Overexpression of human wtTDP-43 causes impairment in hippocampal plasticity and behavioral deficits in CAMKII-tTa transgenic mouse model.

AIMS: The current study utilizes the adeno-associated viral gene transfer system in the CAMKIIα-tTA mouse model to overexpress human wild type TDP-43 (wtTDP-43) and α-synuclein (α-Syn) proteins. The co-existence of these proteins is evident in the pathology of neurodegenerative disorders such as frontotemporal lobar degeneration (FTLD), Parkinson disease (PD), and dementia with Lewy bodies (DLB). METHODS: The novel bicistronic recombinant adeno-associated virus (rAAV) serotype 9 drives wtTDP-43 and α-Syn expression in the hippocampus via "TetO" CMV promoter. Behavior, electrophysiology, and biochemical and histological assays were used to validate neuropathology. RESULTS: We report that overexpression of wtTDP-43 but not α-Syn contributes to hippocampal CA2-specific pyramidal neuronal loss and overall hippocampal atrophy. Further, we report a reduction of hippocampal long-term potentiation and decline in learning and memory performance of wtTDP-43 expressing mice. Elevated wtTDP-43 levels induced selective degeneration of Purkinje cell protein 4 (PCP-4) positive neurons while both wtTDP-43 and α-Syn expression reduced subsets of the glutamate receptor expression in the hippocampus. CONCLUSIONS: Overall, our findings suggest the significant vulnerability of hippocampal neurons toward elevated wtTDP-43 levels possibly via PCP-4 and GluR-dependent calcium signaling pathways. Further, we report that wtTDP-43 expression induced selective CA2 subfield degeneration, contributing to the deterioration of the hippocampal-dependent cognitive phenotype.

TDP-43 mediated blood-brain barrier permeability and leukocyte infiltration promote neurodegeneration in a low-grade systemic inflammation mouse model.

BACKGROUND: Neuronal cytoplasmic inclusions containing TAR DNA-binding protein 43 (TDP-43) are a neuropathological feature of several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and Alzheimer's Disease (AD). Emerging evidence also indicates that systemic inflammation may be a contributor to the pathology progression of these neurodegenerative diseases. METHODS: To investigate the role of systemic inflammation in the progression of neuronal TDP-43 pathology, AAV9 particles driven by the UCHL1 promoter were delivered to the frontal cortex of wild-type aged mice via intracranial injections to overexpress TDP-43 or green fluorescent protein (GFP) in corticospinal motor neurons. Animals were then subjected to a low-dose (500 µg/kg) intraperitoneal E. coli lipopolysaccharide (LPS) administration challenge for 2 weeks to mimic a chronically altered low-grade systemic inflammatory state. Mice were then subjected to neurobehavioral studies, followed by biochemical and immunohistochemical analyses of the brain tissue. RESULTS: In the present study, we report that elevated neuronal TDP-43 levels induced microglial and astrocytic activation in the cortex of injected mice followed by increased RANTES signaling. Moreover, overexpression of TDP-43 exerted abundant mouse immunoglobulin G (IgG), CD3, and CD4+ T cell infiltration as well as endothelial and pericyte activation suggesting increased blood-brain barrier permeability. The BBB permeability in TDP-43 overexpressing brains yielded the frontal cortex vulnerable to the systemic inflammatory response following LPS treatment, leading to marked neutrophil infiltration, neuronal loss, reduced synaptosome-associated protein 25 (SNAP-25) levels, and behavioral impairments in the radial arm water maze (RAWM) task. CONCLUSIONS: These results reveal a novel role for TDP-43 in BBB permeability and leukocyte recruitment, indicating complex intermolecular interactions between an altered systemic inflammatory state and pathologically prone TDP-43 protein to promote disease progression.

Accumulation of C-terminal cleaved tau is distinctly associated with cognitive deficits, synaptic plasticity impairment, and neurodegeneration in aged mice.

C-terminal cleaved tau at D421 (∆D421-tau) accumulates in the brains of Alzheimer's disease (AD) patients. However, it is unclear how tau truncation, an understudied tau post-translational modification, contributes to AD pathology and progression. Utilizing an adeno-associated virus (AAV) gene delivery-based approach, we overexpressed full-length tau (FL-tau) and ∆D421-tau in 4- and 12-month-old mice for 4 months to study the neuropathological impact of accumulation in young adult (8-month) and middle-aged (16-month) mice. Overall, we show that independent of the tau species, age was an important factor facilitating tau phosphorylation, oligomer formation, and deposition into silver-positive tangles. However, mice overexpressing ∆D421-tau exhibited a distinct phosphorylation profile to those overexpressing FL-tau and increased tau oligomerization in the middle-age group. Importantly, overexpression of ∆D421-tau, but not FL-tau in middle-aged mice, resulted in pronounced cognitive impairments and hippocampal long-term potentiation deficits. While both FL-tau and ∆D421-tau induced neuronal loss in mice with age, ∆D421-tau led to significant neuronal loss in the CA3 area of the hippocampus and medial entorhinal cortex compared to FL-tau. Based on our data, we conclude that age increases the susceptibility to neuronal degeneration associated with ΔD421-tau accumulation. Our findings suggest that ΔD421-tau accumulation contributes to synaptic plasticity and cognitive deficits, thus representing a potential target for tau-associated pathologies.

Hypusination of Eif5a regulates cytoplasmic TDP-43 aggregation and accumulation in a stress-induced cellular model.

TAR DNA-binding protein 43 (TDP-43) is a nuclear RNA/DNA binding protein involved in mRNA metabolism. Aberrant mislocalization to the cytoplasm and formation of phosphorylated/aggregated TDP-43 inclusions remains the hallmark pathology in a spectrum of neurodegenerative diseases, including frontotemporal disorders and Alzheimer's disease. Eukaryotic Translation Initiation Factor 5A undergoes a unique post-translation modification of lysine to hypusine (K50), which determines eIF5A binding partners. We used a sodium arsenite-induced cellular stress model to investigate the role of hypusinated eIF5A (eIF5AHypK50) in governing TDP-43 cytoplasmic mislocalization and accumulation in stress granule. Our proteomics and functional data provide evidence that eIF5A interacts with TDP-43 in a hypusine-dependent manner. Additionally, we showed that following stress TDP-43 interactions with eIF5AHypK50 were induced both in the cytoplasm and stress granules. Pharmacological reduction of hypusination or mutations of lysine residues within the hypusine loop decreased phosphorylated and insoluble TDP-43 levels. The proteomic and biochemical analysis also identified nuclear pore complex importins KPNA1/2, KPNB1, and RanGTP as interacting partners of eIF5AHypK50. These findings are the first to provide a novel pathway and potential therapeutic targets that require further investigation in models of TDP-43 proteinopathies.

CCL2 Overexpression in the Brain Promotes Glial Activation and Accelerates Tau Pathology in a Mouse Model of Tauopathy.

Innate immune activation is a major contributor to Alzheimer's Disease (AD) pathophysiology, although the mechanisms involved are poorly understood. Chemokine C-C motif ligand (CCL) 2 is produced by neurons and glial cells and is upregulated in the AD brain. Transgene expression of CCL2 in mouse models of amyloidosis produces microglia-induced amyloid ß oligomerization, a strong indication of the role of these activation pathways in the amyloidogenic processes of AD. We have previously shown that CCL2 polarizes microglia in wild type mice. However, how CCL2 signaling contributes to tau pathogenesis remains unknown. To address this question, CCL2 was delivered via recombinant adeno-associated virus serotype 9 into both cortex and hippocampus of a mouse model with tau pathology (rTg4510). We report that CCL2 overexpression aggravated tau pathology in rTg4510 as shown by the increase in Gallyas stained neurofibrillary tangles as well as phosphorylated tau-positive inclusions. In addition, biochemical analysis showed a reduction in the levels of detergent-soluble tau species followed by increase in the insoluble fraction, indicating a shift toward larger tau aggregates. Indeed, increased levels of high molecular weight species of phosphorylated tau were found in the mice injected with CCL2. We also report that worsening of tau pathology following CCL2 overexpression was accompanied by a distinct inflammatory response. We report an increase in leukocyte common antigen (CD45) and Cluster of differentiation 68 (CD68) expression in the brain of rTg4510 mice without altering the expression levels of a cell-surface protein Transmembrane Protein 119 (Tmem119) and ionized calcium-binding adaptor molecule 1 (Iba-1) in resident microglia. Furthermore, the analysis of cytokines in brain extract showed a significant increase in interleukin (IL)-6 and CCL3, while CCL5 levels were decreased in CCL2 mice. No changes were observed in IL-1α, IL-1ß, TNF-α. IL-4, Vascular endothelial growth factor-VEGF, IL-13 and CCL11. Taken together our data report for the first time that overexpression of CCL2 promotes the increase of pathogenic tau species and is associated with glial neuroinflammatory changes that are deleterious. We propose that these events may contribute to the pathogenesis of Alzheimer's disease and other tauopathies.

Tau depletion prevents progressive blood-brain barrier damage in a mouse model of tauopathy.

INTRODUCTION: The blood-brain barrier (BBB) is damaged in tauopathies, including progressive supranuclear palsy (PSP) and Alzheimer's disease (AD), which is thought to contribute to pathogenesis later in the disease course. In AD, BBB dysfunction has been associated with amyloid beta (Aß) pathology, but the role of tau in this process is not well characterized. Since increased BBB permeability is found in tauopathies without Aß pathology, like PSP, we suspected that tau accumulation alone could not only be sufficient, but even more important than Aß for BBB damage. RESULTS: Longitudinal evaluation of brain tissue from the tetracycline-regulatable rTg4510 tau transgenic mouse model showed progressive IgG, T cell and red blood cell infiltration. The Evans blue (EB) dye that is excluded from the brain when the BBB is intact also permeated the brains of rTg4510 mice following peripheral administration, indicative of a bonafide BBB defect, but this was only evident later in life. Thus, despite the marked brain atrophy and inflammation that occurs earlier in this model, BBB integrity is maintained. Interestingly, BBB dysfunction emerged at the same time that perivascular tau emerged around major hippocampal blood vessels. However, when tau expression was suppressed using doxycycline, BBB integrity was preserved, suggesting that the BBB can be stabilized in a tauopathic brain by reducing tau levels. CONCLUSIONS: For the first time, these data demonstrate that tau alone can initiate breakdown of the BBB, but the BBB is remarkably resilient, maintaining its integrity in the face of marked brain atrophy, neuroinflammation and toxic tau accumulation. Moreover, the BBB can recover integrity when tau levels are reduced. Thus, late stage interventions targeting tau may slow the vascular contributions to cognitive impairment and dementia that occur in tauopathies.