Transcriptional changes of genes encoding sarcoplasmic reticulum calcium binding and up-taking proteins in normal and Duchenne muscular dystrophy dogs.

BACKGROUND: Cytosolic calcium overload contributes to muscle degradation in Duchenne muscular dystrophy (DMD). The sarcoplasmic reticulum (SR) is the primary calcium storage organelle in muscle. The sarco-endoplasmic reticulum ATPase (SERCA) pumps cytosolic calcium to the SR during muscle relaxation. Calcium is kept in the SR by calcium-binding proteins. METHODS: Given the importance of the canine DMD model in translational studies, we examined transcriptional changes of SERCA (SERCA1 and SERCA2a), SERCA regulators (phospholamban, sarcolipin, myoregulin, and dwarf open reading frame), and SR calcium-binding proteins (calreticulin, calsequestrin 1, calsequestrin 2, and sarcalumenin) in skeletal muscle (diaphragm and extensor carpi ulnaris) and heart (left ventricle) in normal and affected male dogs by droplet digital PCR before the onset (≤ 2-m-old), at the active stage (8 to 16-m-old), and at the terminal stage (30 to 50-m-old) of the disease. Since many of these proteins are expressed in a fiber type-specific manner, we also evaluated fiber type composition in skeletal muscle. RESULTS: In affected dog skeletal muscle, SERCA and its regulators were down-regulated at the active stage, but calcium-binding proteins (except for calsequestrin 1) were upregulated at the terminal stage. Surprisingly, nominal differences were detected in the heart. We also examined whether there exists sex-biased expression in 8 to 16-m-old dogs. Multiple transcripts were significantly downregulated in the heart and extensor carpi ulnaris muscle of female dogs. In fiber type analysis, we found significantly more type I fiber in the diaphragm of 8 to 16-m-old affected dogs, and significantly more type II fibers in the extensor carpi ulnaris of 30 to 50-m-old affected dogs. However, no difference was detected between male and female dogs. CONCLUSIONS: Our study adds new knowledge to the understanding of muscle calcium regulation in normal and dystrophic canines.

Graded exposure therapy for adults with persistent symptoms after mTBI: A historical comparison study.

Fear avoidance behaviour is associated with slow recovery from mild traumatic brain injury (mTBI). This study is a preliminary evaluation of graded exposure therapy (GET), which directly targets fear avoidance behaviour, for reducing post-concussion symptoms (PCS) and disability following mTBI. In a historical comparison design, we compared two groups from independent randomized trials. The GET + UC group (N = 34) received GET (delivered over 16 videoconference sessions) in addition to usual care (UC). The historical comparison group (N = 71) received UC only. PCS severity (Rivermead Post Concussion Symptoms Questionnaire; RPQ) and disability (World Health Organization Disability Assessment Schedule; WHODAS 2.0 12-item) were measured at clinic intake (M = 2.7, SD = 1.1 months after injury) and again at M = 4.9 (SD = 1.1) months after injury. Between-group differences were estimated using linear mixed effects regression, with a sensitivity analysis controlling for injury-to-assessment intervals. The estimated average change on the RPQ was -14.3 in the GET + UC group and -5.3 in the UC group. The estimated average change on the WHODAS was -5.3 in the GET + UC group and -3.2 in the UC group. Between-group differences post-treatment were -5.3 on the RPQ and -1.5 on the WHODAS. Treatment effects were larger in sensitivity analyses. Findings suggest that a randomized controlled trial is warranted.

Graded Exposure Therapy for Fear Avoidance Behaviour After Concussion (GET FAB): protocol for a multisite Canadian randomised controlled trial.

INTRODUCTION: Persistent symptoms after mild traumatic brain injury (mTBI) negatively affect daily functioning and quality of life. Fear avoidance behaviour, a coping style in which people avoid or escape from activities or situations that they expect will exacerbate their symptoms, maybe a particularly potent and modifiable risk factor for chronic disability after mTBI. This study will evaluate the efficacy of graded exposure therapy (GET) for reducing persistent symptoms following mTBI, with two primary aims: (1) To determine whether GET is more effective than usual care; (2) to identify for whom GET is the most effective treatment option, by evaluating whether baseline fear avoidance moderates differences between GET and an active comparator (prescribed aerobic exercise). Our findings will guide evidence-based care after mTBI and enable better matching of mTBI patients to treatments. METHODS AND ANALYSIS: We will conduct a multisite randomised controlled trial with three arms. Participants (n=220) will be recruited from concussion clinics and emergency departments in three Canadian provinces and randomly assigned (1:2:2 ratio) to receive enhanced usual care, GET or prescribed aerobic exercise. The outcome assessment will occur remotely 14-18 weeks following baseline assessment, after completing the 12-week treatment phase. The primary outcome will be symptom severity (Rivermead Post-concussion Symptoms Questionnaire). ETHICS AND DISSEMINATION: Informed consent will be obtained from all participants. All study procedures were approved by the local research ethics boards (University of British Columbia Clinical Research Ethics Board, University of Calgary Conjoint Health Research Ethics Board, University Health Network Research Ethics Board-Panel D). Operational approvals were obtained for Vancouver Coastal Health Research Institute and Provincial Health Services Authority. If GET proves effective, we will disseminate the GET treatment manual and present instructional workshops for clinicians. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov #NCT05365776.

SERCA2a overexpression improves muscle function in a canine Duchenne muscular dystrophy model.

Excessive cytosolic calcium accumulation contributes to muscle degeneration in Duchenne muscular dystrophy (DMD). Sarco/endoplasmic reticulum calcium ATPase (SERCA) is a sarcoplasmic reticulum (SR) calcium pump that actively transports calcium from the cytosol into the SR. We previously showed that adeno-associated virus (AAV)-mediated SERCA2a therapy reduced cytosolic calcium overload and improved muscle and heart function in the murine DMD model. Here, we tested whether AAV SERCA2a therapy could ameliorate muscle disease in the canine DMD model. 7.83 × 1013 vector genome particles of the AAV vector were injected into the extensor carpi ulnaris (ECU) muscles of four juvenile affected dogs. Contralateral ECU muscles received excipient. Three months later, we observed widespread transgene expression and significantly increased SERCA2a levels in the AAV-injected muscles. Treatment improved SR calcium uptake, significantly reduced calpain activity, significantly improved contractile kinetics, and significantly enhanced resistance to eccentric contraction-induced force loss. Nonetheless, muscle histology was not improved. To evaluate the safety of AAV SERCA2a therapy, we delivered the vector to the ECU muscle of adult normal dogs. We achieved strong transgene expression without altering muscle histology and function. Our results suggest that AAV SERCA2a therapy has the potential to improve muscle performance in a dystrophic large mammal.

Treatment expectations and clinical outcomes following repetitive transcranial magnetic stimulation for treatment-resistant depression.

BACKGROUND: Patient expectations, including both positive (placebo) and negative (nocebo) effects, influence treatment outcomes, yet their impact on acute repetitive transcranial magnetic stimulation (rTMS) for treatment-resistant depression (TRD) is unclear. METHODS: In this single-center retrospective chart review, 208 TRD patients completed the Stanford Expectation of Treatment Scale (SETS) before starting open-label rTMS treatment. Patients were offered two excitatory rTMS protocols (deep TMS or intermittent theta-burst stimulation), which stimulated the left dorsolateral prefrontal cortex. A minimum of 20 once daily treatments were provided, delivered over 4-6 weeks. Primary outcomes were 1) remission, measured by a post-treatment score of <8 on the Hamilton Depression Rating Scale (HAMD-17), and 2) premature discontinuation. The change in HAMD-17 scores over time was used as a secondary outcome. Physicians were blinded to SETS scores. Logistic and linear regression, adjusting for covariates, assessed SETS and HAMD-17 relationships. RESULTS: Of 208 patients, 177 had baseline and covariate data available. The mean positivity bias score (positive expectancy minus negative expectancy subscale averages) was 0.48 ± 2.21, indicating the cohort was neutral regarding the expectations of their treatment on average. Higher positive expectancy scores were significantly associated with greater odds of remission (OR = 1.90, p = 0.003) and greater reduction in HAMD-17 scores (ß = 1.30, p = 0.005) at the end of acute treatment, after adjusting for covariates. Negative expectancy was not associated with decreased odds of remission (p = 0.2) or treatment discontinuation (p = 0.8). CONCLUSIONS: Higher pre-treatment positive expectations were associated with greater remission rates with open-label rTMS in a naturalistic cohort of patients with TRD.

Understanding and including 'pink-collar' workers in employment-based travel demand models.

The segmentation of commuters into either blue or white-collar workers remains is still common in urban transport models. Internationally, models have started to use more elaborate segmentations, more reflective of changes in labour markets, such as increased female participation. Finding appropriate labour market segmentations for commute trip modelling remains a challenge. This paper harnesses a data-driven approach using unsupervised clustering-applied to 2017-20 South East Queensland Travel Survey (SEQTS) data. Commuter types are grouped by occupational, industry, and socio-demographic variables (i.e., gender, age, household size, household vehicle ownership and worker skill score). The results show that at a large number of clusters (i.e., k = 8) a highly distinct set of commuter types can be observed. But model run times tend to require a much smaller number of market segments. When only three clusters are formed (k = 3) a market segmentation emerges with one female-dominated type ('pink collar'), one male-dominated type ('blue collar') and one with both genders almost equally involved ('white collar'). There are nuances as to which workers are included in each segment, and differences in travel behaviours across the three types. 'Pink collar' workers are mostly comprised of female clerical and administrative workers, community and personal service workers and sales workers. They have the shortest median commutes for both private motorised and active transport modes. The approach and methods should assist transport planners to derive more accurate and robust market segmentations for use in large urban transport models, and, better predict the value of alternative transport projects and policies for all types of commuters.

Harnessing Placebo Effects for the Treatment of Functional Cognitive Disorder: A Feasibility Pilot Study.

OBJECTIVE: Limited research has directly investigated whether and how placebo effects can be harnessed for the treatment of functional neurological disorder (FND), despite a long-standing and controversial history of interest in this area. METHODS: A small exploratory study was conducted with adults with a cognitive subtype of FND recruited from a single cognitive neurology center in the United States. Participants were given the expectation of receiving cranial stimulation that could benefit their memory symptoms; however, the intervention was sham transcranial magnetic stimulation (placebo). Outcomes included measures of short-term memory testing, subjective memory rating, and state anxiety before and after stimulation. After the study, the true objective and rationale for investigating placebo effects were explained in a scripted debriefing session. Acceptability of the study design and qualitative feedback were collected. Institutional ethics approval and signed consent were obtained. RESULTS: Three patients (female, N=2; male, N=1; average age=57 years) were recruited. Outcome data were analyzed descriptively at the patient level. Trends of improvement in subjective memory rating, but not objective cognitive test scores, and decreases in state anxiety were observed. After the debriefing session, all patients found the study design to be acceptable (ratings of 70%, 90%, and 100%), and two of the three patients believed that withholding mechanistic information about the intervention was needed to leverage placebo effects as treatment. CONCLUSIONS: In the first study to prospectively investigate the feasibility of harnessing placebo effects for the treatment of FND, promising preliminary findings were obtained, and methods and resources for use in larger future studies are offered.

CONNECTing Concussion Care with Research Across Ontario.

Common data elements (CDEs) for concussion, as established by international bodies, are not being widely used in Ontario, resulting in significant variability in the data being assessed and collected across clinics. CDEs support standardization of care as well as large-scale data sharing for high impact research. A collaborative network - Concussion Ontario Network: Neuroinformatics to Enhance Clinical care and Translation (CONNECT) - comprised of health care professionals, researchers, members from advocacy groups, and patients was formed to establish and implement CDEs for concussion care and research. While the seeds have been planted and initial effectiveness demonstrated, future challenges exist.

A randomized sham-controlled trial of high-dosage accelerated intermittent theta burst rTMS in major depression: study protocol.

BACKGROUND: Intermittent theta burst stimulation (iTBS), a novel form of repetitive transcranial magnetic stimulation (rTMS), can be administered in 1/10th of the time of standard rTMS (~ 3 min vs. 37.5 min) yet achieves similar outcomes in depression. The brief nature of the iTBS protocol allows for the administration of multiple iTBS sessions per day, thus reducing the overall course length to days rather than weeks. This study aims to compare the efficacy and tolerability of active versus sham iTBS using an accelerated regimen in patients with treatment-resistant depression (TRD). As a secondary objective, we aim to assess the safety, tolerability, and treatment response to open-label low-frequency right-sided (1 Hz) stimulation using an accelerated regimen in those who do not respond to the initial week of treatment. METHODS: Over three years, approximately 230 outpatients at the Centre for Addiction and Mental Health and University of British Columbia Hospital, meeting diagnostic criteria for unipolar MDD, will be recruited and randomized to a triple blind sham-controlled trial. Patients will receive five consecutive days of active or sham iTBS, administered eight times daily at 1-hour intervals, with each session delivering 600 pulses of iTBS. Those who have not achieved response by the week four follow-up visit will be offered a second course of treatment, regardless of whether they initially received active or sham stimulation. DISCUSSION: Broader implementation of conventional iTBS is limited by the logistical demands of the current standard course consisting of 4-6 weeks of daily treatment. If our proposed accelerated iTBS protocol enables patients to achieve remission more rapidly, this would offer major benefits in terms of cost and capacity as well as the time required to achieve clinical response. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT04255784.

Quantification and characterization of microplastics in coastal environments: Insights from laser direct infrared imaging.

The study identified and quantified nine plastic polymers frequently detected in the environment by collecting sediment and seawater samples from coastal areas in Auckland, New Zealand. Polymer types, size distributions, and number of microplastics (MPs) were analyzed using a laser direct infrared (LDIR) imaging technique. Compared to conventional spectroscopic or microscopic methods, LDIR enabled capturing and quantifying MPs in much lower size ranges (20-5000 µm). The results demonstrated the widespread occurrence of MPs in the Auckland coastal environment, with polyethylene terephthalate (PET) being the most frequently detected plastic polymer. MP contamination levels ranged from 13 to 83 particles per liter of coastal water and from 1200 to 3400 particles/kg of dry sand in beach sediments. Six additional locations were investigated to assess the contribution of MPs from stormwater drains to the coastal environment. The total count of identified MPs extracted from sediments near stormwater drains reached a maximum of 18,000 particles/kg of dry sand, representing an order of magnitude increase compared to MP levels found in beach sediments at the same location. In contrast to the prevalence of PET and polyamide observed in beach sediments and coastal waters, polyurethane and polyethylene emerged as the predominant plastic polymers in the vicinity of stormwater drain sediments, implying that the variation could potentially stem from distinct sources of plastics. This significant disparity in quality and quantity underscored the potential link between urban runoff and MP pollution in marine ecosystems. A sample preparation method using 100 g sediment samples was developed and used to assess and compare MPs detection in sediment samples. The commonly used 5 g sample method showed higher extraction efficiency and better detection of the most abundant MPs, but the new 100 g method enabled the detection of previously missed, less abundant plastics.

Dynamic regulation of airway surface liquid pH by TMEM16A and SLC26A4 in cystic fibrosis nasal epithelia with rare mutations.

In cystic fibrosis (CF), defects in the CF transmembrane conductance regulator (CFTR) channel lead to an acidic airway surface liquid (ASL), which compromises innate defence mechanisms, predisposing to pulmonary failure. Restoring ASL pH is a potential therapy for people with CF, particularly for those who cannot benefit from current highly effective modulator therapy. However, we lack a comprehensive understanding of the complex mechanisms underlying ASL pH regulation. The calcium-activated chloride channel, TMEM16A, and the anion exchanger, SLC26A4, have been proposed as targets for restoring ASL pH, but current results are contradictory and often utilise nonphysiological conditions. To provide better evidence for a role of these two proteins in ASL pH homeostasis, we developed an efficient CRISPR-Cas9-based approach to knock-out (KO) relevant transporters in primary airway basal cells lacking CFTR and then measured dynamic changes in ASL pH under thin-film conditions in fully differentiated airway cultures, which better simulate the in vivo situation. Unexpectantly, we found that both proteins regulated steady-state as well as agonist-stimulated ASL pH, but only under inflammatory conditions. Furthermore, we identified two Food and Drug Administration (FDA)-approved drugs which raised ASL pH by activating SLC26A4. While we identified a role for SLC26A4 in fluid absorption, KO had no effect on cyclic adenosine monophosphate (cAMP)-stimulated fluid secretion in airway organoids. Overall, we have identified a role of TMEM16A in ASL pH homeostasis and shown that both TMEM16A and SLC26A4 could be important alternative targets for ASL pH therapy in CF, particularly for those people who do not produce any functional CFTR.

Auditory brainstem response (ABR) waveform analysis program.

Auditory brainstem responses (ABR) are a high-throughput assessment of auditory function. Many studies determine changes to the threshold at frequencies that span the normal hearing range of their test subjects, but fewer studies evaluate changes in waveform morphology. The goal of developing this program was to make a user-friendly semiautomatic peak-detection algorithm to encourage widespread analysis of the amplitudes and latencies of the ABR, which may yield informative details about the integrity of the auditory system with development, aging, genetic manipulations, or damaging conditions. This method incorporates automated peak detection with manual override and inter-rater validation to calculate the amplitude and latency for waves 1-5, as well as interpeak latencies and amplitude ratios between waves. The output includes raw data and calculations in a format compatible with graphical and statistical software.•The method yields a high-throughput peak-detection algorithm with manual override and inter-rater capabilities to streamline ABR waveform analysis.•Data output includes amplitudes, latencies, amplitude ratios, and interpeak latencies for generation of input-output curves.•While complete automation of peak detection with this tool is dependent on good signal-to-noise ratios, relevant amplitude and latency calculations are fully automated, and manual spot-checking is simplified to significantly reduce the time to analyze waveforms.

Characterizing the profiles of patients with acute concussion versus prolonged post-concussion symptoms in Ontario.

Identifying vulnerability factors for developing persisting concussion symptoms is imperative for determining which patients may require specialized treatment. Using cross-sectional questionnaire data from an Ontario-wide observational concussion study, we compared patients with acute concussion (≤ 14 days) and prolonged post-concussion symptoms (PPCS) (≥ 90 days) on four factors of interest: sex, history of mental health disorders, history of headaches/migraines, and past concussions. Differences in profile between the two groups were also explored. 110 patients with acute concussion and 96 patients with PPCS were included in our study. The groups did not differ on the four factors of interest. Interestingly, both groups had greater proportions of females (acute concussion: 61.1% F; PPCS: 66.3% F). Patient profiles, however, differed wherein patients with PPCS were significantly older, more symptomatic, more likely to have been injured in a transportation-related incident, and more likely to live outside a Metropolitan city. These novel risk factors for persisting concussion symptoms require replication and highlight the need to re-evaluate previously identified risk factors as more and more concussions occur in non-athletes and different risk factors may be at play.

Litigation, Performance Validity Testing, and Treatment Outcomes in Adults With Mild Traumatic Brain Injury.

OBJECTIVE: To investigate whether involvement in litigation and performance validity test (PVT) failure predict adherence to treatment and treatment outcomes in adults with persistent symptoms after mild traumatic brain injury (mTBI). SETTING: Outpatient concussion clinics in British Columbia, Canada. Participants were assessed at intake (average 12.9 weeks postinjury) and again following 3 to 4 months of rehabilitation. PARTICIPANTS: Adults who met the World Health Organization Neurotrauma Task Force definition of mTBI. Litigation status was known for 69 participants (n = 21 reported litigation), and 62 participants completed a PVT (n = 13 failed the Test of Memory Malingering) at clinic intake. DESIGN: Secondary analysis of a clinical trial (ClinicalTrials.gov #NCT03972579). MAIN MEASURES: Outcomes included number of completed sessions, homework adherence, symptoms (Rivermead Post Concussion Symptoms Questionnaire), disability ratings (World Health Organization Disability Assessment Schedule 2.0), and patient-rated global impression of change. RESULTS: We did not observe substantial differences in session and homework adherence associated with litigation or PVT failure. Disability and postconcussion symptoms generally improved with treatment. Involvement in litigation was associated with a smaller improvement in outcomes, particularly disability (B = 2.57, 95% confidence interval [CI] [0.25-4.89], P = .03) and patient-reported global impression of change (odds ratio [OR] = 4.19, 95% CI [1.40-12.57], P = .01). PVT failure was not associated with considerable differences in treatment outcomes. However, participants who failed the PVT had a higher rate of missing outcomes (31% vs 8%) and perceived somewhat less global improvement (OR = 3.47, 95% CI [0.86-14.04]; P = .08). CONCLUSION: Adults with mTBI who are in litigation or who failed PVTs tend to adhere to and improve following treatment. However, involvement in litigation may be associated with attenuated improvements, and pretreatment PVT failure may predict lower engagement in the treatment process.

Assessing the risk of a clinically significant infection from a Microneedle Array Patch (MAP) product.

Microneedle Array Patches (MAPs) are an emerging dosage form that creates transient micron-sized disruptions in the outermost physical skin barrier, the stratum corneum, to facilitate delivery of active pharmaceutical ingredients to the underlying tissue. Numerous MAP products are proposed and there is significant clinical potential in priority areas such as vaccination. However, since their inception scientists have hypothesized about the risk of a clinically significant MAP-induced infection. Safety data from two major Phase 3 clinical trials involving hundreds of participants, who in total received tens of thousands of MAP applications, does not identify any clinically significant infections. However, the incumbent data set is not extensive enough to make definitive generalizable conclusions. A comprehensive assessment of the infection risk is therefore advised for MAP products, and this should be informed by clinical and pre-clinical data, theoretical analysis and informed opinions. In this article, a group of key stakeholders identify some of the key product- and patient-specific factors that may contribute to the risk of infection from a MAP product and provide expert opinions in the context of guidance from regulatory authorities. Considerations that are particularly pertinent to the MAP dosage form include the specifications of the finished product (e.g. microbial specification), it's design features, the setting for administration, the skill of the administrator, the anatomical application site, the target population and the clinical context. These factors, and others discussed in this article, provide a platform for the development of MAP risk assessments and a stimulus for early and open dialogue between developers, regulatory authorities and other key stakeholders, to expedite and promote development of safe and effective MAP products.

Placebo Effects and Neuromodulation: Ethical Considerations and Recommendations.

Placebo-controlled trials are the gold standard of evaluating treatment efficacy in clinical research. Neuromodulation is emerging as an important treatment pathway for many neuropsychiatric conditions, and placebo control arms of these trials require careful design with unique considerations (e.g., sham devices that mimic active stimulation, blinding effectiveness). Inherent to placebo-controlled trials are ethical concerns, such as deception, and potential harm of not receiving the active treatment. In this article, we outline important ethical considerations of placebo-controlled trials across neuromodulation approaches and provide recommendations on how ethical principles can be adhered to going forward. We specifically address issues of autonomy and respect for persons, beneficence, and justice. Within the context of this ethical framework, we also discuss factors influencing placebo effects in neuromodulation, the importance of adequate blinding, and alternative trial designs that could be considered.

COVID-19 vaccination for the prevention and treatment of long COVID: A systematic review and meta-analysis.

Empirical evidence addressing the association between SARS-CoV-2 vaccination and long COVID would guide public health priorities and inform personal health decisions. Herein, the co-primary objectives are to determine the differential risk of long COVID in vaccinated versus unvaccinated patients, and the trajectory of long COVID following vaccination. Of 2775 articles identified via systematic search, 17 were included, and 6 were meta-analyzed. Meta-analytic results determined that at least one vaccine dose was associated with a protective effect against long COVID (OR 0.539, 95% CI 0.295-0.987, p = 0.045, N = 257 817). Qualitative analysis revealed that trajectories of pre-existing long COVID following vaccination were mixed, with most patients reporting no changes. The evidence herein supports SARS-CoV-2 vaccination for the prevention of long COVID, and recommends long COVID patients adhere to standard SARS-CoV-2 vaccination schedules.