Phosphorylated tau fluid biomarker sites recognize earlier neurofibrillary tangle maturity levels in the postmortem Alzheimer’s disease brain

Abstract

Alzheimer’s disease (AD) biomarkers have become increasingly more reliable in predicting AD pathology. While phosphorylated tau fluid biomarkers have been studied for over 20 years, there is a lack of deep characterization of these sites in the postmortem brain. Neurofibrillary tangle-bearing neurons, one of the major neuropathologic hallmarks of AD, undergo morphologic changes that mature along a continuum as hyperphosphorylated tau aggregates. To facilitate interpretation of phosphorylated tau sites as an early fluid biomarker, our goal was to characterize which neurofibrillary tangle maturity levels (pretangle, intermediary 1, mature tangle, intermediary 2, and ghost tangle) they recognize. We queried the Florida Autopsied Multi-Ethnic (FLAME) cohort for cases from Braak stages I-VI. We excluded non-AD pathologies and tauopathies. A total of 24 cases, 2 males and 2 females for each Braak stage, were selected. We performed immunohistochemistry on the posterior hippocampus using antibodies directed towards phospho (p) threonine (T) 181, pT205, pT217, and pT231. Slides were digitized to enable quantification of tau burden. To examine differences in regional vulnerability between CA1 and subiculum, we developed a semi-quantitative system to rank the frequency of each neurofibrillary tangle maturity level. We identified all neurofibrillary tangle maturity levels at least once for each phosphorylated tau site. Primarily earlier neurofibrillary tangle maturity levels (pretangle, intermediary 1, mature tangle) were recognized for all phosphorylated tau sites. There was an increase in tau burden in the subiculum compared to CA1; however, this was attenuated compared to thioflavin-S positive tangle counts. On a global scale, tau burden generally increased with each Braak stage. These results provide neurobiologic evidence that these phosphorylated tau fluid biomarker sites are present during earlier neurofibrillary tangle maturity levels. This may help explain why these phosphorylated tau biomarker sites are observed before symptom onset in fluids.

Cellular localization of p-tau217 in brain and its association with p-tau217 plasma levels

“…Our study shows that p-tau217 can be found within NFTs and NTs also containing the p-tau variants p-tau181, 231, 202, 202/205, and 369/404, a finding suggesting that phosphorylation of tau at site Thr217 often occurs at the same time as phosphorylation at other tau sites. This idea is supported, not only by previous studies demonstrating strong correlations between levels of p-tau217 and various p-tau variants in plasma and CSF [4,10,11,19,20] but also by a recently published study showing that p-tau217 can be found in the same early tangle maturation stages as many of the other p-tau variants including p-tau181, 231, 205 [18]. Our study adds to these studies by demonstrating subtle differences in the coverage of p-tau217 immunoreactivity in NFT and NT compared to the other p-tau variants.…”

Localization, induction, and cellular effects of tau-phospho-threonine 217

“…Doing so required optimization of an anti-tau pT217 antibody that can recognize other proteins with the identical epitope, an issue that likely applies to other antibodies directed against tau pT217 . Consistent with prior immunohistological studies of human brain [12, 13], we found that tau pT217 first appears in limbic structure neurons at Braak stages I-II, that its presence in the somatodendritic compartment and neuropil threads increases with disease progression, and that its cellular and subcellular distribution overlaps partially with those of other phospho-tau variants associated with AD and other tauopathies. An immunofluorescence study of aged CVN mice indicates that the tau pT217 distribution in this particular AD model mouse strain is similar to what is seen in mid to late-stage human AD, highlighting the potential utility of CVN mice for further studies of tau pT217 .…”

“…Our immunohistological analysis of tau pT217 in human brain yielded results similar to those reported earlier by two other groups [12, 13]. At Braak stage 0 we did not detect any tau pT217 (Supplementary Figure 5), but occasional immunostaining of axons, neuropil threads and neuronal cell bodies was evident by Braak stages I-II (Figure 1), and more widespread by Braak stages III-VI (Figures 2 and 3).…”

“…Indeed, a study that was reported at the Tau 2022 conference but has not been published yet prompted a tentative conclusion that tau pT217 levels in CSF define a “clock” that predicts AD progression over a 30 year time span (P-Tau217 Clock). Despite these encouraging advances about the diagnostic value of tau pT217 for AD, to the best of our knowledge only one peer-reviewed paper [12] and one published pre-print [13] have addressed the histopathology of tau pT217 , and not a single study published until now has investigated what tau pT217 does to neurons or what provokes its intracellular accumulation.…”

“…Prime examples of promising plasma biomarkers are phosphorylated forms of the neuron-specific, axon-enriched, microtubule-associated protein, tau, particularly tau pT181 , tau pT217 and tau pT231 [6–9]. Though several studies have focused on the localization and functional properties of tau pT181 [10] and tau pT231 [11], little is known about tau pT217 other than its promise as an early fluid-based biomarker for AD, and two reports about its histological distribution in AD brain [12, 13].…”