Mn2+ -Phos-Tag Polyacrylamide for the Quantification of Protein Phosphorylation Levels.

This paper provides a guideline for optimizing and utilizing Mn2+ Phos-tag gel technology to separate phosphorylated proteins from their unphosphorylated counterparts. It provides key insights into methods for careful sample preparation and experimental directions for determining the appropriate Phos-tag gel compositions and electrophoresis and western blotting conditions. This protocol has been used to successfully resolve proteins extracted from cardiac and skeletal muscles. The guidelines can be extended for optimizing protocols to resolve proteins from other cells or tissue sources. With this, phosphoproteomics and the elucidation of underlying mechanisms of disease progression can be accelerated. © 2021 The Authors. Current Protocols published by Wiley Periodicals LLC.

Regulatory Light Chains in Cardiac Development and Disease.

The role of regulatory light chains (RLCs) in cardiac muscle function has been elucidated progressively over the past decade. The RLCs are among the earliest expressed markers during cardiogenesis and persist through adulthood. Failing hearts have shown reduced RLC phosphorylation levels and that restoring baseline levels of RLC phosphorylation is necessary for generating optimal force of muscle contraction. The signalling mechanisms triggering changes in RLC phosphorylation levels during disease progression remain elusive. Uncovering this information may provide insights for better management of heart failure patients. Given the cardiac chamber-specific expression of RLC isoforms, ventricular RLCs have facilitated the identification of mature ventricular cardiomyocytes, opening up possibilities of regenerative medicine. This review consolidates the standing of RLCs in cardiac development and disease and highlights knowledge gaps and potential therapeutic advancements in targeting RLCs.

Functional and Molecular Characterisation of Heart Failure Progression in Mice and the Role of Myosin Regulatory Light Chains in the Recovery of Cardiac Muscle Function.

Heart failure (HF) as a result of myocardial infarction (MI) is a major cause of fatality worldwide. However, the cause of cardiac dysfunction succeeding MI has not been elucidated at a sarcomeric level. Thus, studying the alterations within the sarcomere is necessary to gain insights on the fundamental mechansims leading to HF and potentially uncover appropriate therapeutic targets. Since existing research portrays regulatory light chains (RLC) to be mediators of cardiac muscle contraction in both human and animal models, its role was further explored In this study, a detailed characterisation of the physiological changes (i.e., isometric force, calcium sensitivity and sarcomeric protein phosphorylation) was assessed in an MI mouse model, between 2D (2 days) and 28D post-MI, and the changes were related to the phosphorylation status of RLCs. MI mouse models were created via complete ligation of left anterior descending (LAD) coronary artery. Left ventricular (LV) papillary muscles were isolated and permeabilised for isometric force and Ca2+ sensitivity measurement, while the LV myocardium was used to assay sarcomeric proteins' (RLC, troponin I (TnI) and myosin binding protein-C (MyBP-C)) phosphorylation levels and enzyme (myosin light chain kinase (MLCK), zipper interacting protein kinase (ZIPK) and myosin phosphatase target subunit 2 (MYPT2)) expression levels. Finally, the potential for improving the contractility of diseased cardiac papillary fibres via the enhancement of RLC phosphorylation levels was investigated by employing RLC exchange methods, in vitro. RLC phosphorylation and isometric force potentiation were enhanced in the compensatory phase and decreased in the decompensatory phase of HF failure progression, respectively. There was no significant time-lag between the changes in RLC phosphorylation and isometric force during HF progression, suggesting that changes in RLC phosphorylation immediately affect force generation. Additionally, the in vitro increase in RLC phosphorylation levels in 14D post-MI muscle segments (decompensatory stage) enhanced its force of isometric contraction, substantiating its potential in HF treatment. Longitudinal observation unveils potential mechanisms involving MyBP-C and key enzymes regulating RLC phosphorylation, such as MLCK and MYPT2 (subunit of MLCP), during HF progression. This study primarily demonstrates that RLC phosphorylation is a key sarcomeric protein modification modulating cardiac function. This substantiates the possibility of using RLCs and their associated enzymes to treat HF.

Digital health professions education on chronic wound management: A systematic review.

BACKGROUND: Continuing education is crucial for healthcare professionals to keep up with research but attending classroom lectures is a major barrier. Chronic wound management is increasingly relevant for continuous professional training. Digital education offers learning tailored to individual needs and could be an effective alternative to healthcare professionals' training. However, the effectiveness of digital education for chronic wound management training has not been explored. OBJECTIVES: To assess the effectiveness of digital education in improving healthcare professionals' knowledge, attitudes, practical skills and behaviour change on chronic wound management, and their satisfaction with the intervention. DESIGN: This systematic review follows Cochrane methodology and is one of a series of reviews on the use of digital education for health professions education. Protocol registration: PROSPERO CRD42018109971 DATA SOURCES: Searches were conducted in MEDLINE, Embase, Web of Science, ERIC, PsycINFO, CINAHL, CENTRAL, and ProQuest Dissertation and Theses Database. REVIEW METHODS: We included randomised control trials, cluster randomised control trials and quasi-randomised control trials comparing digital or blended education with traditional learning, no intervention or other forms of digital or blended education for pre- or post-registration healthcare professionals in chronic wound management. A narrative summary of findings is presented. RESULTS: Seven studies (1,404 participants) were included. All studies investigated interventions for nursing students or professionals working in hospitals or community settings, and all but one study focused on pressure ulcers. Five studies (935 participants) assessed post-intervention knowledge, and indicated that digital education was more effective than no intervention, while blended learning was superior to exclusive digital education. Three studies (543 participants) assessed post-intervention skills and reported mixed results. One study (140 participants) compared post-intervention behaviour change and satisfaction with blended and online digital education, and reported no difference in behaviour between the groups, and higher satisfaction with blended education. For knowledge retention up to six months, digital education was more effective than no intervention, while blended learning was superior to digital education. The risk of bias in included studies was mostly high or unclear. CONCLUSIONS: Digital education on chronic wound management appears to be less effective than blended education and more effective than no intervention in improving knowledge among nurses and nursing students. Data for other outcomes is scarce and inconclusive. Future studies should assess participants' skills, attitudes, satisfaction and behaviour change; cost-effectiveness and potential untoward effects of digital education, compare digital education to other learning modalities and include other healthcare professionals in diverse clinical settings.

Digital Education for the Management of Chronic Wounds in Health Care Professionals: Protocol for a Systematic Review by the Digital Health Education Collaboration.

BACKGROUND: Digital education is "the act of teaching and learning by means of digital technologies." Digital education comprises a wide range of interventions that can be broadly divided into offline digital education, online digital education, digital game-based learning, massive open online courses (MOOCs), psychomotor skills trainers, virtual reality environments, virtual patient simulations, and m-learning. Chronic wounds pose an immense economic and psychosocial burden to patients and the health care system, as caring for them require highly specialized personnel. Current training strategies face significant barriers, such as lack of time due to work commitments, distance from provider centers, and costs. Therefore, there is an increased need to synthesize evidence on the effectiveness of digital education interventions on chronic wounds management in health care professionals. OBJECTIVE: Our main objective is to assess the effectiveness of digital education as a stand-alone approach or as part of a blended-learning approach in improving pre- and postregistration health care professionals' knowledge, attitudes, practical skills, and behavior in the management of chronic wounds, as well as their satisfaction with the intervention. Secondary objectives are to evaluate patient-related outcomes, cost-effectiveness of the interventions, and any unfavorable or undesirable outcomes that may arise. METHODS: This systematic review will follow the methodology as described in the Cochrane Handbook for Systematic Reviews of Interventions. As our systematic review is one of a series of reviews on digital education for health professionals' education, we will use a previously developed search strategy. This search includes the following databases: the Cochrane Central Register of Controlled Trials (CENTRAL) (Cochrane Library), MEDLINE (Ovid), Embase (Ovid), Web of Science, the Educational Resource Information Centre (ERIC) (Ovid), PsycINFO (Ovid), the Cumulative Index to Nursing and Allied Health Literature (CINAHL) (EBSCO), the ProQuest Dissertation and Theses database, and trial registries. Databases will be searched for studies published from January 1990 to August 2018. Two independent reviewers will screen the library for included studies. We will describe the screening process using a flowchart as per the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. We will extract the data using a previously developed, structured data extraction form. Included studies will be quality-assessed using the Risk of Bias tool from Cochrane. We will narratively summarize the data and, if possible, we will conduct a meta-analysis. We will use Cochrane's RevMan 5.3 software for data analysis. RESULTS: We have completed the screening of titles and abstracts for this systematic review and are currently selecting papers against our inclusion and exclusion criteria through full-text revision. We are expecting to complete our review by the end of April 2019. CONCLUSIONS: This systematic review will provide an in-depth analysis of digital education strategies to train health care providers in the management of chronic wounds. We consider this topic particularly relevant given the current challenges facing health care systems worldwide, including shortages of skilled personnel and a steep increase in the population of older adults as a result of a prolonged life expectancy. TRIAL REGISTRATION: PROSPERO CRD42018109971; https://www.crd.york.ac.uk/prospero/display_record.php?RecordID=109971. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/12488.