The Microglial Transcriptome of Age-Associated Deep Subcortical White Matter Lesions Suggests a Neuroprotective Response to Blood–Brain Barrier Dysfunction
Abstract
:1. Introduction
2. Results
2.1. Histological Characterization Confirms the Classification of Sampled WM
2.2. RNA-Sequencing Is Not Compatible with Immuno-LCM-ed Post-Mortem Tissue in the CFAS Cohort
2.3. Microarray Transcriptomic Profiling Is Compatible with Immuno-LCM in the CFAS Cohort
2.4. Validation of Candidate Gene Expression
3. Discussion
4. Materials and Methods
4.1. Case Selection and Histological Characterization
4.2. Laser Capture Microdissection (LCM) of Microglia
4.3. RNA-Sequencing
RNA-Seq Data Analysis
4.4. Microarray
Microarray Data Analysis
4.5. Immunohistochemical Validation of Protein Changes
Statistical Analysis
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Sample ID | Length | M Seq (×106) | % Fail | % GC | % Dup | % Over-Representative |
---|---|---|---|---|---|---|
Sample 1_fastq | 101 bp | 3.1 | 45% | 87% | 76.5% | 23.5% |
Sample 2_fastq | 101 bp | 6 | 36% | 87% | 77.7% | 23.5% |
Sample 3_fastq | 101 bp | 6.9 | 36% | 90% | 82.3% | 25.0% |
Sample 4_fastq | 101 bp | 7.8 | 36% | 88% | 80.0% | 25.0% |
Sample 5_fastq | 101 bp | 16.5 | 45% | 83% | 67.3% | 21.5% |
Sample 6_fastq | 101 bp | 6.3 | 36% | 89% | 80.9% | 25.0% |
Sample 7_fastq | 101 bp | 6.1 | 45% | 90% | 81.7% | 23.5% |
Sample 8_fastq | 101 bp | 9.5 | 45% | 80% | 66.8% | 19.7% |
Sample 9_fastq | 101 bp | 16.8 | 45% | 83% | 78.8% | 21.5% |
Sample 10_fastq | 101 bp | 8.4 | 36% | 89% | 80.1% | 19.7% |
Sample 11_fastq | 101 bp | 9.6 | 36% | 89% | 81.2% | 25.0% |
Sample 12_fastq | 101 bp | 8.9 | 45% | 90% | 80.8% | 19.7% |
Annotated Cluster | Enrichment Score | Gene Count | Benjamini p-Value | Directional Change | Differentially Expressed Genes |
---|---|---|---|---|---|
Haptoglobin binding | 2.5 | 3 | 1.7 × 10−2 | Up | Haemoglobin-α1 (HBA1) Haemoglobin-α2 (HBA2) Haemoglobin-β (HBB) |
Haptoglobin–haemoglobin complex | 3 | 2.4 × 10−2 | |||
TBC | 2.66 | 7 | 6.3 × 10−6 | Down | TBC1 domain family member: (TBC1D3B, TBC1D3E, TBC1D3F, TBC1D3G, TBC1D3H, TBC1D3I, TBC1D3L) |
Rab-GTPase-TBC domain | 4.3 × 10−3 |
Annotated Cluster | Enrichment Score | Gene Count | Benjamini p-Value | Directional Change | Differentially Expressed Genes |
---|---|---|---|---|---|
Protein deubiquitination | 5.14 | 9 | 1.3 × 10−4 | Down | Ubiquitin specific peptidase 17-like family member: USP17L5, USP17L19, USP17L24, USP17L25, USP17L26, USP17L27, USP17L28, USP17L29, USP17L30, Ring finger protein 8 (RNF8) Toll interaction protein (TOLLIP) |
Ubiquitin-dependent protein catabolic process | 11 | 1.3 × 10−4 |
WM Group | Control (n = 4) | NAWM (n = 4) | DSCLs (n = 4) |
---|---|---|---|
CD163 | 0.47 (0.29–0.85) | 1.41 (0.43–3.24) | 4.07 (2.12–6.41) |
SRBI | 0.46 (0.22–0.70) | 0.83 (0.74–0.93) | 7.02 (1.85–15.09) * |
Dub3 | 0.25 (0.20–0.28) | 0.44 (0.30–0.66) | 4.44 (1.09–7.89) * |
Primary Antibody | Species | Clonality | Isotype | Dilution | Supplier |
---|---|---|---|---|---|
CD163 | Rabbit | Monoclonal | IgG | 1:500 | Abcam, UK |
Dub3 | Rabbit | Polyclonal | IgG | 1:100 | Novusbio, UK |
SRBI | Rabbit | Monoclonal | IgG | 1:500 | Abcam, UK |
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Almansouri, T.; Waller, R.; Wharton, S.B.; Heath, P.R.; Matthews, F.E.; Brayne, C.; van Eeden, F.; Simpson, J.E. The Microglial Transcriptome of Age-Associated Deep Subcortical White Matter Lesions Suggests a Neuroprotective Response to Blood–Brain Barrier Dysfunction. Int. J. Mol. Sci. 2024, 25, 4445. https://doi.org/10.3390/ijms25084445
Almansouri T, Waller R, Wharton SB, Heath PR, Matthews FE, Brayne C, van Eeden F, Simpson JE. The Microglial Transcriptome of Age-Associated Deep Subcortical White Matter Lesions Suggests a Neuroprotective Response to Blood–Brain Barrier Dysfunction. International Journal of Molecular Sciences. 2024; 25(8):4445. https://doi.org/10.3390/ijms25084445
Chicago/Turabian StyleAlmansouri, Taghreed, Rachel Waller, Stephen B. Wharton, Paul R. Heath, Fiona E. Matthews, Carol Brayne, Fredericus van Eeden, and Julie E. Simpson. 2024. "The Microglial Transcriptome of Age-Associated Deep Subcortical White Matter Lesions Suggests a Neuroprotective Response to Blood–Brain Barrier Dysfunction" International Journal of Molecular Sciences 25, no. 8: 4445. https://doi.org/10.3390/ijms25084445