel, Germany). MB1 cells were established from tumors of Atoh1-cre:SmoM2 mice at P12. Cells were maintained at 37°C and 5% CO2 in DMEM/F-12 medium containing B-27 and N-2 supplements (Gibco, Thermo Fisher Scientific), 20 ng/mL of recombinant murine EGF and basic fibroblast growth factor (bFGF) (PeproTech Germany), and 1% penicillin/streptomycin (Gibco). For co-culture experiments 30 mm Millicell cell culture inserts with hydrophilic PTFE membranes (0.4 µm pore size) were used (Millipore, Merck). Two biological replicates for each of the following experimental conditions were used: Oli-neu cells, MB1 cells and D341 cells grown in mono-culture (sample designations O_d1/2, M_d1/2, D_d1/2, respectively), Oli-neu cells grown in co-culture with MB1 cells (sample designations Om_d1/2 and Mo_d1/2, respectively), or Oli-neu cells grown in co-culture with D341 cells (sample designations Od_d1/2 and Do_d1/2, respectively). mRNA was extracted using the NEBNext Poly(A) mRNA Magnetic Isolation module, libraries were generated with the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs), and sequencing was done using Illumina’s High Output Kit v2.5 with 75 cycles on an NextSeq500 instrument.
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| Contributor(s) |
Kerl K, Albert T, Interlandi M |
| Citation(s) |
33175161 |
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| Submission date |
Oct 21, 2020 |
| Last update date |
Mar 12, 2021 |
| Contact name |
Kornelius Kerl |
| E-mail(s) |
kornelius.kerl@ukmuenster.de
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| Organization name |
University Children's Hospital Münster
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| Department |
Department of Pediatric Hematology and Oncology
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| Street address |
Albert-Schweitzer-Campus 1
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| City |
Münster |
| ZIP/Postal code |
48149 |
| Country |
Germany |
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| Platforms (2) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
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| Samples (14)
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| This SubSeries is part of SuperSeries: |
| GSE160003 |
An extracellular vesicle-related gene expression signature identifies high-risk patients in medulloblastoma |
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| Relations |
| BioProject |
PRJNA670439 |
| SRA |
SRP287993 |
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