Faculty Bibliography

Diagnosis of asthma in obese individuals frequently relies on clinical history, as airflow by spirometry may remain normal. This study hypothesised that obese subjects with self-reported asthma and normal spirometry will demonstrate distinct clinical characteristics, metabolic comorbidities and enhanced small airway dysfunction as compared with healthy obese subjects. Spirometry, plethysmography and oscillometry data pre/post-bronchodilator were obtained in 357 obese subjects in three groups as follows: no asthma group (n=180), self-reported asthma normal spirometry group (n=126), and asthma obstructed spirometry group (n=51). To assess the effects of obesity related to reduced lung volume, oscillometry measurements were repeated during a voluntary inflation to predicted functional residual capacity (FRC). Dyspnoea was equally prevalent in all groups. In contrast, cough, wheeze and metabolic comorbidities were more frequent in the asthma normal spirometry and asthma obstructed spirometry groups versus the no asthma group (p<0.05). Despite similar body size, oscillometry measurements demonstrated elevated R_5-20 (difference between resistance at 5 and 20 Hz) in the no asthma and asthma normal spirometry groups (0.19±0.12; 0.23±0.13 kPa/(L·s^-1), p<0.05) but to a lesser degree than the asthma obstructed spirometry group (0.34±0.20 kPa/(L·s^-1), p<0.05). Differences between groups persisted post-bronchodilator (p<0.05). Following voluntary inflation to predicted FRC, R_5-20 in the no asthma and asthma normal spirometry groups fell to similar values, indicating a reversible process (0.11±0.07; 0.12±0.08 kPa/(L·s^-1), p=NS). Persistently elevated R_5-20 was seen in the asthma obstructed spirometry group, suggesting chronic inflammation and/or remodelling (0.17±0.11 kPa/(L·s^-1), p<0.05). Thus, small airway abnormalities of greater magnitude than observations in healthy obese people may be an early marker of asthma in obese subjects with self-reported disease despite normal airflow. Increased metabolic comorbidities in these subjects may have provided a milieu that impacted airway function.

Background: Alzheimer's disease (AD) is a progressive neurodegenerative disease that currently accounts for over 70% of cases of dementia in adults over 65 worldwide, and is the only cause of death among the top ten with no effective treatments. Clinically, AD is characterized by progressive deterioration in memory and other areas of cognitive function. Neuropathologically, the disease is characterized by extracellular aggregations of amyloid-B (AB) and intraneuronal neurofibrillary tangles (NFTS) composed of abnormally phosphorylated tau, causing progressive neuronal death. The aim of this study was to investigate whether the affibody ZSYM73-ABD (a portion of the active antibody molecule) can reverse AD pathology in an AD mouse model, without also causing significant neuroinflammation and/or microhemorrhage.
Method(s): APP/PS1 double transgenic mice were injected twice weekly with either ZSYM-ABD or a non-AB specific affibody, Ztaq2, as a control. Mice underwent behavioral testing and their brains were then sacrificed for immunohistochemistry.
Result(s): Semi-quantitative analysis of amyloid burden, performed using 6E10/4G8 antibodies, showed a statistically significant reduction in amyloid burden in the hippocampus, and a trend towards reduction in amyloid burden in the cortex. Inflammation was assessed using GFAP and Iba1(markers of gliosis) which showed a statistically significant reduction of GFAP in the cortex and in the hippocampus, and a slight reduction of microgliosis in ZSYM73-ABD affibody treated mice. Finally, mice treated with ZSYM73-ABD performed significantly better on a novel object recognition task than control mice, suggesting a correlation between the histological findings above and improvement in cognitive function.
Conclusion(s): In conclusion, this study demonstrates that passive immunization with an affibody molecule improves cognitive function and significantly decreases amyloid burden in the hippocampus of a transgenic mouse model of AD, without inducing inflammation. This has potential implications for treatment of AD in humans

BACKGROUND:Dent disease type 1 (DD1) is a rare X-linked disorder caused mainly by CLCN5 mutations. Patients may present with nephrotic-range proteinuria leading to erroneous diagnosis of focal segmental glomerulosclerosis (FSGS) and unnecessary immunosuppressive treatments. METHODS:M/TP, and A/TP from the CKiD cohort were compared with DD1 and DC. RESULTS:M/Cr of ≥ 120 mg/g (> 13.6 mg/mmol) creatinine were good screens for Dent disease. CONCLUSIONS:M/Cr ≥ 120 mg/g (> 13.6 mg/mmol) had the highest sensitivity and specificity when differentiating DD1 and studied CKiD populations.

The emergence of CRISPR-Cas9 gene-editing technologies and genome-wide CRISPR-Cas9 libraries enables efficient unbiased genetic screening that can accelerate the process of therapeutic discovery for genetic disorders. Here, we demonstrate the utility of a genome-wide CRISPR-Cas9 loss-of-function library to identify therapeutic targets for facioscapulohumeral muscular dystrophy (FSHD), a genetically complex type of muscular dystrophy for which there is currently no treatment. In FSHD, both genetic and epigenetic changes lead to misexpression of DUX4, the FSHD causal gene that encodes the highly cytotoxic DUX4 protein. We performed a genome-wide CRISPR-Cas9 screen to identify genes whose loss-of-function conferred survival when DUX4 was expressed in muscle cells. Genes emerging from our screen illuminated a pathogenic link to the cellular hypoxia response, which was revealed to be the main driver of DUX4-induced cell death. Application of hypoxia signaling inhibitors resulted in increased DUX4 protein turnover and subsequent reduction of the cellular hypoxia response and cell death. In addition, these compounds proved successful in reducing FSHD disease biomarkers in patient myogenic lines, as well as improving structural and functional properties in two zebrafish models of FSHD. Our genome-wide perturbation of pathways affecting DUX4 expression has provided insight into key drivers of DUX4-induced pathogenesis and has identified existing compounds with potential therapeutic benefit for FSHD. Our experimental approach presents an accelerated paradigm toward mechanistic understanding and therapeutic discovery of a complex genetic disease, which may be translatable to other diseases with well-established phenotypic selection assays.

The combination of electrocyclizations and cycloadditions accounts for the formation of a range of fascinating natural products. Cascades consisting of 8Ï€ electrocyclizations followed by a 6Ï€ electrocyclization and a cycloaddition are relatively common. We now report the synthesis of the tetramic acid PF-1018 through an 8Ï€ electrocyclization, the product of which is immediately intercepted by a Diels-Alder cycloaddition. The success of this pericyclic cascade was critically dependent on the substitution pattern of the starting polyene and could be rationalized through DFT calculations. The completion of the synthesis required the instalment of a trisubstituted double bond by radical deoxygenation. An unexpected side product formed through 4-exo-trig radical cyclization could be recycled through an unprecedented triflation/fragmentation.

The prefrontal cortex is vital for a range of cognitive processes, including working memory, attention, and decision-making. Notably, its absence impairs the performance of tasks requiring the maintenance of information through a delay period. In this paper, we formulate a rodent task-which requires maintenance of delay-period activity-as a Markov decision process and treat optimal task performance as an (active) inference problem. We simulate the behavior of a Bayes optimal mouse presented with 1 of 2 cues that instructs the selection of concurrent visual and auditory targets on a trial-by-trial basis. Formulating inference as message passing, we reproduce features of neuronal coupling within and between prefrontal regions engaged by this task. We focus on the micro-circuitry that underwrites delay-period activity and relate it to functional specialization within the prefrontal cortex in primates. Finally, we simulate the electrophysiological correlates of inference and demonstrate the consequences of lesions to each part of our in silico prefrontal cortex. In brief, this formulation suggests that recurrent excitatory connections-which support persistent neuronal activity-encode beliefs about transition probabilities over time. We argue that attentional modulation can be understood as the contextualization of sensory input by these persistent beliefs.

Supported lipid bilayer (SLB) membranes are key elements to mimic membrane interfaces on a planar surface. Here, we demonstrate that azobenzene photolipids (azo-PC) form fluid, homogeneous SLBs. Diffusion properties of azo-PC within SLBs were probed by fluorescence microscopy and fluorescence recovery after photobleaching. At ambient conditions, we find that the trans-to-cis isomerization causes an increase of the diffusion constant by a factor of two. Simultaneous excitation with two wavelengths and variable intensities furthermore allows to adjust the diffusion constant D continuously. X-ray reflectometry and small-angle scattering measurements reveal that membrane photoisomerization results in a bilayer thickness reduction of ∼0.4 nm (or 10%). While thermally induced back-switching is not observed, we find that the trans bilayer fluidity is increasing with higher temperatures. This change in diffusion constant is accompanied by a red-shift in the absorption spectra. Based on these results, we suggest that the reduced diffusivity of trans-azo-PC is controlled by intermolecular interactions that also give rise to H-aggregate formation in bilayer membranes.

Offline gains in motor performance after initial motor learning likely depend on sleep, but the molecular mechanisms by which this occurs are understudied. Regulation of mRNA translation via p70 S6 kinase 1 (S6K1) signaling represents one potential mechanism, as protein synthesis is thought to be increased during sleep compared to wake and is necessary for several forms of long-term memory. Using phosphorylation of ribosomal protein S6 (RpS6) as a readout of S6K1 activity, we demonstrate that a period of 10 hours of acute sleep disruption impairs both S6K1 signaling and offline gains in motor performance on the rotarod in adult wild type C57/Bl6 mice. Rotarod motor learning results in increased abundance of RpS6 in the striatum, and inhibition of S6K1 either indirectly with rapamycin or directly with PF-4708671 diminished the offline improvement in motor performance without affecting the initial acquisition of rotarod motor learning when sleep is normal. In sum, S6K1 activity is required for sleep-dependent offline gains in motor performance and is inhibited following acute sleep disruption, while motor learning increases the abundance of striatal RpS6. Thus, S6K1 signaling represents a plausible mechanism mediating the beneficial effects of sleep on motor performance.

Responses of sensory neurons are often modeled using a weighted combination of rectified linear subunits. Since these subunits often cannot be measured directly, a flexible method is needed to infer their properties from the responses of downstream neurons. We present a method for maximum likelihood estimation of subunits by soft-clustering spike-triggered stimuli, and demonstrate its effectiveness in visual neurons. For parasol retinal ganglion cells in macaque retina, estimated subunits partitioned the receptive field into compact regions, likely representing aggregated bipolar cell inputs. Joint clustering revealed shared subunits between neighboring cells, producing a parsimonious population model. Closed-loop validation, using stimuli lying in the null space of the linear receptive field, revealed stronger nonlinearities in OFF cells than ON cells. Responses to natural images, jittered to emulate fixational eye movements, were accurately predicted by the subunit model. Finally, the generality of the approach was demonstrated in macaque V1 neurons.

The ε4 allele of apolipoprotein E (APOE) is the dominant genetic risk factor for late-onset Alzheimer's disease (AD). However, the reason for the association between APOE4 and AD remains unclear. While much of the research has focused on the ability of the apoE4 protein to increase the aggregation and decrease the clearance of Aβ, there is also an abundance of data showing that APOE4 negatively impacts many additional processes in the brain, including bioenergetics. In order to gain a more comprehensive understanding of APOE4's role in AD pathogenesis, we performed a transcriptomics analysis of APOE4 vs. APOE3 expression in the entorhinal cortex (EC) and primary visual cortex (PVC) of aged APOE mice. This study revealed EC-specific upregulation of genes related to oxidative phosphorylation (OxPhos). Follow-up analysis utilizing the Seahorse platform showed decreased mitochondrial respiration with age in the hippocampus and cortex of APOE4 vs. APOE3 mice, but not in the EC of these mice. Additional studies, as well as the original transcriptomics data, suggest that multiple bioenergetic pathways are differentially regulated by APOE4 expression in the EC of aged APOE mice in order to increase the mitochondrial coupling efficiency in this region. Given the importance of the EC as one of the first regions to be affected by AD pathology in humans, the observation that the EC is susceptible to differential bioenergetic regulation in response to a metabolic stressor such as APOE4 may point to a causative factor in the pathogenesis of AD.