Structural Insights into Hearing Loss Genetics from Polarizable Protein Repacking.

Hearing loss is associated with ∼8100 mutations in 152 genes, and within the coding regions of these genes are over 60,000 missense variants. The majority of these variants are classified as "variants of uncertain significance" to reflect our inability to ascribe a phenotypic effect to the observed amino acid change. A promising source of pathogenicity information is biophysical simulation, although input protein structures often contain defects because of limitations in experimental data and/or only distant homology to a template. Here, we combine the polarizable atomic multipole optimized energetics for biomolecular applications force field, many-body optimization theory, and graphical processing unit acceleration to repack all deafness-associated proteins and thereby improve average structure MolProbity score from 2.2 to 1.0. We then used these optimized wild-type models to create over 60,000 structures for missense variants in the Deafness Variation Database, which are being incorporated into the Deafness Variation Database to inform deafness pathogenicity prediction. Finally, this work demonstrates that advanced polarizable atomic multipole force fields are efficient enough to repack the entire human proteome.

Association of Patient Characteristics and Tumor Genomics With Clinical Outcomes Among Patients With Non-Small Cell Lung Cancer Using a Clinicogenomic Database.

Importance: Data sets linking comprehensive genomic profiling (CGP) to clinical outcomes may accelerate precision medicine. Objective: To assess whether a database that combines EHR-derived clinical data with CGP can identify and extend associations in non-small cell lung cancer (NSCLC). Design, Setting, and Participants: Clinical data from EHRs were linked with CGP results for 28 998 patients from 275 US oncology practices. Among 4064 patients with NSCLC, exploratory associations between tumor genomics and patient characteristics with clinical outcomes were conducted, with data obtained between January 1, 2011, and January 1, 2018. Exposures: Tumor CGP, including presence of a driver alteration (a pathogenic or likely pathogenic alteration in a gene shown to drive tumor growth); tumor mutation burden (TMB), defined as the number of mutations per megabase; and clinical characteristics gathered from EHRs. Main Outcomes and Measures: Overall survival (OS), time receiving therapy, maximal therapy response (as documented by the treating physician in the EHR), and clinical benefit rate (fraction of patients with stable disease, partial response, or complete response) to therapy. Results: Among 4064 patients with NSCLC (median age, 66.0 years; 51.9% female), 3183 (78.3%) had a history of smoking, 3153 (77.6%) had nonsquamous cancer, and 871 (21.4%) had an alteration in EGFR, ALK, or ROS1 (701 [17.2%] with EGFR, 128 [3.1%] with ALK, and 42 [1.0%] with ROS1 alterations). There were 1946 deaths in 7 years. For patients with a driver alteration, improved OS was observed among those treated with (n = 575) vs not treated with (n = 560) targeted therapies (median, 18.6 months [95% CI, 15.2-21.7] vs 11.4 months [95% CI, 9.7-12.5] from advanced diagnosis; P < .001). TMB (in mutations/Mb) was significantly higher among smokers vs nonsmokers (8.7 [IQR, 4.4-14.8] vs 2.6 [IQR, 1.7-5.2]; P < .001) and significantly lower among patients with vs without an alteration in EGFR (3.5 [IQR, 1.76-6.1] vs 7.8 [IQR, 3.5-13.9]; P < .001), ALK (2.1 [IQR, 0.9-4.0] vs 7.0 [IQR, 3.5-13.0]; P < .001), RET (4.6 [IQR, 1.7-8.7] vs 7.0 [IQR, 2.6-13.0]; P = .004), or ROS1 (4.0 [IQR, 1.2-9.6] vs 7.0 [IQR, 2.6-13.0]; P = .03). In patients treated with anti-PD-1/PD-L1 therapies (n = 1290, 31.7%), TMB of 20 or more was significantly associated with improved OS from therapy initiation (16.8 months [95% CI, 11.6-24.9] vs 8.5 months [95% CI, 7.6-9.7]; P < .001), longer time receiving therapy (7.8 months [95% CI, 5.5-11.1] vs 3.3 months [95% CI, 2.8-3.7]; P < .001), and increased clinical benefit rate (80.7% vs 56.7%; P < .001) vs TMB less than 20. Conclusions and Relevance: Among patients with NSCLC included in a longitudinal database of clinical data linked to CGP results from routine care, exploratory analyses replicated previously described associations between clinical and genomic characteristics, between driver mutations and response to targeted therapy, and between TMB and response to immunotherapy. These findings demonstrate the feasibility of creating a clinicogenomic database derived from routine clinical experience and provide support for further research and discovery evaluating this approach in oncology.

Assessment of Compassion in Physician Assistant Students.

PURPOSE: It has been shown that empathy erodes in the later phases of education of both medical students and physician assistant (PA) students. This brief study assessed compassion in PA students at different levels of training to explore if a similar phenomenon occurs. METHODS: Students at entry, midway, and at the end of PA training were surveyed using Pommier's Compassion Scale. RESULTS: Although overall compassion scores were lower when comparing students further along in training, differences were not significant. Females scored significantly higher on the kindness subscale overall. Scores for the common humanity subscale declined for both males and females. CONCLUSIONS: Unlike scales measuring empathy have shown, scores using a compassion rating scale did not decline with progression through PA school. Further research is needed to elucidate differences in subscales of compassion.

Effect of a Flipped Classroom Compared With a Traditional Lecture on Physician Assistant Students' Exam Performance.

PURPOSE: The purpose of this study was to retrospectively compare the exam performance of physician assistant (PA) students given asthma instruction in a flipped classroom with PA students in a traditional lecture setting while controlling for students' previous academic performance and clinical asthma experience. METHODS: Three cohorts of PA students (n = 146) from the years 2017 (traditional-lecture setting) and 2018 and 2019 (flipped-classroom setting) were included in the study. Academic performance across cohorts was compared using answers to 11 exam questions reflective of the asthma content. RESULTS: Findings demonstrated significantly greater performance in the flipped classroom compared with traditional lecture. The 2018 and 2019 cohorts scored 9.4% and 13.2% higher, respectively, compared with the 2017 cohort. Exam performance of students with a low likelihood of clinical exposure to asthmatic patients before PA school was similar to those with a high likelihood. CONCLUSION: This study found improved exam performance with a flipped classroom. The flipped classroom represents a potential opportunity to maximize similar performances by both less experienced students and more experienced students.

A framework to evaluate whether to pool or separate behaviors in a multilayer network.

A multilayer network approach combines different network layers, which are connected by interlayer edges, to create a single mathematical object. These networks can contain a variety of information types and represent different aspects of a system. However, the process for selecting which information to include is not always straightforward. Using data on 2 agonistic behaviors in a captive population of monk parakeets (Myiopsitta monachus), we developed a framework for investigating how pooling or splitting behaviors at the scale of dyadic relationships (between 2 individuals) affects individual- and group-level social properties. We designed 2 reference models to test whether randomizing the number of interactions across behavior types results in similar structural patterns as the observed data. Although the behaviors were correlated, the first reference model suggests that the 2 behaviors convey different information about some social properties and should therefore not be pooled. However, once we controlled for data sparsity, we found that the observed measures corresponded with those from the second reference model. Hence, our initial result may have been due to the unequal frequencies of each behavior. Overall, our findings support pooling the 2 behaviors. Awareness of how selected measurements can be affected by data properties is warranted, but nonetheless our framework disentangles these efforts and as a result can be used for myriad types of behaviors and questions. This framework will help researchers make informed and data-driven decisions about which behaviors to pool or separate, prior to using the data in subsequent multilayer network analyses.

Osteoblasts response to microstructured and nanostructured polyimide film, processed by the use of silica bead microlenses.

The surface of polyimide films was modified by the use of silica microspheres as microlenses to focus radiation emitted by an excimer laser. The resultant surface had both microstructures and nanostructures. Physical and chemical characterization was performed by atomic force and Fourier transform-infrared microscopy. Laser processing resulted in surfaces that had similar roughness but different component frequencies. Chemical changes were not observed with the techniques used. The response of osteoblasts to the surface was assayed by measuring their metabolic activity and the enzyme alkaline phosphatase activity, after 24 hours of growth. Cytoskeleton and expression were both investigated. Metabolic activity was similar on treated and untreated samples. Total cell number and size were increased on microstructured polymer, where specific structures were observed (protrusions). Adhesion was noted, and the actin cytoskeleton showed normal morphology. Cells on nanostructured samples had a diffuse actin network and less mature adhesions as compared with the control. FROM THE CLINICAL EDITOR: Polyimide films with microstructure and nanostructure surface elements were studied from the standpoint of osteoblast response. Total cell number and size were increased on microstructured polymer and protrusions were also observed. Adhesion was noted and the actin cytoskeleton exhibited normal morphology. Cells on nanostructured samples had a diffuse actin network and less mature adhesions.

Sedation Patterns and Hyperosmolar Therapy in Emergency Departments were Associated with Blood Pressure Variability and Outcomes in Patients with Spontaneous Intracranial Hemorrhage.

BACKGROUND: Spontaneous intracranial hemorrhage (sICH) is associated with high mortality. Little information exists to guide initial resuscitation in the emergency department (ED) setting. However, blood pressure variability (BPV) and mechanical ventilation (MV) are known risk factors for poor outcome in sICH. OBJECTIVES: The objective was to examine the associations between BPV and MV in ED (EDMV) and between two ED interventions - post-MV sedation and hyperosmolar therapy for elevated intracranial pressure - and BPV in the ED and in-hospital mortality. METHODS: We retrospectively studied adults with sICH and external ventricular drainage who were transferred to a quaternary academic medical center from other hospitals between January 2011 and September 2015. We used multivariable linear and logistic regressions to measure associations between clinical factors, BPV, and outcomes. RESULTS: We analyzed ED records from 259 patients. There were 143 (55%) EDMV patients who had more severe clinical factors and significantly higher values of all BPV indices than NoEDMV patients. Two clinical factors and none of the severity scores (i.e., Hunt and Hess, World Federation of Neurological Surgeons Grades, ICH score) correlated with BPV. Hyperosmolarity therapy without fluid resuscitation positively correlated with all BPV indices, whereas propofol infusion plus a narcotic negatively correlated with one of them. Two BPV indices, i.e., successive variation of blood pressure (BPSV) and absolute difference in blood pressure between ED triage and departure (BPDepart - Triage), were significantly associated with increased mortality rate. CONCLUSION: Patients receiving MV had significantly higher BPV, perhaps related to disease severity. Good ED sedation, hyperosmolar therapy, and fluid resuscitation were associated with less BPV and lower likelihood of death.

Sorption kinetic of aroma compounds by edible bio-based films from marine-by product macromolecules: Effect of relative humidity conditions.

Edible films based on gelatin and chitosan have high gas and aroma barrier properties. This study focused on their capability to sorbed/retain aroma compounds (1-hexanal, 2-hexen-1-ol, 1-hexanol, 3-hexanone and phenol) at three relative humidity level (≤2%, 53% or 84% RH). Whatever the relative humidity condition, the order of sorption is keton (3-hexanone) < aldehyde (1-hexanal) < aliphatic alcohols (2-hexen-1-ol and 1-hexanol) < phenol. This order could be related to the intrinsic chemical properties of aroma compounds. The increase in moisture enhanced the sorption at the highest RH for all the aroma compounds. However, a competition between water and aliphatic alcohols is observed at 53%RH. All compounds have an ideal sorption behaviour (logarithmic increase) except 1-hexanal. The sorption of 1-hexanal, 1-hexanol, 2-hexen-1-ol and 3-hexanone induced an antiplasticization of the network by increasing the film Tg by more than 5 °C. On the contrary, phenol was an efficient plasticizer at least as high as moisture.

Real-World Evidence In Support Of Precision Medicine: Clinico-Genomic Cancer Data As A Case Study.

The majority of US adult cancer patients today are diagnosed and treated outside the context of any clinical trial (that is, in the real world). Although these patients are not part of a research study, their clinical data are still recorded. Indeed, data captured in electronic health records form an ever-growing, rich digital repository of longitudinal patient experiences, treatments, and outcomes. Likewise, genomic data from tumor molecular profiling are increasingly guiding oncology care. Linking real-world clinical and genomic data, as well as information from other co-occurring data sets, could create study populations that provide generalizable evidence for precision medicine interventions. However, the infrastructure required to link, ensure quality, and rapidly learn from such composite data is complex. We outline the challenges and describe a novel approach to building a real-world clinico-genomic database of patients with cancer. This work represents a case study in how data collected during routine patient care can inform precision medicine efforts for the population at large. We suggest that health policies can promote innovation by defining appropriate uses of real-world evidence, establishing data standards, and incentivizing data sharing.

Cyanine5-doped silica nanoparticles as ultra-bright immunospecific labels for model circulating tumour cells in flow cytometry and microscopy.

In this work, ultra-bright fluorescent silica nanoparticles (NPs) labels have been shown to selectively bind to a model circulating tumour cell (CTC) line, MCF-7, a metastatic breast cancer by targeting epithelial cellular adhesion molecule (EpCAM) present on the MCF-7 cell membrane. Silica NPs approximately 40nm in diameter were doped with different concentrations of Cyanine5 dye molecules, using the reverse microemulsion method. The NPs were two orders of magnitude brighter than Cyanine5 free dye and the measured fluorescence intensity matched a homo-Förster Resonance Energy Transfer model. NPs were conjugated with anti-EpCAM antibody to the NP surface for immunospecific targeting. In flow cytometry experiments the NPs were twice as bright as two commercial anti-EpCAM red fluorophore conjugates, APC and AlexaFluor®647. This increase is achieved while keeping non-specific binding low as established in control tests with a non-metastatic cancer cell line (HeLa). The NPs were also immunospecific in fluorescence microscopy experiments performed at room temperature on non-fixed cells. Confocal microscopy was used to confirm the NPs were located on the surface of the cells, matching with the location of the EpCAM marker. These NPs labels have excellent potential in biomedical diagnostics, particularly when high signal to noise and good photostability are needed, for example, in point-of-care testing.

Investigating the colloidal stability of fluorescent silica nanoparticles under isotonic conditions for biomedical applications.

Fluorescent silica nanoparticle (NP) labels are of great interest in biomedical diagnostics, however, when used in bioassays under physiological conditions they rapidly agglomerate and precipitate from solution leading to high levels of non-specific binding. In this work, using size and zeta-potential data obtained from Dynamic and Electrophoretic Light Scattering analysis, the improvement in colloidal stability of silica NPs under physiological conditions was correlated with an increase in the concentration of three additives: (1) a protein, bovine serum albumin (BSA); (2) a neutral surfactant, Tween 20®; and (3) a charged surfactant, sodium dodecyl sulfate (SDS). The number of BSA molecules present in the NP corona at each concentration was calculated using UV-Vis spectroscopy and a bicinchoninic acid protein assay (BCA). The optimal concentration of each additive was also effective in stabilizing antibody labeled fluorescent nanoparticles (αNPs) under physiological conditions. Using a fourth additive, trehalose, the colloidal stability of αNPs after freeze-drying and long-term storage also significantly improved. Both as-prepared and freeze-dried αNPs were tested in a standard fluorescence immunoassay for the detection of human IgG. The as-prepared assay showed a higher sensitivity at low concentration and a lower limit of detection when compared to a free dye assay. Assays performed with freeze dried αNPs after 4 and 22 days also showed good reproducibility.

The effect of intraluminal contact mediated guidance signals on axonal mismatch during peripheral nerve repair.

The current microsurgical gold standard for repairing long gap nerve injuries is the autograft. Autograft provides a protective environment for repair and a natural internal architecture, which is essential for regeneration. Current clinically approved hollow nerve guidance conduits allow provision of this protective environment; however they fail to provide an essential internal architecture to the regenerating nerve. In the present study both structured and unstructured intraluminal collagen fibres are investigated to assess their ability to enhance conduit mediated nerve repair. This study presents a direct comparison of both structured and unstructured fibres in vivo. The addition of intraluminal guidance structures was shown to significantly decrease axonal dispersion within the conduit and reduced axonal mismatch of distal nerve targets (p < 0.05). The intraluminal fibres were shown to be successfully incorporated into the host regenerative process, acting as a platform for Schwann cell migration and axonal regeneration. Ultimately the fibres were able to provide a platform for nerve regeneration in a long term regeneration study (16 weeks) and facilitated increased guidance of regenerating axons towards their distal nerve targets.

Straightforward, one-step fabrication of ultrathin thermoresponsive films from commercially available pNIPAm for cell culture and recovery.

The use of thermoresponsive surfaces as platforms for cell culture and cell regeneration has been explored over the last couple of decades. Poly-N-isopropylacrylamide (pNIPAm) is a well characterized thermoresponsive polymer which has an aqueous lower critical solution temperature (LCST) in a physiologically useful range, which allows it to reversibly attract (T < 32 °C) and repel water (T > 32 °C). It is this phenomenon that is exploited in temperature-controlled cell harvesting. pNIPAm coatings are generally poorly cell compatible and a number of complex or expensive techniques have been developed in order to overcome this issue. This study seeks to design a simple one-step system whereby commercially sourced pNIPAm is used to achieve similar results. Films were deposited using the operationally simple but rheologically complex spin coating technique. Reversible temperature modulated cell adhesion was achieved using a variety of different cell lines. This system offers a simplistic and cheaper alternative to methods used elsewhere.

Enhancing transformative learning in physician assistant education.

Much of what educators try to impart to PA students involves learning that is not objective or easily measured. An important part of PA training involves personal growth and experiencing new ideas, beliefs, cultures, and ways of life that are, at times, antithetical to a student's frame of reference. Transformative learning is a theory that helps structure this part of education. Utilizing the framework and parameters of transformative learning can help the PA educator meet the goals of personal growth in the students.

Effect of functionalized micropatterned PLGA on guided neurite growth.

When coaptation is not possible in the repair of nerve injuries, a bridge of biomaterial scaffold provides a structural support for neuronal cell growth and guides nerve regeneration. Poly(lactide-co-glycolide) (PLGA) scaffolds have been widely investigated for neural tissue engineering applications. In order to investigate guided neurite growth, we have fabricated micropatterns on PLGA films using laser ablation methods. The micropatterned PLGA films were coated with collagen type I or laminin peptide (PPFLMLLKGSTR) to promote axon growth. Micropatterned PLGA films provide a guidance effect on both early stage neurite outgrowth and elongation. Small (5 microm) grooves showed more statistically significant parallel neurite growth compared with larger size grooves (10 microm). Micropatterned PLGA films coated with laminin peptide showed more parallel neurite growth compared with those coated with collagen type I. Primary neurite number and total neurite length per cell decreased on micropatterned PLGA films compared with the controls. Neurites showed a preference for growth in the microgrooves rather than on the spaces. This study indicates that surface micropatterned structures with conjugated functional molecules can be used to guide neurite growth.

Exploring bullying: Implications for nurse educators.

This article examines briefly the issue of workplace violence and bullying in the hospital environment, but more importantly how the same and different styles of bullying and intra-staff bullying are emerging in nurse education. The content describes the aetiology of violence and bullying and their place in the National Health Service (NHS) including nursing. It explores bullying as the principle form of intimidation in nurse education, the different types and subtle forms of bullying, why individuals become bullies, dealing with and the consequences of bullying. The legislation, guidelines, policies are part of the recommendations for practice.

Comparison of cell interactions with laser machined micron- and nanoscale features in polymer.

Control of cell responses to artificial surfaces is a research goal for much of the biomaterials community. The role that the micron scale topography of a surface can play in controlling cell responses has been well documented and recent advances in nanofabrication techniques have lead to an interest in cells' responses to submicron-scale surface features. The study described here compares the relative influences that nanoscale and micron-scale features exert on cells by examining cytoskeletal organisation. Micron-scale structures were generated on the polyamide Kapton using a 193 nm ArF Excimer laser, at 400 mJ/cm2 fluence. Nanoscale features were generated on Kapton using the excimer laser with a phase mask. Osteoblasts were seeded onto surfaces for 24 h, then the cell membranes were detergent-extracted, and the cells were applied with a primary antibody to actin and a colloidal gold-conjugated secondary antibody. Samples to be examined using the confocal were mounted in glycerol, those for electron microscopy were carbon-coated. The organisation of actin was examined on micron- and nano-scale structures by scoring sections for order of branching and angles of branching to relate changes in the cytoskeleton relative to the control. Although there was a strong influence of micron-scale structures, the cytoskeleton of cells on the nanoscale structures were similar to the controls.