Pathological aggregation of the microtubule-associated protein tau and subsequent accumulation of neurofibrillary tangles (NFTs) or other tau-containing inclusions are defining histopathological features of many neurodegenerative diseases, which are collectively known as tauopathies. Due to conflicting results regarding a correlation between the presence of NFTs and disease progression, the mechanism linking pathological tau aggregation with cell death is poorly understood. An emerging view is that NFTs are not the toxic entity in tauopathies; rather, tau intermediates between monomers and NFTs are pathogenic. Several proteins associated with neurodegenerative diseases, such as β-amyloid (Aβ) and α-synuclein, have the tendency to form pore-like amyloid structures (annular protofibrils, APFs) that mimic the membrane-disrupting properties of pore-forming protein toxins. The present study examined the similarities of tau APFs with other tau amyloid species and showed for the first time the presence of tau APFs in brain tissue from patients with progressive supranuclear palsy (PSP) and dementia with Lewy bodies (DLB), as well as in the P301L mouse model, which overexpresses mutated tau. Furthermore, we found that APFs are preceded by tau oligomers and do not go on to form NFTs, evading fibrillar fate. Collectively, our results demonstrate that in vivo APF formation depends on mutations in tau, phosphorylation levels, and cell type. These findings establish the pathological significance of tau APFs in vivo and highlight their suitability as therapeutic targets for several neurodegenerative tauopathies.

Alzheimer's disease (AD) is the most common cause of age-related dementia. Increasing evidence suggests that neuroinflammation mediated by microglia and astrocytes contributes to disease progression and severity in AD and other neurodegenerative disorders. During AD progression, resident microglia undergo proinflammatory activation, resulting in an increased capacity to convert resting astrocytes to reactive astrocytes. Therefore, microglia are a major therapeutic target for AD and blocking microglia-astrocyte activation could limit neurodegeneration in AD. Here we report that NLY01, an engineered exedin-4, glucagon-like peptide-1 receptor (GLP-1R) agonist, selectively blocks β-amyloid (Aβ)-induced activation of microglia through GLP-1R activation and inhibits the formation of reactive astrocytes as well as preserves neurons in AD models. In two transgenic AD mouse models (5xFAD and 3xTg-AD), repeated subcutaneous administration of NLY01 blocked microglia-mediated reactive astrocyte conversion and preserved neuronal viability, resulting in improved spatial learning and memory. Our study indicates that the GLP-1 pathway plays a critical role in microglia-reactive astrocyte associated neuroinflammation in AD and the effects of NLY01 are primarily mediated through a direct action on Aβ-induced GLP-1R+ microglia, contributing to the inhibition of astrocyte reactivity. These results show that targeting upregulated GLP-1R in microglia is a viable therapy for AD and other neurodegenerative disorders.

Greater than 160 missense mutations in copper-zinc superoxide dismutase-1 (SOD1) can cause amyotrophic lateral sclerosis (ALS). These mutations produce conformational changes that reveal novel antibody binding epitopes. A monoclonal antibody, clone C4F6 - raised against the ALS variant G93A of SOD1, has been identified as specifically recognizing a conformation shared by many ALS mutants of SOD1. Attempts to determine whether non-mutant SOD1 adopts a C4F6-reactive conformation in spinal tissues of sporadic ALS (sALS) patients has produced inconsistent results. To define the epitope recognized by C4F6, we tested its binding to a panel of recombinant ALS-SOD1 proteins expressed in cultured cells, producing data to suggest that the C4F6 epitope minimally contains amino acids 90-93, which are normally folded into a tight hairpin loop. Multiple van der Waals interactions between the 90-93 loop and a loop formed by amino acids 37-42, particularly a leucine at position 38, form a stable structure termed the β-plug. Based on published modeling predictions, we suggest that the binding of C4F6 to multiple ALS mutants of SOD1 occurs when the local structure within the β-plug, including the loop at 90-93, is destabilized. In using the antibody to stain tissues from transgenic mice or humans, the specificity of the antibody for ALS mutant SOD1 was influenced by antigen retrieval protocols. Using conditions that showed the best discrimination between normal and misfolded mutant SOD1 in cell and mouse models, we could find no obvious difference in C4F6 reactivity to spinal motor neurons between sALS and controls tissues.

Down syndrome (DS) is the leading genetic cause of intellectual disability and causes early-onset dementia and cerebellar hypoplasia. The prevalence of attention deficit hyperactivity disorder is elevated in children with DS. The aneuploid DS mouse model "Ts65Dn" shows prominent brain phenotypes, including learning and memory deficits, cerebellar hypoplasia, and locomotor hyperactivity. Previous studies indicate that impaired Sonic hedgehog (Shh) signaling contributes to neurological phenotypes associated with DS and neurodegenerative diseases. However, because of a lack of working inducible Shh knock-in mice, brain region-specific Shh overexpression and its effects on cognitive function have not been studied in vivo. Here, with Gli1-LacZ reporter mice, we demonstrated that Ts65Dn had reduced levels of Gli1, a sensitive readout of Shh signaling, in both hippocampus and cerebellum at postnatal day 6. Through site-specific transgenesis, we generated an inducible human Shh knock-in mouse, TRE-bi-hShh-Zsgreen1 (TRE-hShh), simultaneously expressing dually-lipidated Shh-Np and Zsgreen1 marker in the presence of transactivator (tTA). Double transgenic mice "Camk2a-tTA;TRE-hShh" and "Pcp2-tTA;TRE-hShh" induced Shh overexpression and activated Shh signaling in a forebrain and cerebellum, respectively, specific manner from the perinatal period. Camk2a-tTA;TRE-hShh normalized locomotor hyperactivity and improved learning and memory in 3-month-old Ts65Dn, mitigated early-onset severe cognitive impairment in 7-month-old Ts65Dn, and enhanced spatial cognition in euploid mice. Camk2a-tTA;TRE-hShh cohort maintained until 600days old showed that chronic overexpression of Shh in forebrain from the perinatal period had no effect on longevity of euploid or Ts65Dn. Pcp2-tTA;TRE-hShh did not affect cognition but mitigated the phenotype of cerebellar hypoplasia in Ts65Dn. Our study provides the first in vivo evidence that Shh overexpression from the perinatal period protects DS brain integrity and enhances learning and memory in normal mice, indicating the broad therapeutic potential of Shh ligand for other neurological conditions. Moreover, the first inducible hShh site-specific knock-in mouse could be widely used for spatiotemporal Shh signaling regulation.