Chromatographic Phospholipid Trapping for Automated H/D Exchange Mass Spectrometry of Membrane Protein-Lipid Assemblies.

Lipid interactions modulate the function, folding, structure, and organization of membrane proteins. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) has emerged as a useful tool to understand the structural dynamics of these proteins within lipid environments. Lipids, however, have proven problematic for HDX-MS analysis of membrane-embedded proteins due to their presence of impairing proteolytic digestion, causing liquid chromatography column fouling, ion suppression, and/or mass spectral overlap. Herein, we describe the integration of a chromatographic phospholipid trap column into the HDX-MS apparatus to enable online sample delipidation prior to protease digestion of deuterium-labeled protein-lipid assemblies. We demonstrate the utility of this method on membrane scaffold protein-lipid nanodisc─both empty and loaded with the ∼115 kDa transmembrane protein AcrB─proving efficient and automated phospholipid capture with minimal D-to-H back-exchange, peptide carry-over, and protein loss. Our results provide insights into the efficiency of phospholipid capture by ZrO2-coated and TiO2 beads and describe how solution conditions can be optimized to maximize not only the performance of our online but also the existing offline, delipidation workflows for HDX-MS. We envision that this HDX-MS method will significantly ease membrane protein analysis, allowing to better interrogate their dynamics in artificial lipid bilayers or even native cell membranes.

The influence of community health on hospitals attainment of Magnet designation: Implications for policy and practice.

AIMS: To determine if there is an association between better County Health Rankings and the increased odds of a hospital gaining Magnet designation in subsequent years (2014-2019) compared with counties with lower rankings. BACKGROUND: The Magnet hospital model is recognized to have a great effect on nurses, patients and organizational outcomes. Although Magnet hospital designation is a well-established structural marker for nursing excellence, the effect of County Health Rankings and subsequent hospital achievement of Magnet status is unknown. DESIGN: A descriptive, cross-sectional quantitative approach was adopted for this study. METHODS: Data were derived from 2010 to 2019 U.S. County Health Rankings, American Hospital Association, and American Nursing Credentialing Center databases. Logistic regression models were utilized to determine associations between county rankings for health behaviours, clinical care, social and economic factors, physical environment and counties with a new Magnet hospital after 2014. RESULTS: Counties with the worst rankings for clinical care and socio-economic status had reduced odds of obtaining a Magnet hospital designation compared with best-ranking counties. While middle-ranking counties for the physical environment ranking had increased odds of having Magnet designation compared with best-ranking counties. Additionally, having an increased percent of government non-federal hospital or a higher percentage of critical access hospitals in the county reduced the odds of having a Magnet-designated facility after 2014. CONCLUSION: The findings underscore the important associations between Magnet-designated facilities' location and the health of its surrounding counties. This study is the first to examine the relationship between County Health Rankings and a hospital's likelihood of obtaining Magnet status and points to the need for future research to explore outcomes of care previously identified as improved in Magnet-designated hospitals. IMPLICATIONS: Recognizing the benefits of Magnet facilities, it is important for health care leaders and policy makers to seek opportunities to promote centres of excellence in higher need communities through policy and financial intervention.

Grandparents' Mental Health and Lived Experiences while Raising Their Grandchildren at the Forefront of COVID-19 in Saudi Arabia.

Understanding grandparents' lived experiences and healthy aging is essential to designing efficient, effective, and safe services to support a family structure in which grandparents care for their grandchildren. However, no study to date has explored this concept in an Arab and Muslim country during a pandemic. The purpose of this study was to examine grandparents' experiences raising their grandchildren to provide recommendations for needed mental health interventions during and after COVID-19. We used a phenomenological approach to gain a detailed and in-depth understanding of the lived experiences of 15 grandparents caring for their grandchildren. This study shows the need for support service interventions (support groups, health professional support, and respite care) for grandparents in Saudi Arabia, especially during global crises like COVID-19, that enhance social distance and social isolation. Raising grandchildren affects the physical, mental, and social wellbeing of the grandparents.

Expression profile based gene clusters for ischemic stroke detection.

In microarray studies alterations in gene expression in circulating leukocytes have shown utility for ischemic stroke diagnosis. We studied forty candidate markers identified in three gene expression profiles to (1) quantitate individual transcript expression, (2) identify transcript clusters and (3) assess the clinical diagnostic utility of the clusters identified for ischemic stroke detection. Using high throughput next generation qPCR 16 of the 40 transcripts were significantly up-regulated in stroke patients relative to control subjects (p<0.05). Six clusters of between 5 and 7 transcripts were identified that discriminated between stroke and control (p values between 1.01e-9 and 0.03). A 7 transcript cluster containing PLBD1, PYGL, BST1, DUSP1, FOS, VCAN and FCGR1A showed high accuracy for stroke classification (AUC=0.854). These results validate and improve upon the diagnostic value of transcripts identified in microarray studies for ischemic stroke. The clusters identified show promise for acute ischemic stroke detection.

Structural Equation Modeling: Effects of Master Adaptive Learner and Clinical Learning Environment Attributes on Career Satisfaction and Intention to Stay.

OBJECTIVE: The objective of this observational study was to test a proposed conceptual model depicting the influence of personal and environmental attributes of clinical learning environments on rehabilitation professionals' career satisfaction and intention to stay with their current organization. The mediating effects of 2 loci of engagement were also assessed. METHODS: Rehabilitation professionals (physical therapists, occupational therapists, speech-language pathologists, and registered nurses; n = 436) from 4 health care organizations in the Northeast, Southeast, and Midwest regions in the USA completed a battery of questionnaires related to personal attributes (resilience and orientation toward lifelong learning), environmental attributes (innovation support and basic psychological need satisfaction at work), loci of engagement (professional and organizational), career satisfaction, and intention to stay. Data were analyzed using structural equation modeling. RESULTS: The measurement and structural model fit was acceptable. The greatest total effects on career satisfaction (r = .459) and intention to stay (r = .462) were observed through the influences of basic psychological need satisfaction, professional engagement, and organizational engagement. Orientation toward lifelong learning was negatively associated with career satisfaction (r = -.208) and intention to stay (r = -.154), but this was mitigated by organizational engagement (r = -.060 and - .134, respectively). Small but significant total effects were also observed between innovation support, professional and organizational engagement, and intention to stay (r = .144) and resilience, professional and organizational engagement, and career satisfaction (r = .110). CONCLUSION: Clinical learning environments that support rehabilitation professionals' basic psychological needs (autonomy, competency, and relatedness) are associated with greater career satisfaction and intention to stay. This association is further enhanced by organizational engagement and innovation support. IMPACT: Rehabilitation organizations interested in developing and retaining master adaptive learners should create clinical learning environments that reinforce rehabilitation professionals' sense of autonomy, competency, and relatedness, and are supportive on innovative behaviors.

Student Physical Therapists' Orientation to Postprofessional Education and Perceptions of Strain: Identification of 4 Subgroups of Students Based on Cluster Analysis.

INTRODUCTION: Determining why physical therapists choose certain pathways to expertise is not well understood. Developing an understanding of these different choices is important for the physical therapy profession and the future of postprofessional education. REVIEW OF LITERATURE: Pathways to expertise as a physical therapist have evolved over the history of the profession, including the most recent emergence of residency education. Regardless of the path taken, physical therapists have a societal duty to be lifelong learners. The decision on which path to pursue is influenced by many factors, including financial stress, personal factors, and relational obligations. Developing an understanding of these different pathways and barriers for physical therapists is essential if the physical therapy profession is to advance with the evolving needs of society. SUBJECTS: Participants were from a stratified purposive sample of Doctor of Physical Therapy Students (n = 124) across the eastern, central, and western regions of the United States. METHODS: This study was a secondary analysis using hierarchical cluster analysis based on students' orientation toward lifelong learning and future professional engagement, self-rated likelihood of pursuing various postprofessional education pathways, and perception of personal and interpersonal strains as a barrier to postprofessional education. RESULTS: The cluster analysis resulted in a 4-cluster solution representing different student subtypes: clinically oriented average learners; resilient, clinically oriented lifelong learners; academically oriented lifelong learners; and strain-limited, clinically oriented lifelong learners. DISCUSSION AND CONCLUSION: Results from this study describe important differences in how student physical therapists view postprofessional education and the pathways they choose to pursue. Factors such as lack of awareness and the perception of personal or interpersonal strain may influence these decisions. Understanding postprofessional education decision making of student physical therapists is of particular interest to the physical therapy profession given the current debates related to student debt and the necessity of residency education. Further research is needed to understand how to increase awareness and diminish barriers to quality postprofessional education for physical therapists.

Covid-19 Impacts on Florida's Healthcare Professionals.

This study described how COVID-19 impacted employment, telehealth usage, and interprofessional collaboration. A cross-sectional survey was deployed in June 2020 to healthcare professionals in Florida. Job status was uniquely separated by profession, with more nurses and medical doctors reported having no effect, and more mental health counselors transitioned to telehealth. Over a third of rehabilitation providers reported being furloughed. Over forty percent of providers had no training in telehealth, yet 33.1% reported an increase in usage. Interprofessional interactions are lower across professions during the pandemic, compared with before. This study shows the need for additional training on telehealth and interprofessional collaboration.

Older and Wiser? The Need to Reexamine the Impact of Health Professionals Age and Experience on Competency-Based Practices.

INTRODUCTION: Delivery of healthcare services makes up a complex system and it requires providers to be competent and to be able to integrate each of the institute of medicine's (IOM) 5 core competencies into practice. However, healthcare providers are challenged with the task to be able to understand and apply the IOM core competencies into practice. OBJECTIVE: The purpose of the study was to examine the factors that influence health professional's likelihood of accomplishing the IOM core competencies. METHODS: A cross-sectional study design was used to administer a validated online survey to health providers. This survey was distributed to physicians, nursing professionals, specialists, and allied healthcare professionals. The final sample included 3,940 participants who completed the survey. RESULTS: The study findings show that younger health professionals more consistently practice daily competencies than their older counterparts, especially in the use of evidence-based practice, informatics, and working in interdisciplinary teams. Less experienced health professionals more consistently applied quality improvement methods but less consistently used evidence-based practice compared to their more experienced counterparts. CONCLUSION: There is a need to understand how health professionals' age and experience impact their engagement with IOM's core competencies. This study highlights the need for educational resources on the competencies to be tailored to health providers' age and experience.

Pre-existing hypertension dominates γδT cell reduction in human ischemic stroke.

T lymphocytes may play an important role in the evolution of ischemic stroke. Depletion of γδT cells has been found to abrogate ischemia reperfusion injury in murine stroke. However, the role of γδT cells in human ischemic stroke is unknown. We aimed to determine γδT cell counts and γδT cell interleukin 17A (IL-17A) production in the clinical setting of ischemic stroke. We also aimed to determine the associations of γδT cell counts with ischemic lesion volume, measures of clinical severity and with major stroke risk factors. Peripheral blood samples from 43 acute ischemic stroke patients and 26 control subjects matched on race and gender were used for flow cytometry and complete blood count analyses. Subsequently, cytokine levels and gene expression were measured in γδT cells. The number of circulating γδT cells was decreased by almost 50% (p = 0.005) in the stroke patients. γδT cell counts did not correlate with lesion volume on magnetic resonance diffusion-weighted imaging or with clinical severity in the stroke patients, but γδT cells showed elevated levels of IL-17A (p = 0.048). Decreased γδT cell counts were also associated with older age (p = 0.004), pre-existing hypertension (p = 0.0005) and prevalent coronary artery disease (p = 0.03), with pre-existing hypertension being the most significant predictor of γδT cell counts in a multivariable analysis. γδT cells in human ischemic stroke are reduced in number and show elevated levels of IL-17A. A major reduction in γδT lymphocytes also occurs in hypertension and may contribute to the development of hypertension-mediated stroke and vascular disease.

Next-generation qPCR for the high-throughput measurement of gene expression in multiple leukocyte subsets.

Clinical studies of gene expression are increasingly using the whole blood, peripheral blood mononuclear cells, and leukocyte subsets involved in the innate and adaptive immune responses. However, the small amount of RNA available in the clinical setting is a limitation for commonly used methods such as quantitative polymerase chain reactions (qPCR) and microarrays. Our aim was to design 96 gene assays to simultaneously measure gene expression in the whole blood and seven leukocyte subsets using a new-generation qPCR method--high-throughput nanofluidic reverse transcription qPCR (HT RT-qPCR). The leukocyte subset purity was 94% to 98% for seven subsets and was less for the γδ T-cell receptor subset (80%). The HT RT-qPCR replicate sample measurements were highly reproducible (r = 0.997, p < 2.2 × 10(-16)), and the ΔΔCt values from HT RT-qPCR correlated significantly with those from qPCR. The control genes were differentially expressed across the eight leukocyte subsets in the control subjects (p = 1.3 × 10(-5), analysis of variance). Two analytical methods, absolute and relative, gave concordant results and were significantly correlated (p = 1.9 × 10(-9)). HT RT-qPCR permits the rapid, reproducible, and quantitative measurement of multiple transcripts using minimal sample amounts. The protocol described yielded leukocyte subsets of high purity and identified two analytic methods for use.