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. 2025 Jan 7:15:1470441.
doi: 10.3389/fneur.2024.1470441. eCollection 2024.

Molecular profiling of frontal and occipital subcortical white matter hyperintensities in Alzheimer's disease

Sulochan Malla #  1   2 , Annie G Bryant #  1   3 , Rojashree Jayakumar  1 , Benjamin Woost  1 , Nina Wolf  1 , Andrew Li  1 , Sudeshna Das  1   2 , Susanne J van Veluw  1   2 , Rachel E Bennett  1   2
Affiliations

Molecular profiling of frontal and occipital subcortical white matter hyperintensities in Alzheimer's disease

Sulochan Malla et al. Front Neurol. .

Abstract

White matter hyperintensities (WMHs) are commonly detected on T2-weighted magnetic resonance imaging (MRI) scans, occurring in both typical aging and Alzheimer's disease (AD). Despite their frequent appearance and their association with cognitive decline in AD, the molecular factors contributing to WMHs remain unclear. In this study, we investigated the transcriptomic profiles of two commonly affected brain regions with coincident AD pathology-frontal subcortical white matter (frontal-WM) and occipital subcortical white matter (occipital-WM)-and compared with age-matched cognitively intact controls. Through RNA-sequencing in frontal- and occipital-WM bulk tissues, we identified an upregulation of genes associated with brain vasculature function in AD white matter. To further elucidate vasculature-specific transcriptomic features, we performed RNA-seq analysis on blood vessels isolated from these white matter regions, which revealed an upregulation of genes related to protein folding pathways. Finally, comparing gene expression profiles between AD individuals with high- versus low-WMH burden showed an increased expression of pathways associated with immune function. Taken together, our study characterizes the diverse molecular profiles of white matter changes in AD and provides mechanistic insights into the processes underlying AD-related WMHs.

Keywords: Alzheimer’s disease; angiogenesis; blood vessels; brain vasculature; heat shock proteins (HSPs); protein folding; white matter hyperintensities (WMHs).

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.

Figures

Figure 1
Figure 1
Experimental outline and characterization of WMH in control and AD cases. (A) Schematic representation of experimental outline and conditions. Tissues were obtained from the Massachusetts ADRC. One hemisphere of the brain was fixed in 10% formalin and was used for ex vivo 7T MRI and histopathology. The other hemisphere was freshly frozen, and tissues were used for RNA-seq from the bulk tissues and isolated blood vessels. Not all donors used for RNA-seq had the alternate hemisphere and corresponding brain regions available for histology. (B) Examples of ex vivo T2-weighted TSE MRI scan showing the frontal- and occipital-WM from control and AD tissues. WMHs were visible as subtle periventricular changes only (yellow arrow) or, in the most affected brains, as hyperintense areas extending into deep subcortical WM (red arrows). (C) Dissection of region of interest (ROI) showing WMH in the frontal- and occipital-WM as indicated by the ex vivo MRI images in (B). (D) Luxol Fast Blue (LFB) staining of the control and AD samples in frontal- and occipital-WM. (E) The percentage area of LFB staining is shown in control and AD for frontal- and occipital-WM.
Figure 2
Figure 2
Gene expression changes in frontal-and occipital-WM bulk tissues. (A,D) Volcano plot showing differential gene expression in (A) frontal-WM bulk tissues and (D) occipital-WM bulk tissues. The color of dots indicates the direction of significantly altered genes, with red indicating upregulation and blue indicating downregulation. (B,C,E,F) Bar plot showing the enriched biological processes (GO/KEGG terms, canonical pathways) generated by Metascape for (B,E) upregulated genes, and (C,F) downregulated genes. The enriched pathways associated with brain vasculature in (E,F) and synaptic functions in (C) are marked by green boxes. (G) Venn diagram showing unique and shared DEGs in the frontal- and occipital-WM bulk tissues for upregulated genes (upper panel), and downregulated genes (lower panel). (H) Biological processes associated with upregulated genes common to frontal- and occipital-WM bulk tissues, generated by ShinyGO. The enriched pathways associated with brain vasculature (B,E) and synaptic functions in (C) are marked by green boxes.
Figure 3
Figure 3
Gene expression changes in isolated blood vessels from frontal-and occipital-WM in AD cases. (A,D) Volcano plots showing differential gene expression in blood vessels (BV) of (A) frontal-WM and (D) occipital-WM. The color of dots indicates the direction of significantly altered genes, with red indicating upregulation and blue indicating downregulation. (B,C,E,F) Bar plot showing the enriched biological processes (GO/KEGG terms, canonical pathways) generated by Metascape for (B,E) upregulated genes, and (C,F) downregulated genes. Enriched pathways associated with protein folding in (B,E) are indicated by green boxes. (G) Venn diagram showing unique and shared DEGs in the isolated blood vessels from frontal- and occipital-WM for upregulated genes (upper panel) and downregulated genes (lower panel). (H) Enrichment network visualization for results from the significantly upregulated gene lists in AD versus control, color code represents that HSPs are generally shared among all RNA-seq samples.
Figure 4
Figure 4
Histopathological validation of heat shock proteins. Immunohistochemistry of frontal-WM tissue from AD (A–C) and Control (D–F) donors. (A,D) An overview of tissue sections with gray matter (dashed line) and white matter (solid line) outlined showing the approximate location of higher magnification views shown in (B,C,E,F). (B,E) HSP90AA1 and (C,F) HSPA6 labeling in an adjacent tissue section. Samples correspond to AD-8 and Control-5 (see Table 1). Arrowheads highlight vascular cells with heat shock protein positivity. (G) An image of tissue that was handled following the same procedure as in panels (A–F) but without the overnight incubation with primary antibody (experimental control).
Figure 5
Figure 5
Characterization of low- and high-WMH in frontal- and occipital-WM in AD cases. (A) Examples of in vivo T2-weighted FLAIR MRI scans showing segmented WMH volumes (red pixels) in AD in frontal-WM (A,B) and occipital-WM (C,D). Tissue sections indicated by green boxes corresponding to the in vivo WMH (red and gray pixels) from AD presenting low-WMH (A,C) and high-WMH (B,D) were dissected. (E) Comparison of high- and low-WMH samples based on volume of WMH region. (F) Pearson correlation between the in vivo MRI (volume) and age at the time of the scan. (G–I) Volcano plot showing differential gene expression in low- versus high-WMH in occipital-WM from (G) bulk tissues, and (I) blood vessels. (H–J) Bar plot showing the enriched biological processes (GO/KEGG terms, canonical pathways) generated by Metascape for low- versus high-WMH in occipital-WM from (H) bulk tissues, and (J) blood vessels.
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