Active Learning Among Health Professions' Educators: Perceptions, Barriers, and Use.

Introduction: Active learning engages students in the learning process through meaningful learning activities. Despite evidence that active learning can improve student's comprehension and problem solving, many educators remain reluctant to adopt it. The goal of this study was to explore health professions' educators' perceptions of active learning and identify implementation barriers. Materials and Methods: We developed a 25-question survey based on the Miller and Metz "perceptions of active learning" survey. We added 12 single-response demographics questions to the original 13 survey questions. Results: One hundred three respondents completed the survey. We found positive perceptions of active learning significantly correlated with gender, rank, teaching FTE, and full-time employment. The use of specific active learning modalities significantly correlated with gender, terminal degree, institutional appointment, academic rank, and role. Lack of time to develop materials and lack of class time were the most common personal barriers identified, while being lecture-accustomed and lack of training were the most common perceived barriers to the implementation of active learning by their peers. Conclusion: Despite overwhelmingly positive perceptions of active learning among US health professions' educators and desire to incorporate it, a gap still exists between institutional and educators' support of active learning due to implementation barriers for resource-intensive active learning.

Counteracting Vaccine Misinformation: An Active Learning Module.

Rises in vaccine hesitancy and the incidence of vaccine-preventable illnesses is, in part, due to an increase in vaccine misinformation. Consequently, many patients express skepticism and mistrust of vaccines. It is important that future clinicians are well equipped to understand vaccine-related literature to prepare them for difficult conversations with patients. This module incorporated various active learning approaches to evaluate vaccine-related literature, discuss true contraindications for vaccination, and aid students in approaching patient-clinician conversations about vaccines. Data gained from delivery of this module suggests that students benefit from gaining knowledge and cultivating communication skills about vaccines early in health professions education.

Measuring vaccine acceptance and knowledge within health professions education.

BACKGROUND: Recent rises in the incidence of vaccine-preventable illnesses and suboptimal vaccine acceptance are considered a consequence of accumulating misinformation. Evidence-based approaches to patient-provider communication are key to addressing vaccine hesitancy. OBJECTIVES: The aim of this study was to assess vaccination attitudes and foundational knowledge among healthcare professions students. METHODS: A 72-item survey was developed to assess vaccine attitudes and knowledge about vaccination among health professions students. The survey incorporated 14 demographics questions, 41 attitude questions, and 17 knowledge questions. 16 of the attitude questions, derived from a set of core questions used to diagnose vaccine hesitancy from the WHO Strategic Advisory Group of Experts on Immunization (SAGE) Vaccine Hesitancy Matrix, were analyzed together to derive a vaccine acceptance score. RESULTS: 295 anonymous survey responses were collected between July 2019 and November 2020. Respondents represented students enrolled in medical, dental, pharmacy, optometry, and biomedical science health professions programs. Respondents scored 82.0% ± 0.8% (mean ± standard error of the mean) on questions that gauged vaccine acceptance. The mean vaccine acceptance score was 85.4% ± 1.0% for medical students and 88.0% ± 1.6% for biomedical science students. The mean knowledge score across all programs was 67.7% ± 1.1%. The greatest proficiency in knowledge scores was seen amongst medical students (79.0% ± 1.3%). CONCLUSIONS: Amongst the different health professions, students in the fields of medicine and biomedical sciences had the highest levels of vaccine acceptance attitudes and knowledge. The vaccine acceptance score can be utilized by health professions educators to guide vaccine education for future health professionals to better prepare them to address vaccine hesitancy and educate patients on vaccination.

Immune-Mediated Renal Diseases: A Team-Based Learning Module for Preclinical Medical Students.

INTRODUCTION: The underlying immune mechanisms of renal diseases are complex, and medical students benefit from learning these complexities in the setting of team-based learning (TBL), where they can apply foundational immunologic concepts to clinical problems. METHODS: This immune-mediated renal disease TBL module focusing on glomerular disease was delivered over a period of 2 consecutive years in an integrated cardiovascular, pulmonary, and renal block within the second semester of the first year of medical school. Each class consisted of 53 students, for a total of 106 students. The TBL module consisted of a seven-question individual and team readiness assurance test (iRAT and tRAT) and two clinically relevant team application activities. Following the TBL module, students completed an 18-item survey that collected information on student perspectives of engagement. RESULTS: For the 2-year period, the iRAT average scores were 73% and 74%, and the tRAT average scores were 100% and 99%, respectively. The survey assessing student perceptions of engagement yielded positive feedback. DISCUSSION: In summary, this TBL module effectively integrates the foundational and clinical sciences to teach higher-order concepts in immunology and renal disease pathogenesis to medical students.

A Tool for Evaluating Session-Level Integration in Medical Education.

Integration in medical education is generally defined as a multidisciplinary approach to the delivery of the basic and applied sciences. In medical education, the number of integrated curricula reported has steadily increased, yet the degree of integration often varies. The purpose of this study was to evaluate a tool for medical educators to assess and improve the level of trans-disciplinary integration within an individual academic session. The Session Integration Tool (SIT) is an analytic, developmental rubric that allows the user to assess session-level integration given three criteria: development, delivery, and outcomes. To evaluate the tool, participants utilized the SIT to assess two exemplar case studies, and then their own educational session. A survey was administered to assess user satisfaction along with the instrument's usefulness and impact on practice. Statistical analysis demonstrated a significant difference between user scores of the exemplar case studies for each paired set of criteria (p < 0.000) and for total scores (p = 0.043). Users also successfully used the instrument to assess their own session. The SIT was highly rated by users in terms of satisfaction, usefulness, and impact on practice, with average scores for each category ranging from agree to strongly agree. The SIT has statistical validity in its ability to discriminate between varied levels of integration. Users found the SIT to be useful and impactful for medical education practice and believe it to support peer- and self-assessment of curricular integration. The SIT is a useful, evidence-based, theory grounded tool for assessing session-level integration.

Generation of three induced pluripotent stem cell lines (SCVIi014-A, SCVIi015-A, and SCVIi016-A) from patients with LQT1 caused by heterozygous mutations in the KCNQ1 gene.

Congenital long QT syndrome type 1 (LQT1) results from KCNQ1 mutations that cause loss of Kv7.1 channel function, leading to arrhythmias, syncope, and sudden cardiac death. Here, we generated three human-induced pluripotent stem cell (iPSC) lines from peripheral blood mononuclear cells (PBMCs) of LQT1 patients carrying pathogenic variants (c.569 G>A, c.585delG, and c.573_577delGCGCT) in KCNQ1. All lines show typical iPSC morphology, high expression of pluripotent markers, normal karyotype, and are able to differentiate into three germ layers in vitro. These lines are valuable resources for studying the pathological mechanisms of LQT1 caused by KCNQ1 mutations.

Cultivating Patient-Physician Communication About Vaccination Through Vaccine Metaphors.

The ease of access to misinformation online leaves patients vulnerable to poor decision-making and perplexed as to who serves as a reliable authority in the dissemination of health-related truths. Of prominent concern in twenty-first century medicine, is the communication between physicians and patients regarding vaccines. This cultural circumstance presents a challenge to physicians to be effective and trustworthy communicators, a challenge that entails the development of crucial skills at the earliest stages of medical education. We describe a pedagogical intervention through which medical students are given the educational experience of metaphorical construction to communicate the importance of vaccination.

Intrinsic properties and plasma membrane trafficking route of Src family kinase SH4 domains sensitive to retargeting by HIV-1 Nef.

The HIV type 1 pathogenicity factor Nef enhances viral replication by modulating multiple host cell pathways, including tuning the activation state of infected CD4 T lymphocytes to optimize virus spread. For this, Nef inhibits anterograde transport of the Src family kinase (SFK) Lck toward the plasma membrane (PM). This leads to retargeting of the kinase to the trans-Golgi network, whereas the intracellular transport of a related SFK, Fyn, is unaffected by Nef. The 18-amino acid Src homology 4 (SH4) domain membrane anchor of Lck is necessary and sufficient for Nef-mediated retargeting, but other details of this process are not known. The goal of this study was therefore to identify characteristics of SH4 domains responsive to Nef and the transport machinery used. Screening a panel of SFK SH4 domains revealed two groups that were sensitive or insensitive for trans-Golgi network retargeting by Nef as well as the importance of the amino acid at position 8 for determining Nef sensitivity. Anterograde transport of Nef-sensitive domains was characterized by slower delivery to the PM and initial targeting to Golgi membranes, where transport was arrested in the presence of Nef. For Nef-sensitive SH4 domains, ectopic expression of the lipoprotein binding chaperone Unc119a or the GTPase Arl3 or reduction of their endogenous expression phenocopied the effect of Nef. Together, these results suggest that, analogous to K-Ras, Nef-sensitive SH4 domains are transported to the PM by a cycle of solubilization and membrane insertion and that intrinsic properties define SH4 domains as cargo of this Nef-sensitive lipoprotein binding chaperone-GTPase transport cycle.

The Antagonism of HIV-1 Nef to SERINC5 Particle Infectivity Restriction Involves the Counteraction of Virion-Associated Pools of the Restriction Factor.

SERINC3 (serine incorporator 3) and SERINC5 are recently identified host cell inhibitors of HIV-1 particle infectivity that are counteracted by the viral pathogenesis factor Nef. Here we confirm that HIV-1 Nef, but not HIV-1 Vpu, antagonizes the particle infectivity restriction of SERINC5. SERINC5 antagonism occurred in parallel with other Nef activities, including cell surface receptor downregulation, trans-Golgi network targeting of Lck, and inhibition of host cell actin dynamics. Interaction motifs with host cell endocytic machinery and the Nef-associated kinase complex, as well as CD4 cytoplasmic tail/HIV-1 protease, were identified as essential Nef determinants for SERINC5 antagonism. Characterization of antagonism-deficient Nef mutants revealed that counteraction of SERINC5 occurs in the absence of retargeting of the restriction factor to intracellular compartments and reduction of SERINC5 cell surface density is insufficient for antagonism. Consistent with virion incorporation of SERINC5 being a prerequisite for its antiviral activity, the infectivity of HIV-1 particles produced in the absence of a SERINC5 antagonist decreased with increasing amounts of virion SERINC5. At low levels of SERINC5 expression, enhancement of virion infectivity by Nef was associated with reduced virion incorporation of SERINC5 and antagonism-defective Nef mutants failed to exclude SERINC5 from virions. However, at elevated levels of SERINC5 expression, Nef maintained infectious HIV particles, despite significant virion incorporation of the restriction factor. These results suggest that in addition to virion exclusion, Nef employs a cryptic mechanism to antagonize virion-associated SERINC5. The involvement of common determinants suggests that the antagonism of Nef to SERINC5 and the downregulation of cell surface CD4 by Nef involve related molecular mechanisms. IMPORTANCE: HIV-1 Nef critically determines virus spread and disease progression in infected individuals by incompletely defined mechanisms. SERINC3 and SERINC5 were recently identified as potent inhibitors of HIV particle infectivity whose antiviral activity is antagonized by HIV-1 Nef. To address the mechanism of SERINC5 antagonism, we identified four molecular determinants of Nef antagonism that are all linked to the mechanism by which Nef downregulates cell surface CD4. Functional characterization of these mutants revealed that endosomal targeting and cell surface downregulation of SERINC5 are dispensable and insufficient for antagonism, respectively. In contrast, virion exclusion and antagonism of SERINC5 were correlated; however, Nef was also able to enhance the infectivity of virions that incorporated robust levels of SERINC5. These results suggest that the antagonism of HIV-1 Nef to SERINC5 restriction of virion infectivity is mediated by a dual mechanism that is related to CD4 downregulation.

Differences in type I interferon signaling antagonism by dengue viruses in human and non-human primate cell lines.

BACKGROUND/OBJECTIVES: In vitro studies have shown that dengue virus (DENV) can thwart the actions of interferon (IFN)-α/ß and prevent the development of an antiviral state in infected cells. Clinical studies looking at gene expression in patients with severe dengue show a reduced expression of interferon stimulated genes compared to patients with dengue fever. Interestingly, there are conflicting reports as to the ability of DENV or other flaviviruses to inhibit IFN-α/ß signaling. METHODOLOGY/PRINCIPAL FINDINGS: In order to determine the relative inhibition of IFN-α/ß signaling by DENVs, a method combining flow cytometry and a four-parameter logistic regression model was established. A representative isolate from DENV-1, -3 and -4 and seventeen representative isolates encompassing all DENV-2 genotypes were evaluated. All of the DENVs evaluated in this study were capable of inhibiting IFN-α/ß signaling. Most of the strains were able to inhibit IFN-α/ß to a degree similar to DENV strain 16681; however, DENV-2 sylvatic strains demonstrated an increased inhibition of phosphorylated signal transducer and activator of transcription (pSTAT1). Surprisingly, we were unable to observe inhibition of pSTAT1 by DENV-2 sylvatic strains or the Asian strain 16681 in non-human primate (NHP) cell lines. Analysis in primary Rhesus macaque dendritic cells suggests that DENVs are capable of inhibiting IFN signaling in these cells. However, contrary to human dendritic cells, production of IFN-α was detected in the supernatant of DENV-infected Rhesus macaque dendritic cells. CONCLUSIONS: The ability of DENVs to inhibit IFN-α/ß signaling is conserved. Although some variation in the inhibition was observed, the moderate differences may be difficult to correlate with clinical outcomes. DENVs were unable to inhibit pSTAT1 in NHP cell lines, but their ability to inhibit pSTAT1 in primary Rhesus macaque dendritic cells suggests that this may be a cell specific phenomena or due to the transformed nature of the cell lines.

Differential cleavage of IRES trans-acting factors (ITAFs) in cells infected by human rhinovirus.

Human rhinovirus (HRV) is a major causative agent of the common cold, and thus has several important health implications. As a member of the picornavirus family, HRV has a small genomic RNA that utilizes several host cell proteins for RNA replication. Host proteins poly(rC) binding protein 2 (PCBP2) and polypyrimidine tract binding protein (PTB) are cleaved by a viral proteinase during the course of infection by the related picornavirus, poliovirus. The cleavage of PCBP2 and PTB inhibits poliovirus translation and has been proposed to mediate a switch in poliovirus template usage from translation to RNA replication. HRV RNA replication also requires a switch in template usage from translation to RNA replication; however, the mechanism is not yet known. We demonstrate that PCBP2 and PTB are differentially cleaved during HRV infection in different cell lines, suggesting that HRV utilizes a mechanism distinct from PCBP2 or PTB cleavage to mediate a switch in template usage.

Inhibition of poliovirus-induced cleavage of cellular protein PCBP2 reduces the levels of viral RNA replication.

UNLABELLED: Due to their small genome size, picornaviruses must utilize host proteins to mediate cap-independent translation and viral RNA replication. The host RNA-binding protein poly(rC) binding protein 2 (PCBP2) is involved in both processes in poliovirus infected cells. It has been shown that the viral proteinase 3CD cleaves PCBP2 and contributes to viral translation inhibition. However, cleaved PCBP2 remains active in viral RNA replication. This would suggest that both cleaved and intact forms of PCBP2 have a role in the viral RNA replication cycle. The picornavirus genome must act as a template for both translation and RNA replication. However, a template that is actively being translated cannot function as a template for RNA replication, suggesting that there is a switch in template usage from translation to RNA replication. We demonstrate that the cleavage of PCBP2 by the poliovirus 3CD proteinase is a necessary step for efficient viral RNA replication and, as such, may be important for mediating a switch in template usage from translation to RNA replication. IMPORTANCE: Poliovirus, like all positive-strand RNA viruses that replicate in the cytoplasm of eukaryotic cells, uses its genomic RNA as a template for both viral protein synthesis and RNA replication. Given that these processes cannot occur simultaneously on the same template, poliovirus has evolved a mechanism(s) to facilitate the switch from using templates for translation to using them for RNA synthesis. This study explores one possible scenario for how the virus alters the functions of a host cell RNA binding protein to mediate, in part, this important transition.

Viral proteinase requirements for the nucleocytoplasmic relocalization of cellular splicing factor SRp20 during picornavirus infections.

Infection of mammalian cells by picornaviruses results in the nucleocytoplasmic redistribution of certain host cell proteins. These viruses interfere with import-export pathways, allowing for the cytoplasmic accumulation of nuclear proteins that are then available to function in viral processes. We recently described the cytoplasmic relocalization of cellular splicing factor SRp20 during poliovirus infection. SRp20 is an important internal ribosome entry site (IRES) trans-acting factor (ITAF) for poliovirus IRES-mediated translation; however, it is not known whether other picornaviruses utilize SRp20 as an ITAF and direct its cytoplasmic relocalization. Also, the mechanism by which poliovirus directs the accumulation of SRp20 in the cytoplasm of the infected cell is currently unknown. Work described in this report demonstrated that infection by another picornavirus (coxsackievirus B3) causes SRp20 to relocalize from the nucleus to the cytoplasm of HeLa cells, similar to poliovirus infection; however, SRp20 is relocalized to a somewhat lesser extent in the cytoplasm of HeLa cells during infection by yet another picornavirus (human rhinovirus 16). We show that expression of poliovirus 2A proteinase is sufficient to cause the nucleocytoplasmic redistribution of SRp20. Following expression of poliovirus 2A proteinase in HeLa cells, we detect cleavage of specific nuclear pore proteins known to be cleaved during poliovirus infection. We also find that expression of human rhinovirus 16 2A proteinase alone can cause efficient cytoplasmic relocalization of SRp20, despite the lower levels of SRp20 relocalization observed during rhinovirus infection compared to poliovirus. Taken together, these results further define the mechanism of SRp20 cellular redistribution during picornavirus infections, and they provide additional insight into some of the differences observed between human rhinovirus and other enterovirus infections.

Picornavirus modification of a host mRNA decay protein.

UNLABELLED: Due to the limited coding capacity of picornavirus genomic RNAs, host RNA binding proteins play essential roles during viral translation and RNA replication. Here we describe experiments suggesting that AUF1, a host RNA binding protein involved in mRNA decay, plays a role in the infectious cycle of picornaviruses such as poliovirus and human rhinovirus. We observed cleavage of AUF1 during poliovirus or human rhinovirus infection, as well as interaction of this protein with the 5' noncoding regions of these viral genomes. Additionally, the picornavirus proteinase 3CD, encoded by poliovirus or human rhinovirus genomic RNAs, was shown to cleave all four isoforms of recombinant AUF1 at a specific N-terminal site in vitro. Finally, endogenous AUF1 was found to relocalize from the nucleus to the cytoplasm in poliovirus-infected HeLa cells to sites adjacent to (but distinct from) putative viral RNA replication complexes. IMPORTANCE: This study derives its significance from reporting how picornaviruses like poliovirus and human rhinovirus proteolytically cleave a key player (AUF1) in host mRNA decay pathways during viral infection. Beyond cleavage of AUF1 by the major viral proteinase encoded in picornavirus genomes, infection by poliovirus results in the relocalization of this host cell RNA binding protein from the nucleus to the cytoplasm. The alteration of both the physical state of AUF1 and its cellular location illuminates how small RNA viruses manipulate the activities of host cell RNA binding proteins to ensure a faithful intracellular replication cycle.

Viral subversion of host functions for picornavirus translation and RNA replication.

Picornavirus infections lead to symptoms that can have serious health and economic implications. The viruses in this family (Picornaviridae) have a small genomic RNA and must rely on host proteins for efficient viral gene expression and RNA replication. To ensure their effectiveness as pathogens, picornaviruses have evolved to utilize and/or alter host proteins for the benefit of the virus life cycle. This review discusses the host proteins that are subverted during infection to aid in virus replication. It will also describe proteins and functions that are altered during infection for the benefit of the virus.

Impairment of CD4+ T cell polarization by dengue virus-infected dendritic cells.

BACKGROUND: The production of type I interferon alpha/beta (IFN-α/ß) is crucial to viral clearance during dengue virus (DENV) infection; however, in vitro-infected dendritic cells (DCs) exhibit a decreased capacity to respond to IFN-α/ß stimulation, and antigen-presenting cells (APCs) isolated from patients with acute DENV infection exhibit defects in T cell priming. METHODS: In order to ascertain the stimulatory capacity of primary human monocyte-derived DCs infected with wild-type DENV isolates, representing a range of genotypes and disease outcomes, we cocultured infected DCs with allogeneic-naive CD4(+) T cells. The gene expression patterns of IFN-α/ß sensitive genes were quantitated to determine if the infected DCs displayed a blunted IFN-α/ß response. RESULTS: DENV-infected DCs induced the initial proliferation of naive CD4(+) T cells but they remained nonpolarized in effector function. The expression of IFN-α/ß-stimulated genes was downregulated, revealing that the inhibition of IFN-α/ß signaling is conserved among endemic DENV serotype 2 strains. CONCLUSIONS: The failure of naive CD4(+) T cells to differentiate into IFN gamma-producing effector T cells when primed by DENV-infected DCs cannot be explained solely by a block in IFN-α/ß signaling, suggesting that the ability of DENV to evade the early host response is multifaceted.

Preservation of FoxP3+ regulatory T cells in the peripheral blood of human immunodeficiency virus type 1-infected elite suppressors correlates with low CD4+ T-cell activation.

Elite suppressors (ES) are untreated human immunodeficiency virus type 1 (HIV-1)-infected individuals who maintain normal CD4(+) T-cell counts and control viremia to levels that are below the limit of detection of current assays. The mechanisms involved in long-term control of viremia have not been fully elucidated. CD4(+) CD25(+) regulatory T cells (Tregs) downmodulate chronic inflammation by suppressing the activation and proliferation of effector lymphocytes. We found that while Tregs were functional in ES and patients on highly active antiretroviral therapy (HAART), ES maintained high levels of Tregs in peripheral blood mononuclear cells whereas patients on HAART had evidence of Treg depletion. We also demonstrated that Tregs can serve as reservoirs for HIV-1 in vivo. These data suggest that both direct infection by HIV-1 and tissue redistribution are possible explanations for declining FoxP3(+) Tregs in progressive HIV-1 infection. Furthermore, the maintenance of Tregs may be one mechanism associated with the nonprogressive nature of HIV-1 infection in ES.

Severe depletion of CD4+ CD25+ regulatory T cells from the intestinal lamina propria but not peripheral blood or lymph nodes during acute simian immunodeficiency virus infection.

CD4+ CD25+ regulatory T cells (Tregs) suppress the activation and proliferation of effector lymphocytes. In human immunodeficiency virus type 1 (HIV-1) infection, Tregs play a significant role in controlling the apoptotic loss of uninfected CD4+ T cells resulting from high levels of generalized immune activation. During acute HIV-1 infection, more than 50% of CD4+ T cells are depleted from the gastrointestinal lamina propria. To elucidate the role of Tregs in HIV-1-induced depletion of CD4+ T cells in the gut-associated lymphoid tissue (GALT), we first determine the distribution of Tregs in a setting of acute infection using the simian immunodeficiency virus (SIV)/pigtailed macaque model of HIV-1 disease. CD4+ T cells from the GALT, lymph nodes, and peripheral blood were isolated from SIV-infected pigtailed macaques on days 4, 14, and 114 postinoculation. Quantitative real-time reverse transcription-PCR was used to quantitate FOXP3 copy numbers in SIV-infected and uninfected control macaques. Expression of FOXP3 in the ileal lamina propria was significantly decreased at all stages of infection compared to levels in uninfected control macaques. In addition, functional analysis of ileal CD4+ T cells from SIV-infected macaques revealed a lack of suppressive activity suggestive of the absence of Tregs in that compartment. These results indicate that Tregs are rapidly depleted in the GALT of SIV-infected macaques, defining a role for the loss of Treg-mediated suppression in early events in the pathogenesis of the disease.