Disease-associated oligodendrocyte signatures are spatiotemporally dysregulated in spinocerebellar ataxia type 3

Abstract

Spinocerebellar ataxia type 3 (SCA3) is a neurodegenerative disease caused by a CAG repeat expansion in the ATXN3 gene. Though the ATXN3 protein is expressed ubiquitously throughout the CNS, regional pathology in SCA3 patients is observed within select neuronal populations and more recently within oligodendrocyte-rich white matter tracts. We have previously recapitulated these white matter abnormalities in an overexpression mouse model of SCA3 and demonstrated that oligodendrocyte maturation impairments are one of the earliest and most progressive changes in SCA3 pathogenesis. Disease-associated oligodendrocyte signatures have recently emerged as significant contributors to several other neurodegenerative diseases, including Alzheimer's disease, Huntington's disease, and Parkinson's disease, but their role in regional vulnerability and disease progression remains unexplored. Here, we are the first to comparatively assess myelination in human tissue in a region-dependent manner. Translating these findings to SCA3 mouse models of disease, we confirmed endogenous expression of mutant Atxn3 leads to regional transcriptional dysregulation of oligodendrocyte maturation markers in Knock-In models of SCA3. We then investigated the spatiotemporal progression of mature oligodendrocyte transcriptional dysregulation in an overexpression SCA3 mouse model and how it relates to the onset of motor impairment. We further determined that regional reduction in mature oligodendrocyte cell counts in SCA3 mice over time parallels the onset and progression of brain atrophy in SCA3 patients. This work emphasizes the prospective contributions of disease-associated oligodendrocyte signatures to regional vulnerability and could inform timepoints and target regions imperative for biomarker assessment and therapeutic intervention in several neurodegenerative diseases.

Keywords: MJD; Machado-Joseph disease; SCA3; myelination; polyglutamine (polyQ) diseases.

FIGURE 1

Human post-mortem myelin staining is reduced in the cerebellum and unchanged in the frontal cortex of patients with SCA3 relative to control subject samples. Representative images show post-mortem human (A) cerebellum and (B) cortex stained with Luxol Fast Blue from controls subjects (left) relative to patients with SCA3 (right). Scale bar 200 μm. Myelin basic protein (MBP) is significantly decreased in the cerebellum of SCA3 patients compared to approximately age-matched controls (C), while MBP varies immensely and is not significantly different between SCA3 and control samples in the frontal cortex (D). Student’s unpaired t-test performed after verifying normal distribution of data via the Shapiro–Wilk normality test. *p < 0.05.

FIGURE 2

Regional oligodendrocyte transcriptional dysregulation is observed in the Knock-In SCA3 mouse models. QPCR analysis of oligodendrocyte maturation markers in 48-week hemizygous (Q82/WT) KIQ82 mice relative to WT/WT in panel (A) brainstem, (B) spinal cord, (C) cerebellum, and (D) forebrain. QPCR analysis of oligodendrocyte maturation markers in 16-week hemizygous (Q300/WT) KIQ300 mice relative to WT/WT in panel (E) brainstem, (F) spinal cord, (G) cerebellum, and (H) forebrain. Each data point represents an independent mouse per genotype. Unpaired Student’s t-tests or Mann–Whitney tests were performed, depending on normality distribution as determined by Shapiro–Wilk tests: *p < 0.05, ^**p < 0.01, ^***p < 0.001, ns, not significant.

FIGURE 3

Onset of behavioral deficits in Q84 mice associates with mature oligodendrocyte transcript dysregulation. (A) Schematic of the recording timepoints for four phenotypic characterizations of Q84 mice. Tissue was collected for qPCR analysis at 3-, 4-, 8-, and 16-weeks-old. Q84;Mobp-eGFP tissue was collected for immunohistochemical analysis at 4-, 8-, and 16-weeks-old. Recordings of panel (B) weight, (C) total locomotor activity (x-y beam breaks), and (D) rearing activity (z beam breaks) in hemizygous (Q84/WT) and homozygous (Q84/Q84) Q84 mice relative to their WT littermates (WT/WT) at 2-, 3-, 4-, 8-, and 14-weeks-old (n = 16–20 mice/genotype. Mixed-effects analysis with a post-hoc Tukey’s multiple comparison test was performed for behavioral analyses *p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001; no asterisk, not significant. Red asterisks indicate comparison of Q84/Q84 to WT/WT. (E,F) QPCR analysis of oligodendrocyte maturation markers (Plp1, Mal, Mobp) in Q84 mice in the (E) brainstem, (F) spinal cord, (G) cerebellum, and (H) forebrain. Each data points represents an independent mouse per genotype. For qPCR analysis, unpaired Student’s t-tests or Mann–Whitney tests were performed, depending on normality distribution as determined by Shapiro–Wilk tests: *p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001; ns, not significant.

FIGURE 4

Mature oligodendrocytes are reduced in a spatiotemporal pattern that corresponds to disease progression. (A) Mouse cross between Q84 SCA3 mice and Mobp-eGFP reporter mice. (B) Depiction of where images were taken for histologically assessed regions. (C) Representative images of Mobp-eGFP+ mature oligodendrocytes in the pons, DCN, CST, CC, cortex, and striatum at 4 weeks (left) and 16 weeks of age (right). Scale bar 50 μm. (D) Quantification of mature oligodendrocytes in the pons, DCN, CST, CC, cortex, and striatum of Q84;Mobp-eGFP+ mice at 4 (left), 8 (middle), and 16 weeks of age (right) (n = 4–6 mice/genotype, 3 images/mouse). One-way ANOVA with a post-hoc Tukey’s multiple comparisons test was performed: *p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001; ns, not significant.

FIGURE 5

Spatiotemporal progression of mature oligodendrocyte transcript dysregulation and cell count reduction in mouse models of SCA3. (Top) Temporal progression of regional transcriptional downregulation of mature oligodendrocyte markers as observed in mouse models. (Bottom) Temporal reduction of regional mature oligodendrocyte cell counts as observed in mouse models. Horizontal arrow represents disease progression over time. Transparency and shade of regional colors represent magnitude of oligodendrocyte maturation impairments (darker/less transparent = more affected, lighter/more transparent = less affected).