The amyloid-β peptide (Aβ) plays a central pathophysiological role in Alzheimer's disease, but little is known about the concentration and dynamics of this secreted peptide in the extracellular space of the human brain. We used intracerebral microdialysis to obtain serial brain interstitial fluid (ISF) samples in 18 patients who were undergoing invasive intracranial monitoring after acute brain injury. We found a strong positive correlation between changes in brain ISF Aβ concentrations and neurological status, with Aβ concentrations increasing as neurological status improved and falling when neurological status declined. Brain ISF Aβ concentrations were also lower when other cerebral physiological and metabolic abnormalities reflected depressed neuronal function. Such dynamics fit well with the hypothesis that neuronal activity regulates extracellular Aβ concentration.

Objective While amyloid-beta (Aβ) peptide deposition into insoluble plaques is a pathological hallmark of Alzheimer’s disease, soluble oligomeric Aβ has been hypothesized to more directly underlie impaired learning and memory in dementia of the Alzheimer type. However, the lack of a sensitive, specific, and quantitative assay for Aβ oligomers has hampered rigorous tests of this hypothesis. Methods We developed a plate-based single molecule counting fluorescence immunoassay for oligomeric Aβ sensitive to low pg/ml concentrations of synthetic Aβ dimers using the same Aβ-specific monoclonal antibody to both capture and detect Aβ. The Aβ oligomer assay does not recognize monomeric Aβ, amyloid precursor protein, or other non-Aβ peptide oligomers. Results Aβ oligomers were detected in aqueous cortical lysates from patients with dementia of the Alzheimer type and non-demented patients with Aβ plaque pathology. However, Aβ oligomer concentrations in demented patients’ lysates were tightly correlated with Aβ plaque coverage (r=0.88), but this relationship was weaker in those from non-demented patients (r=0.30) despite equivalent Aβ plaque pathology. The ratio of Aβ oligomer levels to plaque density fully distinguished demented from non-demented patients, with no overlap between groups in this derived variable. Other Aβ and plaque measures did not distinguish demented from non-demented patients. Aβ oligomers were not detected in cerebrospinal fluid with this assay. Interpretation The results raise the intriguing hypothesis that the linkage between plaques and oligomers may be a key pathophysiological event underlying dementia of the Alzheimer type. This Aβ oligomer assay may be useful for many tests of the oligomer hypothesis.

It is generally accepted that cytokinin oxidases, which oxidatively remove cytokinin side chains to produce adenine and the corresponding isopentenyl aldehyde, play a major role in regulating cytokinin levels in planta. Partially purified fractions of cytokinin oxidase from various species have been studied for many years, but have yet to clearly reveal the properties of the enzyme or to define its biological significance. Details of the genomic organization of the recently isolated maize (Zea mays) cytokinin oxidase gene (ckx1) and some of its Arabidopsis homologs are now presented. Expression of an intronless ckx1 in Pichia pastoris allowed production of large amounts of recombinant cytokinin oxidase and facilitated detailed kinetic and cofactor analysis and comparison with the native enzyme. The enzyme is a flavoprotein containing covalently bound flavin adenine dinucleotide, but no detectable heavy metals. Expression of the oxidase in maize tissues is described.

Amyloid-beta (Aβ) plays a key role in the pathogenesis of Alzheimer’s disease (AD), but little is known about the proteoforms present in AD brain. We used high-resolution mass spectrometry to analyze intact Aβ from soluble aggregates and insoluble material in brains of six cases with severe dementia and pathologically confirmed AD. The soluble aggregates are especially relevant because they are believed to be the most toxic form of Aβ. We found a diversity of Aβ peptides, with 26 unique proteoforms including various N- and C-terminal truncations. N- and C-terminal truncations comprised 73% and 30%, respectively, of the total Aβ proteoforms detected. The Aβ proteoforms segregated between the soluble and more insoluble aggregates with N-terminal truncations predominating in the insoluble material and C- terminal truncations segregating into the soluble aggregates. In contrast, canonical Aβ comprised the minority of the identified proteoforms (15.3%) and did not distinguish between the soluble and more insoluble aggregates. The relative abundance of many truncated Aβ proteoforms did not correlate with post-mortem interval, suggesting they are not artefacts. This heterogeneity of Aβ proteoforms deepens our understanding of AD and offers many new avenues for investigation into pathological mechanisms of the disease, with implications for therapeutic development.

Summary The apolipoprotein E (APOE) gene is the strongest genetic risk factor for late-onset Alzheimer disease. Previous studies suggest reduction of apoE levels through genetic manipulation can reduce Aβ pathology. However, it is not clear how reduction of apoE levels after birth would affect amyloid deposition. We utilize an antisense oligonucleotide (ASO) to reduce apoE expression in the brains of APP/PS1-21 mice homozygous for the APOE-ε4 or APOE-ε3 allele. ASO treatment starting after birth led to a significant decrease in Aβ pathology when assessed at 4 months. Interestingly, ASO treatment starting at the onset of amyloid deposition led to an increase in Aβ plaque size and a reduction in plaque-associated neuritic dystrophy with no change in overall plaque load. These results suggest that lowering apoE levels prior to plaque deposition can strongly affect the initiation of Aβ pathology while lowering apoE after Aβ seeding modulates plaque size and toxicity.

Little is known about central neuron mechanosensation. Gu et al. report that micromechanical stress preferentially induces rapid and reversible varicosity formation in the axons but not the dendrites of hippocampal neurons through activation of mechanosensitive ion channels and microtubule disassembly.

Axonal injury is believed to be a major determinant of adverse outcomes following traumatic brain injury. However, it has been difficult to assess acutely the severity of axonal injury in human traumatic brain injury patients. We hypothesized that microdialysis-based measurements of the brain extracellular fluid levels of tau and neurofilament light chain, two low molecular weight axonal proteins, could be helpful in this regard. To test this hypothesis, 100 kDa cut-off microdialysis catheters were placed in 16 patients with severe traumatic brain injury at two neurological/neurosurgical intensive care units. Tau levels in the microdialysis samples were highest early and fell over time in all patients. Initial tau levels were >3-fold higher in patients with microdialysis catheters placed in pericontusional regions than in patients in whom catheters were placed in normal-appearing right frontal lobe tissue (P = 0.005). Tau levels and neurofilament light-chain levels were positively correlated (r = 0.6, P = 0.013). Neurofilament light-chain levels were also higher in patients with pericontusional catheters (P = 0.04). Interestingly, initial tau levels were inversely correlated with initial amyloid-β levels measured in the same samples (r = -0.87, P = 0.000023). This could be due to reduced synaptic activity in areas with substantial axonal injury, as amyloid-β release is closely coupled with synaptic activity. Importantly, high initial tau levels correlated with worse clinical outcomes, as assessed using the Glasgow Outcome Scale 6 months after injury (r = -0.6, P = 0.018). Taken together, our data add support for the hypothesis that axonal injury may be related to long-term impairments following traumatic brain injury. Microdialysis-based measurement of tau levels in the brain extracellular space may be a useful way to assess the severity of axonal injury acutely in the intensive care unit. Further studies with larger numbers of patients will be required to assess the reproducibility of these findings and to determine whether this approach provides added value when combined with clinical and radiological information.

There are currently few approved effective treatments for SARS-CoV-2, the virus responsible for the COVID-19 pandemic. Nanobodies are 12–15 kDa single-domain antibody fragments that can be delivered by inhalation and are amenable to relatively inexpensive large scale production compared to other biologicals. We have isolated nanobodies that bind to the SARS-CoV-2 spike protein receptor binding domain and block spike protein interaction with the angiotensin converting enzyme 2 (ACE2) with 1–5 nM affinity. The lead nanobody candidate, NIH-CoVnb-112, blocks SARS-CoV-2 spike pseudotyped lentivirus infection of HEK293 cells expressing human ACE2 with an EC50 of 0.3 µg/mL. NIH-CoVnb-112 retains structural integrity and potency after nebulization. Furthermore, NIH-CoVnb-112 blocks interaction between ACE2 and several high affinity variant forms of the spike protein. These nanobodies and their derivatives have therapeutic, preventative, and diagnostic potential.