Identification of MLKL membrane translocation as a checkpoint in necroptotic cell death using Monobodies.

The necroptosis cell death pathway has been implicated in host defense and in the pathology of inflammatory diseases. While phosphorylation of the necroptotic effector pseudokinase Mixed Lineage Kinase Domain-Like (MLKL) by the upstream protein kinase RIPK3 is a hallmark of pathway activation, the precise checkpoints in necroptosis signaling are still unclear. Here we have developed monobodies, synthetic binding proteins, that bind the N-terminal four-helix bundle (4HB) "killer" domain and neighboring first brace helix of human MLKL with nanomolar affinity. When expressed as genetically encoded reagents in cells, these monobodies potently block necroptotic cell death. However, they did not prevent MLKL recruitment to the "necrosome" and phosphorylation by RIPK3, nor the assembly of MLKL into oligomers, but did block MLKL translocation to membranes where activated MLKL normally disrupts membranes to kill cells. An X-ray crystal structure revealed a monobody-binding site centered on the α4 helix of the MLKL 4HB domain, which mutational analyses showed was crucial for reconstitution of necroptosis signaling. These data implicate the α4 helix of its 4HB domain as a crucial site for recruitment of adaptor proteins that mediate membrane translocation, distinct from known phospholipid binding sites.

Computer-assisted EEG diagnostic review for idiopathic generalized epilepsy.

Epilepsy diagnosis can be costly, time-consuming, and not uncommonly inaccurate. The reference standard diagnostic monitoring is continuous video-electroencephalography (EEG) monitoring, ideally capturing all events or concordant interictal discharges. Automating EEG data review would save time and resources, thus enabling more people to receive reference standard monitoring and also potentially heralding a more quantitative approach to therapeutic outcomes. There is substantial research into the automated detection of seizures and epileptic activity from EEG. However, automated detection software is not widely used in the clinic, and despite numerous published algorithms, few methods have regulatory approval for detecting epileptic activity from EEG. This study reports on a deep learning algorithm for computer-assisted EEG review. Deep convolutional neural networks were trained to detect epileptic discharges using a preexisting dataset of over 6000 labelled events in a cohort of 103 patients with idiopathic generalized epilepsy (IGE). Patients underwent 24-hour ambulatory outpatient EEG, and all data were curated and confirmed independently by two epilepsy specialists (Seneviratne et al., 2016). The resulting automated detection algorithm was then used to review diagnostic scalp EEG for seven patients (four with IGE and three with events mimicking seizures) to validate performance in a clinical setting. The automated detection algorithm showed state-of-the-art performance for detecting epileptic activity from clinical EEG, with mean sensitivity of >95% and corresponding mean false positive rate of 1 detection per minute. Importantly, diagnostic case studies showed that the automated detection algorithm reduced human review time by 80%-99%, without compromising event detection or diagnostic accuracy. The presented results demonstrate that computer-assisted review can increase the speed and accuracy of EEG assessment and has the potential to greatly improve therapeutic outcomes. This article is part of the Special Issue "NEWroscience 2018".

Use of Electrical Stimulation for People With Spinal Cord Injury: A Survey of Occupational Therapy Practitioners.

IMPORTANCE: When working with clients who have experienced spinal cord injury (SCI), occupational therapy practitioners can face challenges in achieving desired results during functional activity when using electrical stimulation (ES) interventions. In an effort to understand current practice, a survey study was conducted. OBJECTIVE: For people with SCI, ES elicits positive physiological effects; however, no implementation guidelines exist for upper extremity application of ES for this population. Therefore, we surveyed occupational therapy practitioners about their use of ES with clients who have cervical-level SCI. DESIGN: A 33-item, 20-min online survey was used. PARTICIPANTS AND SETTING: We queried 57 occupational therapy practitioners with active caseloads in regional rehabilitation centers specializing in SCI, both outpatient and inpatient. RESULTS: For clients with SCI, occupational therapy practitioners used ES most often for grasp-and-release, reaching, and grip or pinch activities using a broad range of parameter settings. Among respondents, 43% did not use a specific treatment protocol; 27% used research evidence to guide selection of parameters. CONCLUSIONS AND RELEVANCE: Findings suggest that ES treatment parameters are not uniformly applied, introducing potential unknown effects on client outcomes and undermining treatment fidelity. WHAT THIS ARTICLE ADDS: Our survey of occupational therapy practitioners regarding their practice and use of ES interventions with this population revealed variation in application of ES treatment parameters. Understanding different treatment approaches and justification used when applying ES to clients with SCI is an important first step in unifying and promoting best practice and maximizing patient outcomes.

Alterations in Aerobic Exercise Performance and Gait Economy Following High-Intensity Dynamic Stepping Training in Persons With Subacute Stroke.

BACKGROUND AND PURPOSE: Impairments in metabolic capacity and economy (O2cost) are hallmark characteristics of locomotor dysfunction following stroke. High-intensity (aerobic) training has been shown to improve peak oxygen consumption in this population, with fewer reports of changes in O2cost. However, particularly in persons with subacute stroke, inconsistent gains in walking function are observed with minimal associations with gains in metabolic parameters. The purpose of this study was to evaluate changes in aerobic exercise performance in participants with subacute stroke following high-intensity variable stepping training as compared with conventional therapy. METHODS: A secondary analysis was performed on data from a randomized controlled trial comparing high-intensity training with conventional interventions, and from the pilot study that formed the basis for the randomized controlled trial. Participants 1 to 6 months poststroke received 40 or fewer sessions of high-intensity variable stepping training (n = 21) or conventional interventions (n = 12). Assessments were performed at baseline (BSL), posttraining, and 2- to 3-month follow-up and included changes in submaximal (Equation is included in full-text article.)O2 ((Equation is included in full-text article.)O2submax) and O2cost at fastest possible treadmill speeds and peak speeds at BSL testing. RESULTS: Significant improvements were observed in (Equation is included in full-text article.)O2submax with less consistent improvements in O2cost, although individual responses varied substantially. Combined changes in both (Equation is included in full-text article.)O2submax and (Equation is included in full-text article.)O2 at matched peak BSL speeds revealed stronger correlations to improvements in walking function as compared with either measure alone. DISCUSSION AND CONCLUSIONS: High-intensity stepping training may elicit significant improvements in (Equation is included in full-text article.)O2submax, whereas changes in both peak capacity and economy better reflect gains in walking function. Providing high-intensity training to improve locomotor and aerobic exercise performance may increase the efficiency of rehabilitation sessions.Video Abstract available for more insights from the authors (see Supplemental Digital Content, http://links.lww.com/JNPT/A142).

Deep learning supported machine vision system to precisely automate the wild blueberry harvester header.

An operator of a wild blueberry harvester faces the fatigue of manually adjusting the height of the harvester's head, considering spatial variations in plant height, fruit zone, and field topography affecting fruit yield. For stress-free harvesting of wild blueberries, a deep learning-supported machine vision control system has been developed to detect the fruit height and precisely auto-adjust the header picking teeth rake position. The OpenCV AI Kit (OAK-D) was used with YOLOv4-tiny deep learning model with code developed in Python to solve the challenge of matching fruit heights with the harvester's head position. The system accuracy was statistically evaluated with R2 (coefficient of determination) and σ (standard deviation) measured on the difference in distances between the berries picking teeth and average fruit heights, which were 72, 43% and 2.1, 2.3 cm for the auto and manual head adjustment systems, respectively. This innovative system performed well in weed-free areas but requires further work to operate in weedy sections of the fields. Benefits of using this system include automated control of the harvester's head to match the header picking rake height to the level of the fruit height while reducing the operator's stress by creating safer working environments.

Water-soluble adhesive for stable long-term ambulatory EEG recordings.

OBJECTIVE: Conventional methods used to adhere EEG electrodes are often uncomfortable. Here, we present a polymer-based water-soluble EEG adhesive that can be maintained for up to 6 days. The primary outcome measure of this study is the median electrode impedance at day 6. METHODS: Impedance measurements for 841 EEG recordings using a 21 channel 10-20 configuration were remotely logged daily for 6 days after connection. A novel electrode adhesive was used to attach EEG electrodes. Patients were instructed to maintain their electrodes on day 4. RESULTS: Median electrode impedances were significantly below 10kOhms for each day of recording, with a median value on day 6 of 4.18kOhms. Impedance values were significantly lower on day 5 than on day 4, demonstrating that the maintenance process can reduce impedance. Except for day 4-5, the median impedance increased each day. No significant difference was found on the first or final day between clinics or residences from areas of different geographic remoteness. CONCLUSIONS: EEG is able to be recorded in patients homes for 6 days with acceptable impedance and no significant effect of regionality or patients age. SIGNIFICANCE: To the best of our knowledge, this is the first report in the literature of impedance data from long-term ambulatory EEG studies.

Crystal structure of the hinge domain of Smchd1 reveals its dimerization mode and nucleic acid-binding residues.

Structural maintenance of chromosomes flexible hinge domain containing 1 (SMCHD1) is an epigenetic regulator in which polymorphisms cause the human developmental disorder, Bosma arhinia micropthalmia syndrome, and the degenerative disease, facioscapulohumeral muscular dystrophy. SMCHD1 is considered a noncanonical SMC family member because its hinge domain is C-terminal, because it homodimerizes rather than heterodimerizes, and because SMCHD1 contains a GHKL-type, rather than an ABC-type ATPase domain at its N terminus. The hinge domain has been previously implicated in chromatin association; however, the underlying mechanism involved and the basis for SMCHD1 homodimerization are unclear. Here, we used x-ray crystallography to solve the three-dimensional structure of the Smchd1 hinge domain. Together with structure-guided mutagenesis, we defined structural features of the hinge domain that participated in homodimerization and nucleic acid binding, and we identified a functional hotspot required for chromatin localization in cells. This structure provides a template for interpreting the mechanism by which patient polymorphisms within the SMCHD1 hinge domain could compromise function and lead to facioscapulohumeral muscular dystrophy.

Cardiopulmonary Responses During Clinical and Laboratory Gait Assessments in People With Chronic Stroke.

Background: The 6-Minute Walk Test (6MWT) is a common clinical assessment used to evaluate locomotor function in patients after stroke. Previous work suggests the 6MWT can estimate peak metabolic capacity (VO2peak) without cardiorespiratory assessments during graded exercise tests (GXTs), which may assist with exercise prescription. However, selected research also indicated increased heart rates (HRs) during 6MWTs beyond levels considered safe without GXTs. Objective: The goal of this study was to examine cardiorespiratory responses during 6MWTs and GXTs in individuals with chronic stroke and their associations with demographic or clinical characteristics. Design: The study used a cross-sectional observational design. Methods: Cardiorespiratory responses were assessed during 6MWTs at self-selected velocity (SSV) and fastest velocity (FV), and during GXTs. Secondary assessments included the lower extremity Fugl-Meyer Assessment, Functional Gait Assessment, gait speeds, and daily stepping activity. Correlation and regression analyses were used to evaluate associations between locomotor performance, cardiorespiratory responses, and clinical and demographic characteristics. Results: Average HRs during 6MWT-FV were 72% to 76% of the age-predicted maximum (HRmax), with 20% of participants exceeding 85% predicted HRmax. When normalized to HRs during GXTs, HRs during 6MWT-FV were 86% to 88% of observed HRmax. Primary predictors of increased HRs during 6MWTs were resting HR, body mass index, and daily stepping. Distance during 6MWT-FV was a significant predictor of VO2peak in combination with other variables. Electrocardiographic abnormalities were observed in >80% of participants at rest and 31% demonstrated distinct abnormalities during GXTs, which were not related to 6MWT or GXT performance. Limitations: In addition to sample size, a primary limitation involved the ability to accurately predict or measure HRmax in patients with motor dysfunction after stroke. Conclusions: Cardiac responses were higher than anticipated during 6MWTs and often exceeded recommended HR thresholds. Clinicians should closely monitor cardiorespiratory responses during 6MWTs.

Altered Sagittal- and Frontal-Plane Kinematics Following High-Intensity Stepping Training Versus Conventional Interventions in Subacute Stroke.

BACKGROUND: Common locomotor deficits observed in people poststroke include decreased speeds and abnormal kinematics, characterized by altered symmetry, reduced sagittal-plane joint excursions, and use of compensatory frontal-plane behaviors during the swing phase of gait. Conventional interventions utilized to mitigate these deficits often incorporate low-intensity, impairment-based or functional exercises focused on normalizing kinematics, although the efficacy of these strategies is unclear. Conversely, higher-intensity training protocols that provide only stepping practice and do not focus on kinematics have demonstrated gains in walking function, although minimal attention toward gait quality may be concerning and has not been assessed. OBJECTIVE: The present study evaluated changes in spatiotemporal and joint kinematics following experimental, high-intensity stepping training compared with conventional interventions. DESIGN: Kinematic data were combined from a randomized controlled trial comparing experimental and conventional training and from a pilot experimental training study. METHODS: Individuals with gait deficits 1 to 6 months poststroke received up to 40 sessions of either high-intensity stepping training in variable contexts or conventional lower-intensity interventions. Analyses focused on kinematic changes during graded treadmill testing before and following training. RESULTS: Significant improvements in speed, symmetry, and selected sagittal-plane kinematics favored experimental training over conventional training, although increases in compensatory strategies also were observed. Changes in many kinematic patterns were correlated with speed changes, and increased compensatory behaviors were associated with both stride length gains and baseline impairments. LIMITATIONS: Limitations include a small sample size and use of multiple statistical comparisons. CONCLUSIONS: Improved speeds and selected kinematics were observed following high-intensity training, although such training also resulted in increased use of compensatory strategies. Future studies should explore the consequences of utilizing these compensatory strategies despite the observed functional gains.

Effects of Training Intensity on Locomotor Performance in Individuals With Chronic Spinal Cord Injury: A Randomized Crossover Study.

BACKGROUND: Many physical interventions can improve locomotor function in individuals with motor incomplete spinal cord injury (iSCI), although the training parameters that maximize recovery are not clear. Previous studies in individuals with other neurologic injuries suggest the intensity of locomotor training (LT) may positively influence walking outcomes. However, the effects of intensity during training of individuals with iSCI have not been tested. OBJECTIVE: The purpose of this pilot, blinded-assessor randomized trial was to evaluate the effects of LT intensity on walking outcomes in individuals with iSCI. METHODS: Using a crossover design, ambulatory participants with iSCI >1 year duration performed either high- or low-intensity LT for ≤20 sessions over 4 to 6 weeks. Four weeks following completion, the training interventions were alternated. Targeted intensities focused on achieving specific ranges of heart rate (HR) or ratings of perceived exertion (RPE), with intensity manipulated by increasing speeds or applying loads. RESULTS: Significantly greater increases in peak treadmill speeds (0.18 vs 0.02 m/s) and secondary measures of metabolic function and overground speed were observed following high- versus low-intensity training, with no effects of intervention order. Moderate to high correlations were observed between differences in walking speed or distances and differences in HRs or RPEs during high- versus low-intensity training. CONCLUSION: This pilot study provides the first evidence that the intensity of stepping practice may be an important determinant of LT outcomes in individuals with iSCI. Whether such training is feasible in larger patient populations and contributes to improved locomotor outcomes deserves further consideration.

Variable Intensive Early Walking Poststroke (VIEWS): A Randomized Controlled Trial.

BACKGROUND: Converging evidence suggests that the amount of stepping practice is an important training parameter that influences locomotor recovery poststroke. More recent data suggest that stepping intensity and variability are also important, although such strategies are often discouraged early poststroke. OBJECTIVE: The present study examined the efficacy of high-intensity, variable stepping training on walking and nonwalking outcomes in individuals 1 to 6 months poststroke as compared with conventional interventions. Methods Individuals with unilateral stroke (mean duration = 101 days) were randomized to receive ≤40, 1-hour experimental or control training sessions over 10 weeks. Experimental interventions consisted only of stepping practice at high cardiovascular intensity (70%-80% heart rate reserve) in variable contexts (tasks or environments). Control interventions were determined by clinical physical therapists and supplemented using standardized conventional strategies. Blinded assessments were obtained at baseline, midtraining, and posttraining with a 2-month follow-up. Results A total of 32 individuals (15 experimental) received different training paradigms that varied in the amount, intensity, and types of tasks performed. Primary outcomes of walking speed (experimental, 0.27 ± 0.22 m/s vs control, 0.09 ± 0.09 m/s) and distances (119 ± 113 m vs 30 ± 32 m) were different between groups, with stepping amount and intensity related to these differences. Gains in temporal gait symmetry and self-reported participation scores were greater following experimental training, without differences in balance or sit-to-stand performance. Conclusion Variable intensive stepping training resulted in greater improvements in walking ability than conventional interventions early poststroke. Future studies should evaluate the relative contributions of these training parameters.

Feasibility of Focused Stepping Practice During Inpatient Rehabilitation Poststroke and Potential Contributions to Mobility Outcomes.

BACKGROUND: Optimal physical therapy strategies to maximize locomotor function in patients early poststroke are not well established. Emerging data indicate that substantial amounts of task-specific stepping practice may improve locomotor function, although stepping practice provided during inpatient rehabilitation is limited (<300 steps/session). OBJECTIVE: The purpose of this investigation was to determine the feasibility of providing focused stepping training to patients early poststroke and its potential association with walking and other mobility outcomes. METHODS: Daily stepping was recorded on 201 patients <6 months poststroke (80% < 1 month) during inpatient rehabilitation following implementation of a focused training program to maximize stepping practice during clinical physical therapy sessions. Primary outcomes included distance and physical assistance required during a 6-minute walk test (6MWT) and balance using the Berg Balance Scale (BBS). Retrospective data analysis included multiple regression techniques to evaluate the contributions of demographics, training activities, and baseline motor function to primary outcomes at discharge. RESULTS: Median stepping activity recorded from patients was 1516 steps/d, which is 5 to 6 times greater than that typically observed. The number of steps per day was positively correlated with both discharge 6MWT and BBS and improvements from baseline (changes; r = 0.40-0.87), independently contributing 10% to 31% of the total variance. Stepping activity also predicted level of assistance at discharge and discharge location (home vs other facility). CONCLUSION: Providing focused, repeated stepping training was feasible early poststroke during inpatient rehabilitation and was related to mobility outcomes. Further research is required to evaluate the effectiveness of these training strategies on short- or long-term mobility outcomes as compared with conventional interventions.

Microparticle dispensers for the controlled release of insect pheromones.

The potential utility of micrometer-sized particles as controlled-release devices for the volatilization of insect pheromones for mating disruption applications is evaluated in this study for two pheromone/model compound systems (codlemone/1-dodecanol and disparlure/1,2-epoxyoctadecane). To expedite the measurement of release rates from these particle devices, two techniques based on thermogravimetric analysis (TGA) have been exploited: isothermal TGA (I-TGA) at elevated temperatures (40-80 degrees C) with N(2) convection and volatilization temperature (VT) by dynamic TGA. A correlation between these two methods has been established. Samples that exhibit a higher VT provide a lower release rate from a particle substrate. Using these techniques, it has been demonstrated that chemical interactions between adsorbed liquids and particle surfaces may play a small role in defining release characteristics under conditions of low surface area, whereas parameters associated with total surface area and micropore structure appear to be much more significant in retarding evaporation for uncoated particles containing an adsorbed liquid. Additional regulation of release rates was achieved by coating the particle systems with water-soluble or water-dispersible polymers. By careful selection of particle porosity and coating composition, it is envisioned that the evaporation rate of pheromones can be tailored to specific insect control applications.

Effects of dynamic stepping training on nonlocomotor tasks in individuals poststroke.

BACKGROUND: During the physical rehabilitation of individuals poststroke, therapists are challenged to provide sufficient amounts of task-specific practice in order to maximize outcomes of multiple functional skills within limited visits. Basic and applied studies have suggested that training of one motor task may affect performance of biomechanically separate tasks that utilize overlapping neural circuits. However, few studies have explicitly investigated the impact of training one functional task on separate, nonpracticed tasks. OBJECTIVE: The purpose of this preliminary study was to investigate the potential gains in specific nonlocomotor assessments in individuals poststroke following only stepping training of variable, challenging tasks at high aerobic intensities. METHODS: Individuals with locomotor deficits following subacute and chronic stroke (n=22) completed a locomotor training paradigm using a repeated-measures design. Practice of multiple stepping tasks was provided in variable environments or contexts at high aerobic intensities for ≥40 sessions over 10 weeks. The primary outcome was timed Five-Times Sit-to-Stand Test (5XSTS) performance, with secondary measures of sit-to-stand kinematics and kinetics, clinical assessment of balance, and isometric lower limb strength. RESULTS: Participants improved their timed 5XSTS performance following stepping training, with changes in selected biomechanical measures. Statistical and clinically meaningful improvements in balance were observed, with more modest changes in paretic leg strength. CONCLUSIONS: The present data suggest that significant gains in selected nonlocomotor tasks can be achieved with high-intensity, variable stepping training. Improvements in nonpracticed tasks may minimize the need to practice multiple tasks within and across treatment sessions.