Cryo-ET and Sub-Volume Analysis of Fibrillar Pathology in the Neurodegenerative Human Brain

Structural analyses of amyloid fibrils sourced from brain tissue have unveiled distinct fibril polymorphs associated with specific diseases. However, the mechanisms behind these polymorphs remain elusive due to the lack of a method that connects the cellular context, composition, and ultrastructure of individual aggregates to their fibril polymorphs.

In this study, we introduce a process for in situ cryo-electron tomography (cryo-ET) to investigate amyloids within singular aggregates within human neurodegenerative postmortem brains. Employing chemically fixed specimens, we examined tau fibrils in the hippocampus of an Alzheimer's disease donor and -synuclein fibrils within a glial nuclear inclusion in the cingulate gyrus of a multiple system atrophy donor.

Through subtomogram averaging and helical reconstruction of fibril subvolumes, we generated low-resolution (~30-35A) density maps, which were juxtaposed with pre-existing ex vivo structures. The maintenance of ultrastructure facilitated a detailed examination of pathology, including a striking instance of an alpha-synuclein fibril intruding the nuclear envelope, offering mechanistic understanding of intranuclear aggregate formation.

By integrating cryo-ET with existing high-resolution fibril structures that lack spatial context, this method provides a potent means to comprehend amyloid polymorphism in neurodegenerative diseases, thereby propelling anti-amyloid therapies and diagnostic tools.