Tau seeding in chronic traumatic encephalopathy parallels disease severity.

Chronic traumatic encephalopathy (CTE), a neurodegenerative tauopathy, is associated with behavioral, mood and cognitive impairment, including dementia. Tauopathies are neurodegenerative diseases whose neuropathological phenotypes are characterized by distinct histopathologic features of tau pathology, which progressively deposit throughout the brain. In certain tauopathies, especially Alzheimer's disease (AD), tau deposition appears to follow brain network connections. Experimental evidence suggests that the progression of tau pathology in humans, mouse and cell models could be explained by tau seeds that adopt distinct conformations and serve as templates for their own amplification to mediate transcellular propagation of pathology. Tau seeds are efficiently detected by the induction of aggregation in cell-based "biosensors" that express tau repeat domain (RD) with a disease-associated mutation (P301S) fused to complementary fluorescent protein tags (cyan and yellow fluorescent protein). Biosensors enable quantification of tau seeding in fixed and fresh-frozen brain tissue. Phospho-tau deposition in CTE follows progressive stages (I-IV), but the relationship of seeding to this deposition is unclear. We have used an established biosensor assay to independently quantify tau seeding as compared to AT8 phospho-tau histopathology in thin sections of fixed tissues of 11 brain regions from 27 patients with CTE, 5 with other tauopathies, and 5 negative controls. In contrast to prior studies of AD, we detected tau seeding late in the course of CTE (predominantly stages III and IV). It was less anatomically prevalent than AT8-positive inclusions, which were relatively widespread. We especially observed seeding in the limbic system (amygdala, thalamus, basal ganglia), which may explain the dominant cognitive and behavior impairments that characterize CTE.

Tau seeding activity begins in the transentorhinal/entorhinal regions and anticipates phospho-tau pathology in Alzheimer's disease and PART.

Alzheimer's disease (AD) is characterized by accumulation of tau neurofibrillary tangles (NFTs) and, according to the prion model, transcellular propagation of pathological "seeds" may underlie its progression. Staging of NFT pathology with phospho-tau antibody is useful to classify AD and primary age-related tauopathy (PART) cases. The locus coeruleus (LC) shows the earliest phospho-tau signal, whereas other studies suggest that pathology begins in the transentorhinal/entorhinal cortices (TRE/EC). The relationship of tau seeding activity, phospho-tau pathology, and progression of neurodegeneration remains obscure. Consequently, we employed an established cellular biosensor assay to quantify tau seeding activity in fixed human tissue, in parallel with AT8 phospho-tau staining of immediately adjacent sections. We studied four brain regions from each of n = 247 individuals across a range of disease stages. We detected the earliest and most robust seeding activity in the TRE/EC. The LC did not uniformly exhibit seeding activity until later NFT stages. We also detected seeding activity in the superior temporal gyrus (STG) and primary visual cortex (VC) at stages before NFTs and/or AT8-immunopositivity were detectable. AD and putative PART cases exhibited similar patterns of seeding activity that anticipated histopathology across all NFT stages. Our findings are consistent with the prion model and suggest that pathological seeding activity begins in the TRE/EC rather than in the LC. In the analysis of tauopathy, quantification of seeding activity may offer an important addition to classical histopathology.

Characterization of tau prion seeding activity and strains from formaldehyde-fixed tissue.

Tauopathies such as Alzheimer's disease (AD) feature progressive intraneuronal deposition of aggregated tau protein. The cause is unknown, but in experimental systems trans-cellular propagation of tau pathology resembles prion pathogenesis. Tau aggregate inoculation into mice produces transmissible pathology, and tau forms distinct strains, i.e. conformers that faithfully replicate and create predictable patterns of pathology in vivo. The prion model predicts that tau seed formation will anticipate neurofibrillary tau pathology. To test this idea requires simultaneous assessment of seed titer and immunohistochemistry (IHC) of brain tissue, but it is unknown whether tau seed titer can be determined in formaldehyde-fixed tissue. We have previously created a cellular biosensor system that uses flow cytometry to quantify induced tau aggregation and thus determine seed titer. In unfixed tissue from PS19 tauopathy mice that express 1 N,4R tau (P301S), we have measured tau seeding activity that precedes the first observable histopathology by many months. Additionally, in fresh frozen tissue from human AD subjects at early to mid-neurofibrillary tangle stages (NFT I-IV), we have observed tau seeding activity in cortical regions predicted to lack neurofibrillary pathology. However, we could not directly compare the same regions by IHC and seeding activity in either case. We now describe a protocol to extract and measure tau seeding activity from small volumes (.04 mm3) of formaldehyde-fixed tissue immediately adjacent to that used for IHC. We validated this method with the PS19 transgenic mouse model, and easily observed seeding well before the development of phospho-tau pathology. We also accurately isolated two tau strains, DS9 and DS10, from fixed brain tissues in mice. Finally, we have observed robust seeding activity in fixed AD brain, but not controls. The successful coupling of classical IHC with seeding and strain detection should enable detailed study of banked brain tissue in AD and other tauopathies.

Tau Prion Strains Dictate Patterns of Cell Pathology, Progression Rate, and Regional Vulnerability In Vivo.

Tauopathies are neurodegenerative disorders that affect distinct brain regions, progress at different rates, and exhibit specific patterns of tau accumulation. The source of this diversity is unknown. We previously characterized two tau strains that stably maintain unique conformations in vitro and in vivo, but did not determine the relationship of each strain to parameters that discriminate between tauopathies such as regional vulnerability or rate of spread. We have now isolated and characterized 18 tau strains in cells based on detailed biochemical and biological criteria. Inoculation of PS19 transgenic tau (P301S) mice with these strains causes strain-specific intracellular pathology in distinct cell types and brain regions, and induces different rates of network propagation. In this system, strains alone are sufficient to account for diverse neuropathological presentations, similar to those that define human tauopathies. Further study of these strains can thus establish a structural logic that governs these biological effects.

Prions and Protein Assemblies that Convey Biological Information in Health and Disease.

Prions derived from the prion protein (PrP) were first characterized as infectious agents that transmit pathology between individuals. However, the majority of cases of neurodegeneration caused by PrP prions occur sporadically. Proteins that self-assemble as cross-beta sheet amyloids are a defining pathological feature of infectious prion disorders and all major age-associated neurodegenerative diseases. In fact, multiple non-infectious proteins exhibit properties of template-driven self-assembly that are strikingly similar to PrP. Evidence suggests that like PrP, many proteins form aggregates that propagate between cells and convert cognate monomer into ordered assemblies. We now recognize that numerous proteins assemble into macromolecular complexes as part of normal physiology, some of which are self-amplifying. This review highlights similarities among infectious and non-infectious neurodegenerative diseases associated with prions, emphasizing the normal and pathogenic roles of higher-order protein assemblies. We propose that studies of the structural and cellular biology of pathological versus physiological aggregates will be mutually informative.

Distinct tau prion strains propagate in cells and mice and define different tauopathies.

Prion-like propagation of tau aggregation might underlie the stereotyped progression of neurodegenerative tauopathies. True prions stably maintain unique conformations ("strains") in vivo that link structure to patterns of pathology. We now find that tau meets this criterion. Stably expressed tau repeat domain indefinitely propagates distinct amyloid conformations in a clonal fashion in culture. Reintroduction of tau from these lines into naive cells reestablishes identical clones. We produced two strains in vitro that induce distinct pathologies in vivo as determined by successive inoculations into three generations of transgenic mice. Immunopurified tau from these mice recreates the original strains in culture. We used the cell system to isolate tau strains from 29 patients with 5 different tauopathies, finding that different diseases are associated with different sets of strains. Tau thus demonstrates essential characteristics of a prion. This might explain the phenotypic diversity of tauopathies and could enable more effective diagnosis and therapy.

Prion-like propagation of protein aggregation and related therapeutic strategies.

Many neurodegenerative diseases are characterized by the progressive accumulation of aggregated protein. Recent evidence suggests the prion-like propagation of protein misfolding underlies the spread of pathology observed in these diseases. This review traces our understanding of the mechanisms that underlie this phenomenon and discusses related therapeutic strategies that derive from it.

Effects of sildenafil on nigrostriatal dopamine neurons in a murine model of Parkinson's disease.

The objective of this study was to determine if the phosphodiesterase 5 (PDE-5) inhibitor, sildenafil, could be used as a neuroprotective agent in a chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) murine model of Parkinson's disease. The underlying hypothesis of these studies is that blockade of PDE-5 catabolism of cGMP will attenuate the loss of nigrostriatal dopamine (NSDA) neurons following chronic neurotoxin exposure. Chronic MPTP-treated mice were administered sildenafil using three different regimens. Animals were: 1) treated with sildenafil and then exposed to chronic MPTP; 2) treated concurrently with sildenafil and MPTP; and 3) first exposed to MPTP and subsequently treated with sildenafil. End points of neurotoxicity included dopamine (DA) and tyrosine hydroxylase (TH) concentrations in NSDA axon terminals in the striatum, and stereological cell counts of TH immunoreactive neurons in the substantia nigra. Results reveal that sildenafil did not prevent neurotoxicity produced by chronic MPTP exposure regardless of the treatment paradigms employed. On the other hand, sildenafil did not produce any deleterious effect on NSDA neuron function nor did it potentiate the neurotoxic effects of MPTP. These results suggest that sildenafil would not accelerate DA cell loss when used as a treatment for erectile dysfunction in men diagnosed with Parkinson's disease.