Pediatric Residents Knowledge of Penicillin Allergy.

Penicillin allergy knowledge has not been evaluated specifically in the pediatric resident population. An anonymous electronic survey was distributed to all the pediatric residents in a single residency program to ascertain knowledge of penicillin allergies and allergy history taking skills. Responses among each resident class were compared using the Fisher exact test, 2-tailed. A total of 46 (52%) of 88 pediatric residents completed the survey. Only 63% reported to have had prior penicillin allergy education. All residents incorrectly identified low-risk symptoms as high-risk symptoms. The knowledge of penicillin allergy was poor across all training levels with no improvement over the duration of training. There is large support in the literature for de-labeling penicillin allergy in patients. Pediatric residents evaluate patients in childhood when most of the allergy labeling occurs. We need to consider strategies for incorporating penicillin allergy education in pediatric residency training.

A MTA2-SATB2 chromatin complex restrains colonic plasticity toward small intestine by retaining HNF4A at colonic chromatin.

Plasticity among cell lineages is a fundamental, but poorly understood, property of regenerative tissues. In the gut tube, the small intestine absorbs nutrients, whereas the colon absorbs electrolytes. In a striking display of inherent plasticity, adult colonic mucosa lacking the chromatin factor SATB2 is converted to small intestine. Using proteomics and CRISPR-Cas9 screening, we identify MTA2 as a crucial component of the molecular machinery that, together with SATB2, restrains colonic plasticity. MTA2 loss in the adult mouse colon activated lipid absorptive genes and functional lipid uptake. Mechanistically, MTA2 co-occupies DNA with HNF4A, an activating pan-intestinal transcription factor (TF), on colonic chromatin. MTA2 loss leads to HNF4A release from colonic chromatin, and accumulation on small intestinal chromatin. SATB2 similarly restrains colonic plasticity through an HNF4A-dependent mechanism. Our study provides a generalizable model of lineage plasticity in which broadly-expressed TFs are retained on tissue-specific enhancers to maintain cell identity and prevent activation of alternative lineages, and their release unleashes plasticity.

Epigenetic disruptions in the offspring hypothalamus in response to maternal infection.

DNA methylation is an epigenetic modification that can alter gene expression, and the incidence can vary across developmental stages, inflammatory conditions, and sexes. The effects of viral maternal viral infection and sex on the DNA methylation patterns were studied in the hypothalamus of a pig model of immune activation during development. DNA methylation at single-base resolution in regions of high CpG density was measured on 24 individual hypothalamus samples using reduced representation bisulfite sequencing. Differential over- and under-methylated sites were identified and annotated to proximal genes and corresponding biological processes. A total of 120 sites were differentially methylated (FDR-adjusted p-value < 0.05) between maternal infection or sex groups. Among the 66 sites differentially methylated between groups exposed to inflammatory signals and control, most sites were over-methylated in the challenged group and included sites in the promoter regions of genes SIRT3 and NRBP1. Among the 54 differentially methylated sites between females and males, most sites were over-methylated in females and included sites in the promoter region of genes TNC and EIF4G1. The analysis of the genes proximal to the differentially methylated sites suggested that biological processes potentially impacted include immune response, neuron migration and ensheathment, peptide signaling, adaptive thermogenesis, and tissue development. These results suggest that translational studies should consider that the prolonged effect of maternal infection during gestation may be enacted through epigenetic regulatory mechanisms that may differ between sexes.

Atacama Clear for Complex 3D Imaging of Organs.

3D reconstructive imaging is a powerful strategy to interrogate the global architecture of tissues. We developed Atacama Clear (ATC), a novel method that increases 3D imaging signal-to-noise ratios (SNRs) while simultaneously increasing the capacity of tissue to be cleared. ATC potentiated the clearing capacity of all tested chemical reagents currently used for optical clearing by an average of 68%, and more than doubled SNRs. This increased imaging efficacy enabled multiplex interrogation of tough fibrous tissue and specimens that naturally exhibit high levels of background noise, including the heart, kidney, and human biopsies. Indeed, ATC facilitated visualization of previously undocumented adjacent nephron segments that exhibit notoriously high autofluorescence, elements of the cardiac conduction system, and the distinct human glomerular tissue layers, at single cell resolution. Moreover, ATC was validated to be compatible with fluorescent reporter proteins in murine, zebrafish, and 3D stem cell model systems. These data establish ATC for 3D imaging studies of challenging tissue types.

SARS-CoV-2 infection causes dopaminergic neuron senescence.

COVID-19 patients commonly present with signs of central nervous system and/or peripheral nervous system dysfunction. Here, we show that midbrain dopamine (DA) neurons derived from human pluripotent stem cells (hPSCs) are selectively susceptible and permissive to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. SARS-CoV-2 infection of DA neurons triggers an inflammatory and cellular senescence response. High-throughput screening in hPSC-derived DA neurons identified several FDA-approved drugs that can rescue the cellular senescence phenotype by preventing SARS-CoV-2 infection. We also identified the inflammatory and cellular senescence signature and low levels of SARS-CoV-2 transcripts in human substantia nigra tissue of COVID-19 patients. Furthermore, we observed reduced numbers of neuromelanin+ and tyrosine-hydroxylase (TH)+ DA neurons and fibers in a cohort of severe COVID-19 patients. Our findings demonstrate that hPSC-derived DA neurons are susceptible to SARS-CoV-2, identify candidate neuroprotective drugs for COVID-19 patients, and suggest the need for careful, long-term monitoring of neurological problems in COVID-19 patients.

Isogenic human trophectoderm cells demonstrate the role of NDUFA4 and associated variants in ZIKV infection.

Population-based genome-wide association studies (GWAS) normally require a large sample size, which can be labor intensive and costly. Recently, we reported a human induced pluripotent stem cell (hiPSC) array-based GWAS method, identifying NDUFA4 as a host factor for Zika virus (ZIKV) infection. In this study, we extended our analysis to trophectoderm cells, which constitute one of the major routes of mother-to-fetus transmission of ZIKV during pregnancy. We differentiated hiPSCs from various donors into trophectoderm cells. We then infected cells carrying loss of function mutations in NDUFA4, harboring risk versus non-risk alleles of SNPs (rs917172 and rs12386620) or having deletions in the NDUFA4 cis-regulatory region with ZIKV. We found that loss/reduction of NDUFA4 suppressed ZIKV infection in trophectoderm cells. This study validated our published hiPSC array-based system as a useful platform for GWAS and confirmed the role of NDUFA4 as a susceptibility locus for ZIKV in disease-relevant trophectoderm cells.

Transanal irrigation in functional bowel disorders and LARS: short-term results from an Italian national study.

PURPOSE: To evaluate the potential benefits associated with the short-term (6 months) treatment with transanal irrigation (TAI) in patients suffering from functional constipation (FC), functional fecal incontinence (FI), and low anterior resection syndrome (LARS). METHODS: A multicenter observational study (12 centers; 369 patients) was conducted to assess the following primary and secondary objectives: to evaluate the level of satisfaction regarding bowel control and quality of life (QoL); to evaluate bowel symptoms severity and dropout frequency and reason. To this aim, validated questionnaires were provided to the patients at baseline (T0) and after 6 months of TAI treatment (T6) performed with the medical device Peristeen® Plus (Coloplast A/S, Denmark). Statistical analyses were conducted to compare the outcomes obtained at T0 and T6. RESULTS: A 6-month treatment with TAI enabled a statistically significant (p < 0.05) improvement of QoL scores, satisfaction scores regarding bowel control, and severity indexes of disorder-related symptoms in patients suffering from FC, FI, and LARS. Globally, 8.0% of patients discontinued the treatment after 6 months as a result of occurrence of symptoms (2.4%) or other justifications (3.8%) such as personal reasons. None of the dropouts were due to treatment inefficacy. CONCLUSION: Results of the present study suggest that short-term TAI treatment is beneficial for patients suffering from functional bowel disorders and LARS. Future analysis of prospective data will focus on the clinical outcomes associated with the long-term use (up to 24 months) of TAI when dealing with these types of medical conditions.

Human pluripotent-stem-cell-derived organoids for drug discovery and evaluation.

Human pluripotent stem cells (hPSCs) and three-dimensional organoids have ushered in a new era for disease modeling and drug discovery. Over the past decade, significant progress has been in deriving functional organoids from hPSCs, which have been applied to recapitulate disease phenotypes. In addition, these advancements have extended the application of hPSCs and organoids for drug screening and clinical-trial safety evaluations. This review provides an overview of the achievements and challenges in using hPSC-derived organoids to conduct relevant high-throughput, high-contentscreens and drug evaluation. These studies have greatly enhanced our knowledge and toolbox for precision medicine.

[Odontogenic infection as cause of orbital cellulitis]. / Een dentale oorzaak van orbitale cellulitis.

BACKGROUND: Orbital cellulitis is a potentially life-threatening condition. Compression of the optical nerve can cause total or partial loss of vision. Early diagnosis is crucial to prevent complications. In case of a unilateral sinusitis as cause of a unilateral orbital cellulitis complete clinical and dental examination combined with imaging are essential in diagnostics. CASE DESCRIPTION: A 53-year-old man presented with left eye movement impairment, intermittent diplopia and moderate swelling of the left lower eyelid. His diagnosis was post septal orbital cellulitis and despite administration of oral antibiotics no clinical improvement was observed. Orbital imaging by CT could not exclude a dental cause of his unilateral maxillary sinusitis. He was referred to the department of oral and maxillofacial surgery where clinical examination showed a dental cause. After removal of two decayed upper molars a complete recovery was accomplished. CONCLUSION: Odontogenic causes for unilateral orbital cellulitis should always be considered in diagnostics in adults. Clinical presentation and dental examination combined with adequate imaging can confirm the diagnosis.

Druggable targets and therapeutic development for COVID-19.

Coronavirus disease (COVID-19), which is caused by SARS-CoV-2, is the biggest challenge to the global public health and economy in recent years. Until now, only limited therapeutic regimens have been available for COVID-19 patients, sparking unprecedented efforts to study coronavirus biology. The genome of SARS-CoV-2 encodes 16 non-structural, four structural, and nine accessory proteins, which mediate the viral life cycle, including viral entry, RNA replication and transcription, virion assembly and release. These processes depend on the interactions between viral polypeptides and host proteins, both of which could be potential therapeutic targets for COVID-19. Here, we will discuss the potential medicinal value of essential proteins of SARS-CoV-2 and key host factors. We summarize the most updated therapeutic interventions for COVID-19 patients, including those approved clinically or in clinical trials.

A human iPSC-array-based GWAS identifies a virus susceptibility locus in the NDUFA4 gene and functional variants.

Population-based studies to identify disease-associated risk alleles typically require samples from a large number of individuals. Here, we report a human-induced pluripotent stem cell (hiPSC)-based screening strategy to link human genetics with viral infectivity. A genome-wide association study (GWAS) identified a cluster of single-nucleotide polymorphisms (SNPs) in a cis-regulatory region of the NDUFA4 gene, which was associated with susceptibility to Zika virus (ZIKV) infection. Loss of NDUFA4 led to decreased sensitivity to ZIKV, dengue virus, and SARS-CoV-2 infection. Isogenic hiPSC lines carrying non-risk alleles of SNPs or deletion of the cis-regulatory region lower sensitivity to viral infection. Mechanistic studies indicated that loss/reduction of NDUFA4 causes mitochondrial stress, which leads to the leakage of mtDNA and thereby upregulation of type I interferon signaling. This study provides proof-of-principle for the application of iPSC arrays in GWAS and identifies NDUFA4 as a previously unknown susceptibility locus for viral infection.

Expanding the precision oncology toolkit with micro-organospheres for early cancer diagnosis.

Using an automatic microfluidics droplet platform, Ding et al. successfully replicated the tumor micro-environment by generating micro-organospheres, which were then used to predict the response to anti-tumor drugs. These miniature models could be obtained within an extremely short time frame of 14 days, amplifying their role in facilitating cancer treatment decisions.

Evaluation of the Sensitivity of Metabolic Profiling by Rapid Evaporative Ionization Mass Spectrometry: Toward More Radical Oral Cavity Cancer Resections.

Radical resection for patients with oral cavity cancer remains challenging. Rapid evaporative ionization mass spectrometry (REIMS) of electrosurgical vapors has been reported for real-time classification of normal and tumor tissues for numerous surgical applications. However, the infiltrative pattern of invasion of oral squamous cell carcinomas (OSCC) challenges the ability of REIMS to detect low amounts of tumor cells. We evaluate REIMS sensitivity to determine the minimal amount of detected tumors cells during oral cavity cancer surgery. A total of 11 OSCC patients were included in this study. The tissue classification based on 185 REIMS ex vivo metabolic profiles from five patients was compared to histopathology classification using multivariate analysis and leave-one-patient-out cross-validation. Vapors were analyzed in vivo by REIMS during four glossectomies. Complementary desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) was employed to map tissue heterogeneity on six oral cavity sections to support REIMS findings. REIMS sensitivity was assessed with a new cell-based assay consisting of mixtures of cell lines (tumor, myoblasts, keratinocytes). Our results depict REIMS classified tumor and soft tissues with 96.8% accuracy. In vivo REIMS generated intense mass spectrometric signals. REIMS detected 10% of tumor cells mixed with 90% myoblasts with 83% sensitivity and 82% specificity. DESI-MSI underlined distinct metabolic profiles of nerve features and a metabolic shift phosphatidylethanolamine PE(O-16:1/18:2))/cholesterol sulfate common to both mucosal maturation and OSCC differentiation. In conclusion, the assessment of tissue heterogeneity with DESI-MSI and REIMS sensitivity with cell mixtures characterized sensitive metabolic profiles toward in vivo tissue recognition during oral cavity cancer surgeries.

Inflammatory responses in the placenta upon SARS-CoV-2 infection late in pregnancy.

The effect of SARS-CoV-2 infection on placental function is not well understood. Analysis of placentas from women who tested positive at delivery showed SARS-CoV-2 genomic and subgenomic RNA in 22 out of 52 placentas. Placentas from two mothers with symptomatic COVID-19 whose pregnancies resulted in adverse outcomes for the fetuses contained high levels of viral Alpha variant RNA. The RNA was localized to the trophoblasts that cover the fetal chorionic villi in direct contact with maternal blood. The intervillous spaces and villi were infiltrated with maternal macrophages and T cells. Transcriptome analysis showed an increased expression of chemokines and pathways associated with viral infection and inflammation. Infection of placental cultures with live SARS-CoV-2 and spike protein-pseudotyped lentivirus showed infection of syncytiotrophoblast and, in rare cases, endothelial cells mediated by ACE2 and Neuropilin-1. Viruses with Alpha, Beta, and Delta variant spikes infected the placental cultures at significantly greater levels.

Human organoid models to study SARS-CoV-2 infection.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is one of the deadliest pandemics in history. SARS-CoV-2 not only infects the respiratory tract, but also causes damage to many organs. Organoids, which can self-renew and recapitulate the various physiology of different organs, serve as powerful platforms to model COVID-19. In this Perspective, we overview the current effort to apply both human pluripotent stem cell-derived organoids and adult organoids to study SARS-CoV-2 tropism, host response and immune cell-mediated host damage, and perform drug discovery and vaccine development. We summarize the technologies used in organoid-based COVID-19 research, discuss the remaining challenges and provide future perspectives in the application of organoid models to study SARS-CoV-2 and future emerging viruses.

SARS-CoV-2 Infection Induces Ferroptosis of Sinoatrial Node Pacemaker Cells.

BACKGROUND: Increasing evidence suggests that cardiac arrhythmias are frequent clinical features of coronavirus disease 2019 (COVID-19). Sinus node damage may lead to bradycardia. However, it is challenging to explore human sinoatrial node (SAN) pathophysiology due to difficulty in isolating and culturing human SAN cells. Embryonic stem cells (ESCs) can be a source to derive human SAN-like pacemaker cells for disease modeling. METHODS: We used both a hamster model and human ESC (hESC)-derived SAN-like pacemaker cells to explore the impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on the pacemaker cells of the heart. In the hamster model, quantitative real-time polymerase chain reaction and immunostaining were used to detect viral RNA and protein, respectively. We then created a dual knock-in SHOX2:GFP;MYH6:mCherry hESC reporter line to establish a highly efficient strategy to derive functional human SAN-like pacemaker cells, which was further characterized by single-cell RNA sequencing. Following exposure to SARS-CoV-2, quantitative real-time polymerase chain reaction, immunostaining, and RNA sequencing were used to confirm infection and determine the host response of hESC-SAN-like pacemaker cells. Finally, a high content chemical screen was performed to identify drugs that can inhibit SARS-CoV-2 infection, and block SARS-CoV-2-induced ferroptosis. RESULTS: Viral RNA and spike protein were detected in SAN cells in the hearts of infected hamsters. We established an efficient strategy to derive from hESCs functional human SAN-like pacemaker cells, which express pacemaker markers and display SAN-like action potentials. Furthermore, SARS-CoV-2 infection causes dysfunction of human SAN-like pacemaker cells and induces ferroptosis. Two drug candidates, deferoxamine and imatinib, were identified from the high content screen, able to block SARS-CoV-2 infection and infection-associated ferroptosis. CONCLUSIONS: Using a hamster model, we showed that primary pacemaker cells in the heart can be infected by SARS-CoV-2. Infection of hESC-derived functional SAN-like pacemaker cells demonstrates ferroptosis as a potential mechanism for causing cardiac arrhythmias in patients with COVID-19. Finally, we identified candidate drugs that can protect the SAN cells from SARS-CoV-2 infection.

Increasing Prenatal Care Compliance in At-Risk Black Women: Findings from a RCT of Patient Navigation and Behavioral Incentives.

BACKGROUND: In the USA, infant mortality remains a major public health concern, particularly for Black women and their infants who continue to experience disproportionately high mortality rates. Prenatal care is a key determinant of infant health, with inadequate prenatal care increasing risk for prematurity, stillbirth, neonatal loss, and infant death. The aim of the present study was to determine if concurrent delivery of patient navigation and behavioral incentives to at-risk Black pregnant women could improve prenatal care attendance and associated maternal and infant outcomes. METHODS: Participants were 150 Black pregnant women recruited at first prenatal visit and screening at risk for adverse maternal and infant outcomes. Women were randomized to either the patient navigation + behavioral incentives intervention (PNBI) or assessment + standard care control (ASC) group. All were followed throughout pregnancy and 12-week postpartum. Group comparisons were made using intention-to-treat and per-protocol sensitivity analyses. RESULTS: While no group differences were found in prenatal care visits, the average number of visits for both groups (9.3 for PNBI and 8.9 for ASC) approached the American College of Obstetricians and Gynecologists (ACOG) recommended guidelines. There were also no group differences in maternal and infant outcomes. Both intention-to-treat and per-protocol sensitivity analyses, however, consistently found PNBI women attended more postpartum visits than ASC controls (p = 0.002). CONCLUSIONS: Given ACOG's redefining of the postpartum period as the fourth trimester, study findings suggest PNBI may facilitate prevention and intervention efforts to more successfully reduce health disparities in outcomes for both mother and infant.

Introductory patient communication training for medical physics graduate students: Pilot experience.

Despite medical physics becoming a more patient-facing part of the radiation oncology team, medical physics graduate students have no training in patient communication. An introductory patient communication training for medical physics graduate students is presented here. This training exposes participants to foundational concepts and effective communication skills through a lecture and it allows them to apply these concepts through realistic simulated patient interactions. The training was conducted virtually, and eight students participated. The impact of the training was evaluated based on changes in both confidence and competence of the participants' patient communication skills. Participants were asked to fill out a survey to assess their confidence on communicating with patients before and after the training. They also underwent a simulated patient interaction pre- and postlecture. Their performance during these was evaluated by both the simulated patient actors and the participants themselves using a rubric. Each data set was paired and analyzed for significance using a Wilcoxon rank-sum test with an alpha of 0.05. Participants reported significantly higher confidence in their feeling of preparedness to interact with patients (mean = 2.38 vs. 3.88, p = 0.008), comfort interacting independently (mean = 2.00 vs. 4.00, p = 0.002), comfort showing patients they are actively listening (mean = 3.50 vs. 4.50, p = 0.005), and confidence handling challenging patient interactions (mean = 1.88 vs. 3.38, p = 0.01), after the training. Their encounter scores, as evaluated by the simulated patient actors, significantly increased (mean = 77% vs. 91%, p = 0.022). Self-evaluation scores increased, but not significantly (mean = 62% vs. 68%, p = 0.184). The difference between the simulated patient and self-evaluation scores for the postinstruction encounter was statistically significant (p = 0.0014). This patient communication training for medical physics graduate students is effective at increasing both the confidence and the competence of the participants in the subject. We propose that similar trainings be incorporated into medical physics graduate training programs prior to students entering into residency.

SARS-CoV-2 infection induces beta cell transdifferentiation.

Recent clinical data have suggested a correlation between coronavirus disease 2019 (COVID-19) and diabetes. Here, we describe the detection of SARS-CoV-2 viral antigen in pancreatic beta cells in autopsy samples from individuals with COVID-19. Single-cell RNA sequencing and immunostaining from ex vivo infections confirmed that multiple types of pancreatic islet cells were susceptible to SARS-CoV-2, eliciting a cellular stress response and the induction of chemokines. Upon SARS-CoV-2 infection, beta cells showed a lower expression of insulin and a higher expression of alpha and acinar cell markers, including glucagon and trypsin1, respectively, suggesting cellular transdifferentiation. Trajectory analysis indicated that SARS-CoV-2 induced eIF2-pathway-mediated beta cell transdifferentiation, a phenotype that could be reversed with trans-integrated stress response inhibitor (trans-ISRIB). Altogether, this study demonstrates an example of SARS-CoV-2 infection causing cell fate change, which provides further insight into the pathomechanisms of COVID-19.