Prion-like α-synuclein pathology in the brain of infants with Krabbe disease

Abstract

Krabbe disease is an infantile neurodegenerative disorder resulting from pathogenic variants in the GALC gene that causes accumulation of the toxic sphingolipid psychosine. GALC variants are also associated with Lewy body diseases, an umbrella term for age-associated neurodegenerative diseases in which the protein α-synuclein aggregates into Lewy bodies. To explore whether α-synuclein in Krabbe disease has pathological similarities to that in Lewy body disease, we performed an observational post-mortem study of Krabbe disease brain tissue (n = 4) compared to infant controls (n = 4) and identified widespread accumulations of α-synuclein. To determine whether α-synuclein in Krabbe disease brain displayed disease-associated pathogenic properties we evaluated its seeding capacity using the real-time quaking-induced conversion assay in two cases for which frozen tissue was available and strikingly identified aggregation into fibrils similar to those observed in Lewy body disease, confirming the prion-like capacity of Krabbe disease-derived α-synuclein. These observations constitute the first report of prion-like α-synuclein in the brain tissue of infants and challenge the putative view that α-synuclein pathology is merely an age-associated phenomenon, instead suggesting it results from alterations to biological pathways, such as sphingolipid metabolism. Our findings have important implications for understanding the mechanisms underlying Lewy body formation in Lewy body disease.

Keywords: Krabbe disease; Lewy body disease; sphingolipids; α-synuclein.

Figure 1

α-Synuclein in Krabbe disease. Photomicrographs of Krabbe disease (KD) cases demonstrating α-synuclein-immunoreactive globoid cells (A) that were variably positive for the autophagic marker p62 (B and C). Occasional neurons manifested punctate α-synuclein staining (D–F) and neurites that were immunoreactive for α-synuclein and p62 were variably present throughout the cortex of all cases (G I). Scale bars = 50 µm (A–C, G and H) 25 µm (D–F) and 20 µm (I).

Figure 2

Western blot and RT-QuIC of Krabbe disease brain tissue lysates. The TEAB-soluble fraction of KD temporal cortex grey matter lysates contained pS129 bands at ∼40 kDa (blue arrow) that were not observed in control cases or the DLB case (A). In white matter samples, KD cases had additional bands of pS129 at ∼20 kDa (blue arrow) that were not present in control or DLB cases (B). RT-QuIC of the TBS-soluble tissue fractions demonstrated that KD1 and KD3 grey matter gave positive reactions, defined as Thioflavin-T fluorescence >5 times the control standard deviation, at ∼55 h while no reaction was observed in white matter samples (C). Transmission electron microscopy of RT-QuIC end-products from grey matter of KD1 [D(i)] and KD3 [D(ii)] confirmed the presence of fibrillar structures. It is notable that KD1 RT-QuIC end-products led to elongated fibrils of ∼200–400 nm while KD3 end-products were typically 100–200 nm [E(i) and F(ii)]. Scale bars = 50 nm [D(i and ii)].