Palmitoyl transferase ZDHHC20 promotes pancreatic cancer metastasis.

Metastasis is one of the defining features of pancreatic ductal adenocarcinoma (PDAC) that contributes to poor prognosis. In this study, the palmitoyl transferase ZDHHC20 was identified in an in vivo short hairpin RNA (shRNA) screen as critical for metastatic outgrowth, with no effect on proliferation and migration in vitro or primary PDAC growth in mice. This phenotype is abrogated in immunocompromised animals and animals with depleted natural killer (NK) cells, indicating that ZDHHC20 affects the interaction of tumor cells and the innate immune system. Using a chemical genetics platform for ZDHHC20-specific substrate profiling, a number of substrates of this enzyme were identified. These results describe a role for palmitoylation in enabling distant metastasis that could not have been detected using in vitro screening approaches and identify potential effectors through which ZDHHC20 promotes metastasis of PDAC.

A palmitoyl transferase chemical-genetic system to map ZDHHC-specific S-acylation.

The 23 human zinc finger Asp-His-His-Cys motif-containing (ZDHHC) S-acyltransferases catalyze long-chain S-acylation at cysteine residues across an extensive network of hundreds of proteins important for normal physiology or dysregulated in disease. Here we present a technology to directly map the protein substrates of a specific ZDHHC at the whole-proteome level, in intact cells. Structure-guided engineering of paired ZDHHC 'hole' mutants and 'bumped' chemically tagged fatty acid probes enabled probe transfer to specific protein substrates with excellent selectivity over wild-type ZDHHCs. Chemical-genetic systems were exemplified for five human ZDHHCs (3, 7, 11, 15 and 20) and applied to generate de novo ZDHHC substrate profiles, identifying >300 substrates and S-acylation sites for new functionally diverse proteins across multiple cell lines. We expect that this platform will elucidate S-acylation biology for a wide range of models and organisms.

Single-copy Snail upregulation causes partial epithelial-mesenchymal transition in colon cancer cells.

BACKGROUND: Epithelial-mesenchymal transition (EMT) is an embryonic programme implicated in cancer stem cells, metastasis and therapeutic resistance. Its role in cancer progression remains controversial because the transition can be partial or complete in different models and contexts. METHODS: Using human colon cancer DLD-1 cells, we engineered a cell line with a single-copy of Snail that was doxycycline-inducible and compared it to existing EMT models in DLD-1. The effect of Snail upregulation was characterised functionally, morphologically, and by transcriptional profiling and protein expression. RESULTS: Induction with doxycycline increased Snail expression to a level similar to that observed in cancer cell lines spontaneously expressing Snail and results in partial EMT. In comparison, higher levels of overexpression arising from introduction of episomal-Snail, results in complete EMT. DLD-1 cells with partial EMT show chemoresistance in vitro, increased tumour growth in vivo and decreased apoptosis. CONCLUSIONS: These findings highlight that the amount of bioavailable Snail can dictate phenotypic outcome and that partial EMT may be a preferred outcome of models operating within a natural range of Snail overexpression.

Noninvasively recorded high-gamma signals improve synchrony of force feedback in a novel neurorehabilitation brain-machine interface for brain injury.

Objective.Brain injury is the leading cause of long-term disability worldwide, often resulting in impaired hand function. Brain-machine interfaces (BMIs) offer a potential way to improve hand function. BMIs often target replacing lost function, but may also be employed in neurorehabilitation (nrBMI) by facilitating neural plasticity and functional recovery. Here, we report a novel nrBMI capable of acquiring high-γ(70-115 Hz) information through a unique post-traumatic brain injury (TBI) hemicraniectomy window model, and delivering sensory feedback that is synchronized with, and proportional to, intended grasp force.Approach. We developed the nrBMI to use electroencephalogram recorded over a hemicraniectomy (hEEG) in individuals with TBI. The nrBMI empowered users to exert continuous, proportional control of applied force, and provided continuous force feedback. We report the results of an initial testing group of three human participants with TBI, along with a control group of three skull- and motor-intact volunteers.Main results. All participants controlled the nrBMI successfully, with high initial success rates (2 of 6 participants) or performance that improved over time (4 of 6 participants). We observed high-γmodulation with force intent in hEEG but not skull-intact EEG. Most significantly, we found that high-γcontrol significantly improved the timing synchronization between neural modulation onset and nrBMI output/haptic feedback (compared to low-frequency nrBMI control).Significance. These proof-of-concept results show that high-γnrBMIs can be used by individuals with impaired ability to control force (without immediately resorting to invasive signals like electrocorticography). Of note, the nrBMI includes a parameter to change the fraction of control shared between decoded intent and volitional force, to adjust for recovery progress. The improved synchrony between neural modulations and force control for high-γsignals is potentially important for maximizing the ability of nrBMIs to induce plasticity in neural circuits. Inducing plasticity is critical to functional recovery after brain injury.

Memory Reactivation during Sleep Improves Execution of a Challenging Motor Skill.

Memory reactivation during sleep reinforces various types of learning. Basic motor skills likely benefit from sleep. There is insufficient evidence, however, on whether memory reactivation during sleep contributes to learning how to execute a novel action. Here, we investigated motor learning in a myoelectric feedback task. Human male and female participants learned to control myoelectric activity in specific arm muscles to move a computer cursor to each of 16 locations. Each location was associated with a unique sound. Half of the sounds were played during slow-wave sleep to reactivate corresponding memories of muscle control. After sleep, movements cued during sleep were performed more quickly and efficiently than uncued movements. These results demonstrated that memory reactivation during sleep contributes to learning of action execution. We conclude that sleep supports learning novel actions, which also maps onto the learning required in certain neurorehabilitation procedures.SIGNIFICANCE STATEMENT Prior literature on motor learning has produced much evidence supporting a role for sleep but scant evidence on the execution component. This aspect of learning is critical for many complex skills that people value in their lives. Our results not only implicate sleep in skill learning but also pinpoint a benefit for motor execution using a method for modifying memory storage during sleep. We used targeted memory reactivation (TMR), whereby a stimulus that has been associated with learning is presented again during sleep to bring on a recapitulation of waking brain activity. Our demonstration that memory reactivation contributed to skilled performance may be relevant for neurorehabilitation as well as fields concerned with motor learning, such as kinesiology and physiology.

Wearable myoelectric interface enables high-dose, home-based training in severely impaired chronic stroke survivors.

BACKGROUND: High-intensity occupational therapy can improve arm function after stroke, but many people lack access to such therapy. Home-based therapies could address this need, but they don't typically address abnormal muscle co-activation, an important aspect of arm impairment. An earlier study using lab-based, myoelectric computer interface game training enabled chronic stroke survivors to reduce abnormal co-activation and improve arm function. Here, we assess feasibility of doing this training at home using a novel, wearable, myoelectric interface for neurorehabilitation training (MINT) paradigm. OBJECTIVE: Assess tolerability and feasibility of home-based, high-dose MINT therapy in severely impaired chronic stroke survivors. METHODS: Twenty-three participants were instructed to train with the MINT and game for 90 min/day, 36 days over 6 weeks. We assessed feasibility using amount of time trained and game performance. We assessed tolerability (enjoyment and effort) using a customized version of the Intrinsic Motivation Inventory at the conclusion of training. RESULTS: Participants displayed high adherence to near-daily therapy at home (mean of 82 min/day of training; 96% trained at least 60 min/day) and enjoyed the therapy. Training performance improved and co-activation decreased with training. Although a substantial number of participants stopped training, most dropouts were due to reasons unrelated to the training paradigm itself. INTERPRETATION: Home-based therapy with MINT is feasible and tolerable in severely impaired stroke survivors. This affordable, enjoyable, and mobile health paradigm has potential to improve recovery from stroke in a variety of settings. Clinicaltrials.gov: NCT03401762.

Hemicraniectomy in Traumatic Brain Injury: A Noninvasive Platform to Investigate High Gamma Activity for Brain Machine Interfaces.

Brain-machine interfaces (BMIs) translate brain signals into control signals for an external device, such as a computer cursor or robotic limb. These signals can be obtained either noninvasively or invasively. Invasive recordings, using electrocorticography (ECoG) or intracortical microelectrodes, provide higher bandwidth and more informative signals. Rehabilitative BMIs, which aim to drive plasticity in the brain to enhance recovery after brain injury, have almost exclusively used non-invasive recordings, such electroencephalography (EEG) or magnetoencephalography (MEG), which have limited bandwidth and information content. Invasive recordings provide more information and spatiotemporal resolution, but do incur risk, and thus are not usually investigated in people with stroke or traumatic brain injury (TBI). Here, in this paper, we describe a new BMI paradigm to investigate the use of higher frequency signals in brain-injured subjects without incurring significant risk. We recorded EEG in TBI subjects who required hemicraniectomies (removal of a part of the skull). EEG over the hemicraniectomy (hEEG) contained substantial information in the high gamma frequency range (65-115 Hz). Using this information, we decoded continuous finger flexion force with moderate to high accuracy (variance accounted for 0.06 to 0.52), which at best approaches that using epidural signals. These results indicate that people with hemicraniectomies can provide a useful resource for developing BMI therapies for the treatment of brain injury.

Myoelectric Computer Interface Training for Reducing Co-Activation and Enhancing Arm Movement in Chronic Stroke Survivors: A Randomized Trial.

BACKGROUND: Abnormal muscle co-activation contributes to impairment after stroke. We developed a myoelectric computer interface (MyoCI) training paradigm to reduce abnormal co-activation. MyoCI provides intuitive feedback about muscle activation patterns, enabling decoupling of these muscles. OBJECTIVE: To investigate tolerability and effects of MyoCI training of 3 muscle pairs on arm motor recovery after stroke, including effects of training dose and isometric versus movement-based training. METHODS: We randomized chronic stroke survivors with moderate-to-severe arm impairment to 3 groups. Two groups tested different doses of isometric MyoCI (60 vs 90 minutes), and one group tested MyoCI without arm restraint (90 minutes), over 6 weeks. Primary outcome was arm impairment (Fugl-Meyer Assessment). Secondary outcomes included function, spasticity, and elbow range-of-motion at weeks 6 and 10. RESULTS: Over all 32 subjects, MyoCI training of 3 muscle pairs significantly reduced impairment (Fugl-Meyer Assessment) by 3.3 ± 0.6 and 3.1 ± 0.7 ( P < 10-4) at weeks 6 and 10, respectively. Each group improved significantly from baseline; no significant differences were seen between groups. Participants' lab-based and home-based function also improved at weeks 6 and 10 ( P ≤ .01). Spasticity also decreased over all subjects, and elbow range-of-motion improved. Both moderately and severely impaired patients showed significant improvement. No participants had training-related adverse events. MyoCI reduced abnormal co-activation, which appeared to transfer to reaching in the movement group. CONCLUSIONS: MyoCI is a well-tolerated, novel rehabilitation tool that enables stroke survivors to reduce abnormal co-activation. It may reduce impairment and spasticity and improve arm function, even in severely impaired patients.

The effect of myoelectric computer interface training on arm kinematics and function after stroke.

Abnormal co-activation patterns of arm muscles is a substantial cause of impaired arm function after stroke. We designed a myoelectric computer interface (MCI) training paradigm to help stroke survivors reduce this abnormal coactivation. Here, we evaluated the effects of MCI training on function and arm kinematics in 32 chronic stroke survivors. We compared the effects of training duration and isometric vs. movement-based training conditions in 3 different groups. All groups reduced abnormal co-activation in targeted muscles, and showed reduced arm impairment after 6 weeks of training. They also showed improvements in arm kinematics as well as functional scores. Moreover, the gains persisted, though most were reduced, at one month after training stopped. These results suggest that MCI training holds promise to improve arm function after stroke.

Phospho-regulation of ATOH1 Is Required for Plasticity of Secretory Progenitors and Tissue Regeneration.

The intestinal epithelium is largely maintained by self-renewing stem cells but with apparently committed progenitors also contributing, particularly following tissue damage. However, the mechanism of, and requirement for, progenitor plasticity in mediating pathological response remain unknown. Here we show that phosphorylation of the transcription factor Atoh1 is required for both the contribution of secretory progenitors to the stem cell pool and for a robust regenerative response. As confirmed by lineage tracing, Atoh1+ cells (Atoh1(WT)CreERT2 mice) give rise to multilineage intestinal clones both in the steady state and after tissue damage. In a phosphomutant Atoh1(9S/T-A)CreERT2 line, preventing phosphorylation of ATOH1 protein acts to promote secretory differentiation and inhibit the contribution of progenitors to self-renewal. Following chemical colitis, Atoh1+ cells of Atoh1(9S/T-A)CreERT2 mice have reduced clonogenicity that affects overall regeneration. Progenitor plasticity maintains robust self-renewal in the intestinal epithelium, and the balance between stem and progenitor fate is directly coordinated by ATOH1 multisite phosphorylation.

Phytochemical profile of Rosmarinus officinalis and Salvia officinalis extracts and correlation to their antioxidant and anti-proliferative activity.

The goal of this study was to monitor the anti-proliferative activity of Rosmarinus officinalis and Salvia officinalis extracts against cancer cells and to correlate this activity with their phytochemical profiles using liquid chromatography/diode array detection/electrospray ion trap tandem mass spectrometry (LC/DAD/ESI-MS(n)). For the quantitative estimation of triterpenic acids in the crude extracts an NMR based methodology was used and compared with the HPLC measurements, both applied for the first time, for the case of betulinic acid. Both extracts exerted cytotoxic activity through dose-dependent impairment of viability and mitochondrial activity of rat insulinoma m5F (RINm5F) cells. Decrease of RINm5F viability was mediated by nitric oxide (NO)-induced apoptosis. Importantly, these extracts potentiated NO and TNF-α release from macrophages therefore enhancing their cytocidal action. The rosemary extract developed more pronounced antioxidant, cytotoxic and immunomodifying activities, probably due to the presence of betulinic acid and a higher concentration of carnosic acid in its phytochemical profile.