Seeding human tau knock‐in neurons with human derived pathological tau results in the formation of pathogenic conformations of tau

Abstract

AbstractBackgroundOne of the pathological hallmarks of Alzheimer’s disease (AD) is the accumulation of pathological tau inclusions. Conformational exposure of an N‐terminal motif in tau known as the phosphatase‐activating domain (PAD) occurs early in AD and other tauopathies and aberrant exposure of this motif disrupts axonal transport. Tau is capable of seeding aggregation in cells; however, the extent to which disease‐associated pathogenic conformations can be effectively seeded remains an area of active investigation. To help address this gap, we seeded aggregation in primary neurons and assessed known pathogenic tau conformations.MethodPrimary hippocampal neurons from human tau knock‐in mice were treated with pathological tau aggregates from human AD‐brain (AD‐tau) according to methods described in Narasimhan and Lee, 2017. The same purification protocol was used with non‐demented human brain tissue (CO) as a control. The AD‐tau and CO samples were characterized by transmission electron microscopy and biochemical assays that measure total tau and pathological modifications, such as phosphorylation (PHF1) and a conformation specific antibody (TNT1) that labels display of PAD. Neurons treated at 5 days in vitro were maintained for up to 28 days post‐treatment. Markers of pathological species of tau (PAD exposed and phospho‐tau) were used to assess the formation of disease‐associated forms of tau and changes in the numbers of neurons (MAP2) and astrocytes (GFAP) were measured.ResultThe AD‐derived material showed morphologies consistent with small pieces of the characteristic paired helical filaments of AD, phosphorylation at the PHF1 site, and the presence of oligomeric tau and PAD‐exposed tau species. We observed a robust dose and time‐dependent increase in TNT1 in AD‐tau‐treated neurons and ongoing studies are evaluating the presence of other pathological tau species. There were no significant effects on cell number in seeded cultures out to 28 days post‐treatment.ConclusionOur results suggest that primary neurons seeded with AD‐tau form inclusions consisting of PAD‐exposed tau, but overt degeneration does not occur within 28 days of seeding. The presence of PAD‐exposed tau but lack of cell loss could indicate that more time is needed and/or that the consequences of these tau species are more subtle.