What's the CATCH? A Participatory Learning Model for Case Review Rounds to Support a Culture of Safety and Quality Improvement in Pediatric Hospital Medicine.

OBJECTIVE: To evaluate the effectiveness of a new model, Case Analysis and Translation to Care in Hospital (CATCH), for the review of pediatric inpatient cases when an adverse event or "close call" had occurred. STUDY DESIGN: The curricular intervention consisted of an introductory podcast/workshop, mentorship of presenters, and monthly CATCH rounds over 16 months. The study was conducted with 22 pediatricians at a single tertiary care center. Intervention assessment occurred using participant surveys at multiple intervals: pre/post the intervention, presenter experience (post), physicians involved and mentors experience (post), and after each CATCH session. Paired t-tests and thematic analysis were used to analyze data. Time required to support the CATCH process was used to assess feasibility. RESULTS: Our overall experience and data revealed a strong preference for the CATCH model, high levels of engagement and satisfaction with CATCH sessions, and positive presenter as well as physicians-involved and mentor experiences. Participants reported that the CATCH model is feasible, engages physicians, promotes a safe learning environment, facilitates awareness of tools for case analysis, and provides opportunities to create "CATCH of the Day" recommendations to support translation of learning to clinical practice. CONCLUSIONS: The CATCH model has significant potential to strengthen clinical case rounds in pediatric hospital medicine. Future research is needed to assess the effectiveness of the model at additional sites and across medical specialities.

Monitoring methane emissions from oil and gas operations‡.

The atmospheric concentration of methane has more than doubled since the start of the Industrial Revolution. Methane is the second-most-abundant greenhouse gas created by human activities and a major driver of climate change. This APS-Optica report provides a technical assessment of the current state of monitoring U.S. methane emissions from oil and gas operations, which accounts for roughly 30% of U.S. anthropogenic methane emissions. The report identifies current technological and policy gaps and makes recommendations for the federal government in three key areas: methane emissions detection, reliable and systematized data and models to support mitigation measures, and effective regulation.

CD73+ CD127high Long-Term Memory CD4 T Cells Are Highly Proliferative in Response to Recall Antigens and Are Early Targets in HIV-1 Infection.

HIV-1 infection rapidly leads to a loss of the proliferative response of memory CD4+ T lymphocytes, when cultured with recall antigens. We report here that CD73 expression defines a subset of resting memory CD4+ T cells in peripheral blood, which highly express the α-chain of the IL-7 receptor (CD127), but not CD38 or Ki-67, yet are highly proliferative in response to mitogen and recall antigens, and to IL-7, in vitro. These cells also preferentially express CCR5 and produce IL-2. We reasoned that CD73+ memory CD4+ T cells decrease very early in HIV-1 infection. Indeed, CD73+ memory CD4+ T cells comprised a median of 7.5% (interquartile range: 4.5-10.4%) of CD4+ T cells in peripheral blood from healthy adults, but were decreased in primary HIV-1 infection to a median of 3.7% (IQR: 2.6-6.4%; p = 0.002); and in chronic HIV-1 infection to 1.9% (IQR: 1.1-3%; p < 0.0001), and were not restored by antiretroviral therapy. Moreover, we found that a significant proportion of CD73+ memory CD4+ T cells were skewed to a gut-homing phenotype, expressing integrins α4 and ß7, CXCR3, CCR6, CD161 and CD26. Accordingly, 20% of CD4+ T cells present in gut biopsies were CD73+. In HIV+ subjects, purified CD73+ resting memory CD4+ T cells in PBMC were infected with HIV-1 DNA, determined by real-time PCR, to the same level as for purified CD73-negative CD4+ T cells, both in untreated and treated subjects. Therefore, the proliferative CD73+ subset of memory CD4+ T cells is disproportionately reduced in HIV-1 infection, but, unexpectedly, their IL-7 dependent long-term resting phenotype suggests that residual infected cells in this subset may contribute significantly to the very long-lived HIV proviral DNA reservoir in treated subjects.

Potassium disturbance associated with an inpatient childhood asthma pathway.

BACKGROUND: Paediatric asthma exacerbations in Alberta are treated via standardized order sets known as the Alberta Acute Childhood Asthma Pathway (ACAP). This pathway is utilized in paediatric tertiary hospitals and in remote and rural locations. Incidence, magnitude, and risk factors for hypokalemia in inpatients receiving salbutamol for asthma exacerbations via this pathway are presently unknown. OBJECTIVE: Establish incidence, magnitude, and risk factors for hypokalemia associated with salbutamol therapy as directed by a paediatric asthma pathway. METHODS: Retrospective cohort study using visit-level electronic medical data. Inpatients aged <18 years old receiving salbutamol via the ACAP with at least one potassium level were included. Hypokalemia was defined as mild (3.0 ≤ [K+] < 3.5 mEq/L), moderate (2.5 ≤ [K+] < 3.0 mEq/L), or severe ([K+] < 2.5 mEq/L), as measured in serum or blood gas. Binomial logistic regression was utilized to examine risk factors for hypokalemia, route of administration, location of lowest [K+], nil per os (NPO) status during admission, potassium supplementation, gender, and age. RESULTS: There were 821 patients screened for analysis and 433 patients were analyzed after exclusions. There was an incidence of hypokalemia of 38.8%. Of patients experiencing hypokalemia, 71.4% were mild, 25.6% moderate, and 3.0% severe. Risk factors included nebulized salbutamol, patient location (emergency department or paediatric intensive care unit), and age (>5 years) although these risk factors may actually represent patients receiving higher doses of salbutamol. CONCLUSIONS: The majority of the 38.8% of children experiencing hypokalemia associated with the ACAP were mild. Routine monitoring of potassium status in children receiving salbutamol per standardized pathway is recommended for children with described risk factors, and ideally within the first 12 hours of presentation.

HIV DNA subspecies persist in both activated and resting memory CD4+ T cells during antiretroviral therapy.

UNLABELLED: The latent HIV reservoir is a major impediment to curing HIV infection. The contribution of CD4(+) T cell activation status to the establishment and maintenance of the latent reservoir was investigated by enumerating viral DNA components in a cohort of 12 individuals commencing antiretroviral therapy (ART) containing raltegravir, an integrase inhibitor. Prior to ART, the levels of total HIV DNA were similar across HLA-DR(+) and HLA-DR(-) (HLA-DR(±)) CD38(±) memory CD4(+) T cell phenotypes; episomal two-long terminal repeat (2-LTR) HIV DNA levels were higher in resting (HLA-DR(-) CD38(-)) cells, and this phenotype exhibited a significantly higher ratio of 2-LTR to integrated HIV DNA (P = 0.002). After 1 year of ART, there were no significant differences across each of the memory phenotypes of any HIV DNA component. The decay dynamics of integrated HIV DNA were slow within each subset, and integrated HIV DNA in the resting HLA-DR(-) CD38(-) subset per mm(3) of peripheral blood exhibited no significant decay (half-life of 25 years). Episomal 2-LTR HIV DNA decayed relative to integrated HIV DNA in resting cells with a half-life of 134 days. Surprisingly, from week 12 on, the decay rates of both total and episomal HIV DNA were lower in activated CD38(+) cells. By weeks 24 and 52, HIV RNA levels in plasma were most significantly correlated with the numbers of resting cells containing integrated HIV DNA. On the other hand, total HIV DNA levels in all subsets were significantly correlated with the numbers of HLA-DR(+) CD38(-) cells containing integrated HIV DNA. These results provide insights into the interrelatedness of cell activation and reservoir maintenance, with implications for the design of therapeutic strategies targeting HIV persistence. IMPORTANCE: It is generally believed that HIV is not cleared by extensive antiretroviral therapy (ART) due to the difficulty in eradicating the latent reservoir in resting CD4(+) T cells. New therapies that attempt to activate this reservoir so that immune or viral cytopathic mechanisms can remove those infected cells are currently being investigated. However, results obtained in this research indicate that activation, at least on some level, already occurs within this reservoir. Furthermore, we are the first to describe the dynamics of different HIV DNA species in resting and activated memory CD4+ T cell subsets that point to the role different levels of activation play in maintaining the HIV reservoir.

Ratios of effector to central memory antigen-specific CD4(+) T cells vary with antigen exposure in HIV+ patients.

The OX40/CD25 assay is a novel technique that assesses antigen-specific CD4(+) T-cell responses. To unequivocally demonstrate that responding cells are memory cells that become activated after secondary stimulation, naïve CD45RA(+) and memory CD45RO(+) populations were stimulated with cytomegalovirus (CMV) lysate and the combined expression of CD25 and OX40 measured. As expected, the naïve population showed very little response, whereas there was a higher response from the memory counterpart. To further elucidate CD4(+) memory T-cell subsets involved in recall responses, CD4(+) T cells were separated into central memory (Tcm) and effector memory (Tem) subsets and stimulated with antigen-pulsed antigen-presenting cells (APCs). CMV responses in healthy donors showed a Tem-dominant response with a Tem/Tcm ratio of 1.2, whereas the tetanus toxoid responses were dominated by a Tcm response with a Tem/Tcm ratio of 0.35. To determine memory response in the chronic of HIV infection, patient samples were used. A similar pattern to healthy donors was observed in seven chronic HIV+ patients at week 4 after anti-retroviral therapy who responded to CMV with a larger response coming from Tem. The pattern was similar after 48 weeks of therapy but the responses were lower in magnitude. In chronic HIV+ patients who respond to Gag peptides, following institution of therapy there was an inversion of the ratio of the responding memory subsets compared with week 4, with a greater response from Tcm at week 48. This result was concordant with reduction in antigen load. As immune activation decreased there was also a decrease in the percentage of responding effector memory cells and maintenance of long-term central memory.

Simian immunodeficiency virus infects follicular helper CD4 T cells in lymphoid tissues during pathogenic infection of pigtail macaques.

T follicular helper (Tfh) cells are a specialized subset of memory CD4(+) T cells that are found exclusively within the germinal centers of secondary lymphoid tissues and are important for adaptive antibody responses and B cell memory. Tfh cells do not express CCR5, the primary entry coreceptor for both human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV), and therefore, we hypothesized that these cells would avoid infection. We studied lymph nodes and spleens from pigtail macaques infected with pathogenic strain SIVmac239 or SIVmac251, to investigate the susceptibility of Tfh cells to SIV infection. Pigtail macaque PD-1(high) CD127(low) memory CD4(+) T cells have a phenotype comparable to that of human Tfh cells, expressing high levels of CXCR5, interleukin-21 (IL-21), Bcl-6, and inducible T cell costimulator (ICOS). As judged by either proviral DNA or cell-associated viral RNA measurements, macaque Tfh cells were infected with SIV at levels comparable to those in other CD4(+) memory T cells. Infection of macaque Tfh cells was evident within weeks of inoculation, yet we confirmed that Tfh cells do not express CCR5 or either of the well-known alternative SIV coreceptors, CXCR6 and GPR15. Mutations in the SIV envelope gp120 region occurred in chronically infected macaques but were uniform across each T cell subset investigated, indicating that the viruses used the same coreceptors to enter different cell subsets. Early infection of Tfh cells represents an unexpected focus of viral infection. Infection of Tfh cells does not interrupt antibody production but may be a factor that limits the quality of antibody responses and has implications for assessing the size of the viral reservoir.

HIV disease progression despite suppression of viral replication is associated with exhaustion of lymphopoiesis.

The mechanisms of CD4(+) T-cell count decline, the hallmark of HIV disease progression, and its relationship to elevated levels of immune activation are not fully understood. Massive depletion of CD4(+) T cells occurs during the course of HIV-1 infection, so that maintenance of adequate CD4(+) T-cell levels probably depends primarily on the capacity to renew depleted lymphocytes, that is, the lymphopoiesis. We performed here a comprehensive study of quantitative and qualitative attributes of CD34(+) hematopoietic progenitor cells directly from the blood of a large set of HIV-infected persons compared with uninfected donors, in particular the elderly. Our analyses underline a marked impairment of primary immune resources with the failure to maintain adequate lymphocyte counts. Systemic immune activation emerges as a major correlate of altered lymphopoiesis, which can be partially reversed with prolonged antiretroviral therapy. Importantly, HIV disease progression despite elite control of HIV replication or virologic success on antiretroviral treatment is associated with persistent damage to the lymphopoietic system or exhaustion of lymphopoiesis. These findings highlight the importance of primary hematopoietic resources in HIV pathogenesis and the response to antiretroviral treatments.

How the COVID-19 pandemic shaped adolescents' future orientations: Insights from a global scoping review.

Throughout adolescence, youth imagine what the future holds and determine plans to achieve their educational, professional, and personal goals. In this article, we review research that explores how adolescents' future orientations were shaped by the societal unpredictability, physical and mental health risks, and educational disruptions brought on by the COVID-19 pandemic. Findings show that the pandemic, which exacerbated existing societal inequities, also heightened adolescents' social awareness, provoked feelings of uncertainty, and altered adolescents' short- and long-term plans for educational and career prospects. Throughout school building closures and program cancellations, families played a greater role in fostering adolescents' plans. With prospects for more societal uncertainty on the horizon, future directions point toward supporting adolescents in developing adaptable and flexible future orientations.

High levels of human antigen-specific CD4+ T cells in peripheral blood revealed by stimulated coexpression of CD25 and CD134 (OX40).

Ag-specific human CD4(+) memory T lymphocytes have mostly been studied using assays of proliferation in vitro. Intracellular cytokine and ELISPOT assays quantify effector cell populations but barely detect responses to certain recall Ags that elicit strong proliferative responses, e.g., tetanus toxoid, that comprise non-Th1 CD4(+) cells. We have found that culturing whole blood with Ag for 40-48 h induces specific CD4(+) T cells to simultaneously express CD25 and CD134. This new technique readily detects responses to well-described CD4(+) T cell recall Ags, including preparations of mycobacteria, CMV, HSV-1, influenza, tetanus toxoid, Candida albicans, and streptokinase, as well as HIV-1 peptides, with high specificity. The assay detects much higher levels of Ag-specific cells than intracellular cytokine assays, plus the cells retain viability and can be sorted for in vitro expansion. Furthermore, current in vitro assays for human CD4(+) memory T lymphocytes are too labor-intensive and difficult to standardize for routine diagnostic laboratories, whereas the whole-blood CD25(+)CD134(+) assay combines simplicity of setup with a straightforward cell surface flow cytometry readout. In addition to revealing the true extent of Ag-specific human CD4(+) memory T lymphocytes, its greatest use will be as a simple in vitro monitor of CD4(+) T cell responses to Ags such as tuberculosis infection or vaccines.

Predicting the Refractive Index of Tissue Models Using Light Scattering Spectroscopy.

In this work, we report the application of Raman microspectroscopy for analysis of the refractive index of a range of tissue phantoms. Using both a custom-developed setup with visible laser source and a commercial microspectrometer with near infrared laser, we measured the Raman spectra of gelatin hydrogels at various concentrations. By building a calibration curve from measured refractometry data and Raman scattering intensity for different vibrational modes of the hydrogel, we were able to predict the refractive indices of the gels from their Raman spectra. This work highlights the importance of a correlative approach through Brillouin-Raman microspectroscopy for the mechano-chemical analysis of biologically relevant samples.

Brillouin microscopy for the evaluation of hair micromechanics and effect of bleaching.

Brillouin microscopy is a new form of optical elastography and an emerging technique in mechanobiology and biomedical physics. It was applied here to map the viscoelastic properties of human hair and to determine the effect of bleaching on hair properties. For hair samples, longitudinal measurements (i.e. along the fibre axis) revealed peaks at 18.7 and 20.7 GHz at the location of the cuticle and cortex, respectively. For hair treated with a bleaching agent, the frequency shifts for the cuticle and cortex were 19.7 and 21.0 GHz, respectively, suggesting that bleaching increases the cuticle modulus and-to a minor extent-the cortex modulus. These results demonstrate the capability of Brillouin spectroscopy to address questions on micromechanical properties of hair and to validate the effect of applied treatments.

Accelerating Rare Disease Drug Development: Lessons Learned from Muscular Dystrophy Patient Advocacy Groups.

With scientific and molecular advancements related to disease pathogenesis, advances in gene and stem cell therapies, and the promise of lucrative markets for biopharmaceutical companies, there has been a rapid expansion in the number of potential new muscular dystrophy (MD) treatments. The first champion for a newly diagnosed MD patient and their caregivers is typically an MD-specific patient advocacy group (PAG). Muscular dystrophy PAGs have been among the most active in the rare disease drug development space. Notable achievements in the last decade include promulgating the first U.S. clinical research guidance, setting up registries and natural history studies, and investing in companies-some of which have brought potentially disease-modifying products to the market. This paper will discuss five key strategies that have been successfully employed by MD PAGs to advance treatments: (1) creating a national registry, (2) understanding the barriers to identifying patients with certain subtypes of muscular dystrophy to participate in clinical trials, (3) partnering with the biopharmaceutical industry, (4) collaborating with the regulators, and (5) incorporating market access and use insights early in clinical development. While clearly helpful within the MD community, these tactics could also be employed by PAGs representing other types of rare diseases.

Preservation of Gastrointestinal Mucosal Barrier Function and Microbiome in Patients With Controlled HIV Infection.

Background: Despite successful ART in people living with HIV infection (PLHIV) they experience increased morbidity and mortality compared with HIV-negative controls. A dominant paradigm is that gut-associated lymphatic tissue (GALT) destruction at the time of primary HIV infection leads to loss of gut integrity, pathological microbial translocation across the compromised gastrointestinal barrier and, consequently, systemic inflammation. We aimed to identify and measure specific changes in the gastrointestinal barrier that might allow bacterial translocation, and their persistence despite initiation of antiretroviral therapy (ART). Method: We conducted a cross-sectional study of the gastrointestinal (GIT) barrier in PLHIV and HIV-uninfected controls (HUC). The GIT barrier was assessed as follows: in vivo mucosal imaging using confocal endomicroscopy (CEM); the immunophenotype of GIT and circulating lymphocytes; the gut microbiome; and plasma inflammation markers Tumour Necrosis Factor-α (TNF-α) and Interleukin-6 (IL-6); and the microbial translocation marker sCD14. Results: A cohort of PLHIV who initiated ART early, during primary HIV infection (PHI), n=5), and late (chronic HIV infection (CHI), n=7) infection were evaluated for the differential effects of the stage of ART initiation on the GIT barrier compared with HUC (n=6). We observed a significant decrease in the CD4 T-cell count of CHI patients in the left colon (p=0.03) and a trend to a decrease in the terminal ileum (p=0.13). We did not find evidence of increased epithelial permeability by CEM. No significant differences were found in microbial translocation or inflammatory markers in plasma. In gut biopsies, CD8 T-cells, including resident intraepithelial CD103+ cells, did not show any significant elevation of activation in PLHIV, compared to HUC. The majority of residual circulating activated CD38+HLA-DR+ CD8 T-cells did not exhibit gut-homing integrins α4ß7, suggesting that they did not originate in GALT. A significant reduction in the evenness of species distribution in the microbiome of CHI subjects (p=0.016) was observed, with significantly higher relative abundance of the genus Spirochaeta in PHI subjects (p=0.042). Conclusion: These data suggest that substantial, non-specific increases in epithelial permeability may not be the most important mechanism of HIV-associated immune activation in well-controlled HIV-positive patients on antiretroviral therapy. Changes in gut microbiota warrant further study.

Seroprevalence of SARS-CoV-2 infection in Cincinnati Ohio USA from August to December 2020.

The world is currently in a pandemic of COVID-19 (Coronavirus disease-2019) caused by a novel positive-sense, single-stranded RNA ß-coronavirus referred to as SARS-CoV-2. Here we investigated rates of SARS-CoV-2 infection in the greater Cincinnati, Ohio, USA metropolitan area from August 13 to December 8, 2020, just prior to initiation of the national vaccination program. Examination of 9,550 adult blood donor volunteers for serum IgG antibody positivity against the SARS-CoV-2 Spike protein showed an overall prevalence of 8.40%, measured as 7.56% in the first 58 days and 9.24% in the last 58 days, and 12.86% in December 2020, which we extrapolated to ~20% as of March, 2021. Males and females showed similar rates of past infection, and rates among Hispanic or Latinos, African Americans and Whites were also investigated. Donors under 30 years of age had the highest rates of past infection, while those over 60 had the lowest. Geographic analysis showed higher rates of infectivity on the West side of Cincinnati compared with the East side (split by I-75) and the lowest rates in the adjoining region of Kentucky (across the Ohio river). These results in regional seroprevalence will help inform efforts to best achieve herd immunity in conjunction with the national vaccination campaign.

Brillouin microspectroscopy data of tissue-mimicking gelatin hydrogels.

Brillouin spectroscopy, based on the inelastic scattering of light from thermally driven acoustic waves or phonons [1], holds great promise in the field of life sciences as it provides functionally relevant micromechanical information in a contactless all-optical manner [2]. Due to the complexity of biological systems such as cells and tissues, which present spatio-temporal heterogeneities, interpretation of Brillouin spectra can be difficult. The data presented here were collected from gelatin hydrogels, used as tissue-mimicking model systems for Brillouin microspectroscopy measurements conducted using a lab-built Brillouin microscope with a dual-stage VIPA spectrometer. By varying the solute concentration in the range 4-18% (w/w), the macroscopic mechanical properties of the hydrogels can be tuned and the corresponding evolution in the Brillouin-derived longitudinal elastic modulus measured. An increase in Brillouin frequency shift with increasing solute concentration was observed, which was found to correlate with an increase in acoustic wave velocity and longitudinal modulus. The gels used here provide a viable model system for benchmarking and standardisation, and the data will be useful for spectrometer development and validation.

Viscoelastic properties of biopolymer hydrogels determined by Brillouin spectroscopy: A probe of tissue micromechanics.

Many problems in mechanobiology urgently require characterization of the micromechanical properties of cells and tissues. Brillouin light scattering has been proposed as an emerging optical elastography technique to meet this need. However, the information contained in the Brillouin spectrum is still a matter of debate because of fundamental problems in understanding the role of water in biomechanics and in relating the Brillouin data to low-frequency macroscopic mechanical parameters. Here, we investigate this question using gelatin as a model system in which the macroscopic physical properties can be manipulated to mimic all the relevant biological states of matter, ranging from the liquid to the gel and the glassy phase. We demonstrate that Brillouin spectroscopy is able to reveal both the elastic and viscous properties of biopolymers that are central to the structure and function of biological tissues.

Image analysis applied to Brillouin images of tissue-mimicking collagen gelatins.

Brillouin spectroscopy is an emerging analytical tool in biomedical and biophysical sciences. It probes viscoelasticity through the propagation of thermally induced acoustic waves at gigahertz frequencies. Brillouin light scattering (BLS) measurements have traditionally been performed using multipass Fabry-Pérot interferometers, which have high contrast and resolution, however, as they are scanning spectrometers they often require long acquisition times in poorly scattering media. In the last decade, a new concept of Brillouin spectrometer has emerged, making use of highly angle-dispersive virtually imaged phase array (VIPA) etalons, which enable fast acquisition times for minimally turbid materials, when high contrast is not imperative. The ability to acquire Brillouin spectra rapidly, together with long term system stability, make this system a viable candidate for use in biomedical applications, especially to probe live cells and tissues. While various methods are being developed to improve system contrast and speed, little work has been published discussing the details of imaging data analysis and spectral processing. Here we present a method that we developed for the automated retrieval of Brillouin line shape parameters from imaging data sets acquired with a dual-stage VIPA Brillouin microscope. We applied this method for the first time to BLS measurements of collagen gelatin hydrogels at different hydration levels and cross-linker concentrations. This work demonstrates that it is possible to obtain the relevant information from Brillouin spectra using software for real-time high-accuracy analysis.